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1. sssssssse 60 test bimodal after Data 1736K 23 mm essent nenn anna 60 Figure 18d The intensity weighted Distribution Analysis result for the test bimodal 61 corresponding to Figure TREE 61 Figure 18e The number weighted Distribution Analysis result for the test bimodal 62 corresponding to Figures 196 0 ca nenne then e de 62 Figure 19a Printout of the intensity weighted Distribution Analysis result for the 3 1 64 91 26 TTA test bImodal n ae M AE 64 Nicomp 380 User Manual PSS 380Nicomp 030806 11 06 Page vi TT Yi iT Ti qm t TABLE OF CONTENTS m Figure for the 19b Printout of the volume weighted Distribution Analysis result 63 91 261nm test bimodal err ee ee een 65 Figure 19c Printout of the number weighted Distribution Analysis result for the 66 91 201 nm test DIMOU Al nenne nennen 66 Figure 20 Log C t B vs t for a widely separated bimodal latex sample 3 1 vol 68 ratio 91 and 1091 rimiz ie eden tas 68 Figure 21 The volume weighted Distribution Analysis result for the 91 1091 sample Figure 20 nm bimodal 69 Figure 22a Printout of volume weighted Distribution Analysis result for the 3 1 70 91 261 bimodal sample after 7 min sse 70 Figure 22b Printo
2. 11 decay Figure 8a Autocorrelation function for 91 nm latex standard ooooooocooocccccnnncccccconancnnncnnnnnnnnnnnnnos 16 Figure 8b Block of raw data corresponding to Figure 8a cccocccconnccoconcccnccnnnccnnnnnnnnnnnonononnnnnnnns 18 Figure 8c Log C t B vs t for data of Figures 8a and 8b uuuuueenssnnnessnnnnnnnnnnnnnnnnnnnnnnn 19 Figure 9a Autocorrelation function for an IV fat emulsion ooooconinononcccnnnnncncnnnnannnnnnnnnnnnnnnnnnns 22 Figure 9b Loge C t B vs t for data of Figure 9a 24 Figure lOa Intensity weighted Gaussian Analysis corresponding to the data of 27 Figure 9a and Dirse en ee M 27 Figure IOb Volume weighted Gaussian Analysis corresponding to Figure l0a 31 and data or Figures 98 andb see ae nad bee lehnten enden M Ea nenne ee 31 Figure lOc Number weighted Gaussian Analysis corresponding to Figure IOa 32 and data 0f Figure 9a and D a ee 32 Figure 11a Printout of volume weighted Gaussian Analysis result for fat emulsion 33 S66 Figure ID P 33 Figure 11b Printout of volume weighted Gaussian Analysis result for fat emulsion 34 See Fig re lOD repe m 34 Nicomp 380 User Manual PSS 380Nicomp 030806 11 06 Page v TT A ay Tm d m TABLE OF CONTENTS Figure 11c Printout of number weighted Gaussian Analysis result for 35
3. F3 F5 Increase sensitivity Used to raise the sensitivity to achieve the optimum Photopulse Rate Stop Auto Save Print Used to stop the automatic saving and printing of particle size results and allows for the clearing of the autocorrelator so that a new sample may be introduced into the instrument Help Menu This option can be accessed anywhere in the software to access help Auto Print Save Menu Please refer to the Particle Sizing section of this manual for additional information concerning this option Control Menu Please refer to the Particle Sizing section of this manual for additional information concerning this option Save Data File This option is used throughout the CW380 software whenever a data file is to be saved Please refer to the File section of this manual for step by step instruction for saving a file ALT F5 Save ASCII File F6 Use this option to save the data collected for a particular sample to an ASCII file format This data can then be imported to a spreadsheet program for presentation Read Data File A data file that has been stored following a measurement can be retrieved to display the resulting particle size distribution PSD with the desired weighting When this option is selected a list of data files will display in the Read Data File window Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 66 ME T T 1 NICOMP SOFTWARE AA F7 Read
4. B A A B A A A A A A A A A A C B B A B A B A B B B B D A A A A A 0 U0Ug U gt U gt U gt gt gt gt gt gt gt 00U0U0O gt UD gt gt gt Dp gt gt gt U0UOU gt P gt gt UUU 1 Di Di DD 1 SOI vie UJ P UPP PPP gt Duw WD gt U UU purga BY OU PPD PPPD B gt gt gt gt gt gt gt Head Material PC PPS A A A A A B A A z D A D A D B A A D A A A D A A A A A A A A A A A A A a A A A A A A A A A D A A A A A A OWWOAPOP gt OUPWWWWPU gt gt gt gt PrwWwWwuwW Pu gt uww um gt wu gt gt gt W Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page C 9 Fluid Sodium Sulfite Steam Stearic Acid Sulfuric Acid med Sulfuric Acid conc Sulfuric Acid dil Sulfurous Acid Tannic Acid Tanning Liquors Tartaric Acid Tin Salts Toluene Toluol Trichloroacetic Acid Trichloroethylene Trisoduim Phosphate Turpentine Urea Uric Acid Water Fresh Water Salt Xylene Zinc Chloride Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page C 10 APPENDIX C WU gt gt gt gt 0 gt U0UUW gt gt gt gt gt gt gt 0 gt 2 P gt PU gt gt gt P gt PU gt PU gt P gt PU gt D gt UD gt DPD gt gt gt DP gt 0 gt Q Tubing YOUOOODU gt gt O0 gt PU gt gt gt gt U gt U WU gt gt gt gt gt gt gt gt gt U gt WO gt 0 WO UOOU gt gt U gt gt gt gt gt gt UUU gt
5. Chi Squared 3 861 Norm Stnd Dev 0 180 Baseline Adj 0 000 Coeff of Var n Z Avg Diff Coeff 1 61E 008 cm2 s Run Time 0 Hr 6 Min 8 Sec W avelength 6328 nm Count Rate 0 KHz Temperature 23 deg C Channel 1 6126 K Viscosity 0 933 cp Channel Width 35 0 uSec Index of Ref 1333 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 6 APPENDIX Al NUMBER WEIGHTED NICOMP Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm 7 3 Data File C Program Files Particle Sizing Systems Nicomp 388 version 1 68 Bimodal 300 NUMBER Weighted NICOMP DISTRIBUTION Analysis Solid Particle NICOMP SUMMARY Peak 1 Mean Diam 230 1 nm S Dev 7 6 nm 3 29 Num 47 72 96 Peak 2 Mean Diam 345 3 nm S Dev 13 8 nm 4 01 Num 52 28 NUMBER WT NICOMP DISTRIBUTION 100 Diam nm gt Bimodal 300 Mean Diameter 293 0 nm Fit Error 7 349 Residual 0 000 NICOMP SCALE PARAMETERS Min Diam 100 nm Plot Size 50 Smoothing 2 Plot Range 10 GAUSSIAN SUMMARY Mean Diameter 263 8 nm Variance P 1 0 033 Stnd Deviation 47 6 nm 18 0 Chi Squared 3 861 Norm Stnd Dev 0 180 Baseline Adj 0 000 Coeff of Var n Z Avg Diff Coeff 1 61E 008 cm2 s Run Time 0 Hr 6 Min 8 Sec W avelength 632 8 nm Count Rate 0 KHz Temperature 23 degC Channel 1 6126 K Viscosity 0 933 cp Channel Width 35 0 uSec index of Ref 1 333 Nicomp 380 Ma
6. Figure 19a Printout of the intensity weighted Distribution Analysis result for the 3 1 91 261nm test bimodal Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 64 A A A A RESULTS Distribution Analysis Peak 1 Diameter 100 9 Intens 22 7 Peak 2 Diameter 283 1 Intens 77 3 Fit Error 1 1 gt Residual Mean Diameter 239 2 nm Standard Deviation 5 0 nm Baussian Analysis Intens Weighting Mean Diameter 198 1 nw Standard Deviation 9 3 nm 45 1 X Chi uared ey Baseline Adjust 0 00 X Mean Diffusion 2 34E 8 cm2 sec Data 1735 8 K DLS THEORY LAAL NICOMP Distribution Analysis Solid Particles 3 Bimodal latex samples 311 91 amp 261 rm nanometers log scale REL VOLUME RESULTS Distribution Analysis Peak 1 Diameter 94 2 Volume 69 9 Peak 2 Diameter 283 4 Volume 30 1 z PoSumn w RESP Fit Error 1 1 TOF ITO TO TO i f mn pes NO Hf 4 0 DO OL HO TOCA 00 To Residual Mean Diameter 150 3 nm Standard Deviation nm Baussian Analysis Volume Weighting Mean Diameter 39 0 nm Standard Deviation 62 7 nw Chi uared 23 Baseline Adjust 0 00 x Mean Diffusion 3 34E 8 cm2 sec Data 1735 8 K Scale Parameters MIN DIAM 10 PLOT SIZE 45 SMOOTHING 3 RANGE 100 RUN TIME O Hours 22 Mins 50 Secs AVG COUNT RATE 306 4 kHz CHANNEL WIDTH USEC
7. Figures 12a and b show the displays for the intensity and volume weighted particle size distributions obtained from the Gaussian Analysis after only 64K of accumulated Data equivalent to between one and two minutes of running By contrast Figures 12c and d show the corresponding results obtained with 455K in Channel 1 after approximately 10 minutes The latter clearly represent results having a much higher confidence level Finally there is a little recognized but significant attribute of the Gaussian Analysis that deserves comment here It often turns out that the computed particle size distribution for a broad unimodal population settles more quickly than for a narrow distribution In the latter case one frequently observes results for 91 nm latex after only 64K of Data Nicomp 380 Manual Gaussian Analysis Solid Particles Chi Squared 2 3 Baseline adjust 0 00 X Data 63 6 K Mean Diffusion 4 96E 8 cm2 sec 42 2220 4 gt Diameter log 10 23 INTENSITY MEAN DIAMETER STD DEVIATION WEIGHTING Figure 12a Intensity weighted Gaussian Analysis PSS 380Nicomp 030806 06 06 Page 2 40 1 1 1 1 1 1 D 3 1 1 1 t i 1 t 1 1 J 1 4 E 4 1 J t 4 1 DLS THEORY Gaussian Analysis Solid Particles Chi Squared 2 3 Baseline adjust 0 00 Data 63 6 K Mean Diffusion 5 70E 8 cm2 sec che AO a gt Diameter 109 0 100 CHI SQUARED LARGE
8. INT VOL Weighted GAU SSIAN DISTRIBUTION Analysis Solid Particle Fit Error 735 Residual 0 00 Chi Squared 3586 Baseline Adj 0 00 96 Run Time 0 Hr 6 Min 8 Sec Wavelength 6328 nm Count Rate 0 KHz Temperature 23 degC Channel 1 6126 K Viscosity 0 933 cp Channel Width 35 0 uSec Index of Ref 1 333 Intensity Weighting Mean Diameter 288 9 nm Stnd Deviation 52 1 nm 18 03 Cumulative Result 25 of distribution lt 251 6 nm 50 of distribution lt 284 1 nm 75 of distribution lt 320 9 nm 90 of distribution lt 358 0 nm 99 of distribution lt 432 2 nm 80 of distribution 330 7 nm Volume Weighting Mean Diameter 2955 nm Stnd Deviation 53 3 nm 18 0396 Cumulative Result 25 96 of distribution 257 3 nm 50 96 of distribution 290 6 nm 75 of distribution lt 328 2 nm 90 96 of distribution 366 1 nm 99 96 of distribution 442 0 nm 80 96 of distribution 338 2 nm Diam nm gt Nicomp 380 Manual PSS 380Nicomp 030806 06 06 PageA 4 APPENDIX VOLUME WEIGHTED NICOMP Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm 7 3 Data File C Program Files Particle Sizing Systems Nicomp 388 version 1 68 Bimodal 300 VOLUME Weighted NICOMP DISTRIBUTION Analysis Solid Particle NICOMP SUMMARY Peak 1 Mean Diam 230 5 nm S Dev 7 5 nm 3 26 Vol 38 52 Peak 2 Mean Diam 345 9 nm S Dev 13 4 nm 3 89 Vol 61 48 VOLUME WT NICO
9. NICOMP 380 DLS User Manual Particle Sizing Systems Inc Particle Sizing Systems makes every effort to ensure that this document is correct However due to Particle Sizing Systems policy of continual product development we are unable to guarantee the accuracy of this or any other document after the date of publication We therefore disclaim all liability for any changes errors or omissions after the date of publication No reproduction or transmission of any part of this publication is allowed without express written permission of Particle Sizing Systems Inc Tg nani DOCUMENT CHANGE HISTORY AM Date Description of Document Revision of Review New Release Number 11 07 06 New Document 01 Particle Sizing Systems Nicomp 380 User Manual PSS 380Nicomp 030806 11 06 TT T ay MEA TABLE OF CONTENTS m GENERAL INFORMATION Erie eec eu ae SECTION 1 REGISTRA TON Ser E c td E cma ad EE Lt E 1 TECHNICAL SUPPORT einen 1 SAFETY CONSIDERATIONS 3 5 tete nen nenne Dr us e c petU tU tuse RE tue sce 2 ue DAA A 3 DES THEORY ti ta SECTION 2 DYNAMIC LIGHT SCATTERING THEORY uuuuuzssansnsnnnannnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn 1 PRINCIPLES OF DLS A QUALITATIVE REVIEW nter iret terea rca tad eR nip ida 1 Dynamic scattering the effects of diffusion nennen 3 Obtaining particle size from the diffusion coefficient esseeeeee 7 Autocorrelation func
10. PSS 380Nicomp 030806 06 06 Page C 11 RE v APPENDIX C In Appendix D a list of some of the more commonly used organic solvents with their viscosities at one or more temperatures as well as their approximate indices of refraction again whose T dependence can be ignored has been provided To operate at some different unlisted temperature the correct viscosity value can be estimated by linear interpolation or extrapolation The values in Appendix D were obtained from the following references 1 R C Weast Ed CRC Handbook of Chemistry and Physics CRC Press Inc Boca Raton FL e g 1982 63rd Ed 2 J Timmermans Physico Chemical Constants of Pure Organic Compounds Vols and Il Elsevier New York 1950 Important Exercise adequate CAUTION whenever using organic solvents In particular be aware of the fact that many solvents are highly FLAMMABLE and constitute a FIRE HAZARD They are often extremely volatile hence all other considerations being equal measurements should generally be carried out at REDUCED TEMPERATURES e g 15 or 20 C Also take necessary PRECAUTIONS to ensure adequate VENTILATION exists in the area where such solvents are used in most cases the FUMES can be assumed to be TOXIC dangerous to the eyes skin and internal organs Also spillage of such organic fluids may remove the paint on the instrument cabinet and otherwise cause damage within the Nicomp Nicomp 380 Manual PSS 380Nicom
11. corresponding to a particular diffusivity D and hence of particle radius R The challenge which we face is to develop fast and efficient mathematical methods of analysis whereby we can deconvolve C t and thereby extract the distribution of D values and hence of particle diameters from the detailed shape of C t The magic behind the DLS Module has to do with its ability to obtain accurately and consistently the most useful information relating to the distribution of particle sizes in solution To do this the 380 must analyze precisely the deviations of autocorrelation function C t from single exponential behavior As we shall discover below these deviations are often surprisingly slight and subtle given the large range of complicated distributions which are encountered The simplest kind of complexity in the particle size distribution that we can introduce is a smooth gaussian like population of sizes having a well defined mean diameter and half width Such an idealized distribution shape is often obtained for emulsions prepared by a variety of processes sonication homogenization and Microfluidization Typically some type of oil and water are caused to be mixed together with the aid of a dispersing agent e g a non ionic surfactant to form a single microscopically homogeneous phase The result tiny droplets of one component e g the oil suspended in the other component or phase e g water The mean
12. 0 WERFRBERPEEFFRPEFPFFESUERERGEFEBERF FE FEFRCRECFEPELFETEFFETEUPLERTEFRELRFREFEFPETFRRIFELERTERFE EPREFPERRFHER ENTER 57 Show Distributions u un en De eco eee ee ete RR HR een ale 57 Time PlotScalo TD C 2 58 WEIGHTING iaa Ren AA irte 59 O O 59 MS es 59 Number to a eT ler s E Me tee Cr wp Dco CU aks 59 azione E 59 Nicomp 380 User Manual PSS 380Nicomp 030806 11 06 Page ii TT Yi iT Ti qm it TABLE OF CONTENTS m PU zl T AIL T UI ile did 60 aie mer EE 60 MSU TAS iscsi Ah TEE ETT NN 60 ADOUECW O86 una A melee eine inane emul 60 COMMAND KEYS sehe dee 61 SAMPLE ANALYSIS RUN a NERERAR EUN CAR VEK GERM SECTION 6 NATERIAES poc T 1 aUi elo IMO je T RE E 1 bleed uel 1 EE ucc EET 2 Procedure A todilution oce cora dert te eee aan Caden ER e ge tne 2 Bree eeu Ern 5 Review of Completed Sample Results Print Sample Results Post Measurement System Flush SAMPLE MAINTENANCE 2 acer sun SECTION 7 MAINTENANCE tacon ne era tede dran A TR t fase de asco xa fers ases ie en debat ders 1 APPENDIX A VOLUME WEIGHTED GAUSSIAN en elek 1 NUMBER WEIGHTED GAUSSIAN snnm ath autor d deg aud UR qa a Rn eod tfe ino Deeds 3 INT VOLUME WEIGHTED GRAUSSIAN tra alarde 4 VOLUME WEIGHTED NICOMP 4 tdt rd 5 INTENSITY WEIGHTED NIGOM Pensionados 6 NUMBER WEIGHTED NICOMP asus ee a 7 NTVOL WEIGHT ED uba rele 8 SUMMAR Y RESUE
13. 16 TE Ti qp at NICOMP SOFTWARE m i Resume Taking Data M pee The autocorrelator resumes collecting data for the sample that has been introduced into the instrument Neutral Density Adjustments Es Used to increase the intensity of the incident laser light by rotating the Neutral Density Filter E Used to decrease the intensity of the incident laser light by rotating the Neutral Density Filter amp Will show a small dialogue box that will prompt the user for a Neutral Density Filter setting e Allows the Neutral Density Filter to automatically search out the optimal scattering intensity for the sample Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 17 NICOMP SOFTWARE Status Bar For Help press F1 Colecling Data The status bar provides pertinent information while running the Nicomp 380 Clock 10 59 52 AM Displays the real time clock that is set up in the windows operating system of the computer This is the clock that is used to date and time stamp the data files that are saved Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 18 NICOMP SOFTWARE SETUP The Setup menu allows for communication to be established between the Nicomp and the computer controller System Setup E CFG File PASOFTWARE PSS 388 version 1 58 cw388 cfg Select Serial Port Multi hngle Option Serial Port 1 Fixed Angle 90 Deg C Serial Port 2 Multi Angle Square Cell C
14. Ch 1 Dats x 000 154 Count Rate X1000 290 uSen gt Channel Width 10 0 uScc INTENSITY WEIGHTED GAUSSIAN SUMMARY Mean Diam nm Coelf of Var n Chi Sq 2 Decays Auto Baseline Adj 87 152 0 049 2 076 2 178 0 552 Figure 8a Autocorrelation function for 91 nm latex standard This particular sample has been chosen for this example because of its high uniformity of particle size i e it is nearly monodisperse Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 16 DLS THEORY Figure 8b shows a printout of the block of raw autocorrelation channel data corresponding to Figure 8a Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 17 RE hi DLS THEORY Figure 8b Block of raw data corresponding to Figure 8a The channel width At for this particular run was 10 usec microseconds It is instructive to verify that this autocorrelation function C t closely approximates the ideal result of a single decaying exponential function To do this we plot C t B B baseline versus t i e channel number on semi logarithmic graph paper A perfectly straight line of negative slope should result according to our previous discussion Equation 7 This is shown in Figure 8c The solid straight line has been drawn by eye to best approximate the slope established by the data points log C t B We have deliberately displaced the line below the points to permit the latter
15. Chi Squared cannot be used to judge the stability and or quality of the fit results if too little data has been acquired Here just 31 seconds into the run the very low value of Chi Squared is potentially misleading in suggesting that the Gaussian Analysis has already produced final results of high quality with settled values of the Mean Diameter and Standard Deviation In fact Chi Squared later increases to a high of 2 8 at the later time of 8 min 4 see when substantial amounts of additional Data have been incorporated into the autocorrelation function 564K in Channel 1 However what is meaningful is the fact that this rise is obviously spurious since it is followed by consistently lower values Chi Squared falls back essentially to unity 1 1 after 13 14 minutes into the run Clearly what matters in establishing the validity of the Gaussian Analysis result for this sample is the fact that Chi Squared remains low with increasing data acquisition showing no tendency to grow with time Second we verify from Table 1 that the intensity weighted Mean Diameter colt 3 settles very quickly to a reliable value After just a couple of minutes all succeeding values are within 1 of the settled value of approximately 226nm On the other hand the exhibits considerably initial few minutes of difference is the fact the settled value of approx 226 nm volume weighted Mean Diameter col 5 more variation up to 4 during the data acqu
16. Hydrocyanic Acid gas 10 Hydrofluoric Acid 50 Hydrofluoric Acid 75 Hydrogen Peroxide dil Hydrogen Peroxide 90 Hypochlorous Acid lodine Solutions Idoform Kerosene Ketones Lacquers Solvents Lactic Acids Lead Acetate Linseed Oil Lithium Hydroxide Manganese Salts Magnesium Chloride Magnesium Sulfate Malic Acid Mercury Salts Methane Methanol Methyl Alcohol Methyl Chloride Methyl Ethyl Ketone Mixed Acid Molydenium disulfide Monoethanolamine Naptha Natural Gas Nickel Salts Nitric Acid dil Nitric Acid med conc Nitric Acid concentrated Nitrobenzene Nitrogen Oxides Nitrous Acid gt U0BPB SPDs O PPPU gt gt UU 0000 PP gt gt gt gt PPP gt gt gt gt UU O gt U gt gt gt gt gt gt 000020 gt gt PPU gt Pwu gt gt u gt gt gt gt H4 OPOU O0Uv dg OUO gt UUOOOUUUU P PU gt PPP D gt O gt PUUP U PDDD I DD gt gt PU gt P gt gt PP gt PUUU PD P gt gt DUr gt rwo U gt U UY gt gt gt UOT OPUS PPDP tgi i WU gt 2 gt 2 gt 2 gt PU gt 2U0DOW gt 0W gt gt gt gt 0 0000 WOUUOOU WU0UO gt gt gt gt gt gt gt gt u gt uDvuo UDUUO gt gt gt gt gt gt gt gt gt gt UDUDu DUD gt gt gt 1 OW DD gt 2 12 gt 00 gt 00 O U00 gt gt U gt 0U00U0 gt U0U0U PDI OOOPS gt gt WP gt gt u DD gt P gt OO0 gt gt gt UDU U0U gt gt gt
17. It is convenient to absorb all of the components above into a new constant C5 given by C 4 3 M Mw CR E 5 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page E 1 T mir rm t m y APPENDIX E The expression for radius R can now be solved R 1 2 Mw E 6 By inserting this expression in for R using equation D 3 it is possible to obtain a relationship which connects D and My D e cys Macs or D C4My E 7 where single constant C4 and C and C2 C3 C4 C2 Equation E 7 has exactly the form which was claimed earlier in Equation E 1 where C3 and R 1 3 The prefactor depends on the temperature T and solvent viscosity n through constant C4 as well as the mass density of the particles through constant C2 Exponent amp depends on the relationship between the hydrodynamic radius R in Equation E 2 and the molecular weight for simple spheres R equals 1 3 Exponent 3 has been computed theoretically for other particle shapes for example R 1 2 for random coil molecules and f 1 for rigid coil molecules From Equation E 1 a simple general expression for the molecular weight My as a function of the diffusivity D Mw D E 8 In the Nicomp Mw is computed from Equation E 8 using the mean diffusivity D intensity weighted obtained from the simple Gaussian Analysis Contents and 3 are obtained for the Input Menu In order to obtain reliable estimates of the mean molecular
18. Press F2 or choose Auto Print Save Menu from the Particle Sizing pull down menu NOTE the menu file location may be different for each computer and is unimportant for this exercise 3 Press F3 or choose Control Menu from the Particle Sizing pull down menu 4 Ensure that parameters are set according to the below Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 6 5 C380 Control Menu ES Menu File CAPSS Software cw388 version 1 68Cw388 tbl Channel Width 1 0 uS ec Autodilution Drop In Drop In Cell 23 veo C C Flow Cell Liquid Viscosity 0 333 CP Iv Autoset Channel Width Liquid Refractive Index 1 333 V Autoset Sensitivity Intensity Setpoint 300 KHz M Auto NICOMP Parameter First Channel Used 2 v Auto Baseline Adj Laser Wavelength nm Cum Set Pt ETS External Fiber Angle l E Y En DE Scattering Angle deg coo NOTE 5 6 10 11 12 13 14 The laser wavelength may differ depending on the instruments hardware setup Click on OK to return to the CW388 Software Window Prepare the sample by gently inverting the bottle of latex material ten times Add one drop of the concentrated latex standard to 25 mL of distilled filtered water The resulting mixture should be manually agitated to provide a uniform suspension Manually agitate sample to provide a uniform suspension This is confirmed by visual Inspection of the sterile beaker The suspension should be turbid slightly
19. Q RESULTS Distribution Analysis Peak Diameter Volume 16 4 54 6 89 1 1 100 0 63 5 29 1 Peak 42 Diameter Volume dh Bet geo grt pee e I I OL CD Fit Error 1 0 mu SE Residual 0 0 Mean Diameter 144 9 nm G3 QJ GIJTO TO TU TO EI ES DS en ooodOu Standard Deviation 85 5 rim A E 2010203 Gaussian Analysis Volume Weighting Mean Diameter 123 8 nm Standard Deviation 58 7 nm 47 4 Chi uared any Baseline Adjust 0 00 Data 2298 6 K ame mam nm mm Scale Parameters MIN DIAM 10 PLOT SIZE 60 SMOOTHING 3 RANGE 100 w Hours 41 Mins 42 Secs kHz 280 USEC DEGREES CENT CENTIPOISE K COUNTS INDEX PRINT AT DATA OF PRINTOUTS Figure 22c Printout of volume weighted Distribution Analysis result for the 3 1 91 261 bimodal sample after 42 min Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 72 DLS THEORY NICOMP Distribution Analysis Solid Particles SIZE nanometers log scale REL VOLUME 85 0 RESULTS 76 Distribution Analysis Peak 1 Diameter 89 5 Volume 76 3 Peak 2 Diameter 271 5 Volume 23 7 Fit Error Residual 0 0 Mean Diameter 140 2 nm Standard Deviation B4 2 nm Gaussian Analysis Volume Weighting Mean Diameter 124 1 run Standard Deviation 58 6 nm 47 3 X Ch uared
20. Serial Port 3 C Multi Angle Round Cell C Serial Port 4 C Multi Angle Model 170 Interrupter Angle 13 5 deg Flow Pump Change Laser Wavelength nm APD Overload Protection Intensity Overshot Factor 12 NICOMP Intens wt Threshold EA Enable Intensity Monitor DualPartilce Sizing DLS Detector O Cancel C Select Serial Port Four serial ports are provided for setting up communications between the Nicomp and the computer Position the cursor over the desired selection and click on the corresponding circle A black circle will display next to the selection and the parameter will appear in the System Setup menu Multi Angle Option This parameter is used to establish the configuration for the detection of scattered light There are four possible configurations to choose from Fixed Angle 90 Deg Selected when using the basic Nicomp in which the scattering angle is set to 90 degrees Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 19 T ri Ts roman AM N NICOMP SOFTWARE Multi Angle Square Cell Selected ifthe Nicomp possesses the multi angle option computer controlled stepper motor 0 9 deg step optical fiber and a square cuvet either normal 1 cm or miniature is used for the sample cell Multi Angle Round Cell Selected when the Nicomp possesses the multi angle option and a cylindrical sample cell Cylindrical Cells The true scattering angle is equal to the external angle of th
21. TEMPERATURE 23 DEGREES CENT VISCOSITY CENTIPOISE INDEX OF REFRRCTION co kHz Wt E 4 BETA Molec Wt Figure for the 19b Printout of the volume weighted Distribution Analysis result 91 261nm test bimodal Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 65 DLS THEORY NICOMP Distribution Analysis Solid Particles 3 Bimodal latex samples 3 1 91 4 261 nm SIZE nanometers log scale REL NUMBER 3 8 RESULTS 65 7 Distribution Analysis Peak 1 Diameter 90 0 Number 98 5 Peak 42 Diameter 265 7 Number 1 5 FAS Error Residual Mean Diameter rm Standard Deviation 25 4 nm Gaussian Analysis Number Weight ing Mean Diameter 61 6 nm Standard Deviation 7 7 nm 45 1 X Chi Squared 24 7 Baseline Adjust 0 00 X Mean Diffusion 7 54E 8 cm2 sec Data 1735 8 K Scale Parameters MIN DIAM 10 PLOT SIZE 45 SMOOTHING 3 RANGE 100 rs 22 Mins 50 Secs Hz yo DEGREES CENT CENTIPOISE kHz E 4 CHANNEL WID E BETA Molec Wt Figure 19c Printout of the number weighted Distribution Analysis result for the 91 261 nm test bimodal Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 66 rity Til iT jT DLS THEORY AAL As mentioned above we can generally conclude that the wider the separation of a bimodal the easier it is for the Distribution Analysis to perform an accurate measurement again provided there is adequate scattering from the weakest of t
22. gt UUUUOO gt gt gt Uu gt gt gt gt uw gt gt UUUD gt gt gt U gt UU gt Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page C 6 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page C 7 Fluid Oils Animal Oils Minerals Oils Vegetable Oleic acid Oxalic Acid cold Oxygen gas Palmitic Acid Perchloric Acid Perchloroethylene Phenol Carbolic Acid Phosphoric Acid gt 50 Phthalic Acid Plating Solutions Polyglycol Potassium Carbonate Potassium Chlorate Potassium Hydroxide caus Potassium Hydroxide med Potassium lodide Propyl Alcohol Pyridine Silicone Oils Silicone Fluids Silver Nitrate Soap Solutions Sodium Bicarbonate Sodium Bisulfate Sodium Bisulfite Sodium Borate Sodium Carbonate Sodium Chlorate Sodium Chloride Sodium Ferrocyanide Sodium Hydrosulfite Sodium Hydroxide dil Sodium Hydroxide 20 Sodium Hydroxide conc Sodium Hypochlorite lt 5 Soduim Hypochlorite gt 5 Sodium Nitrate Sodium Silicate Sodium Sulfide Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page C 8 APPENDIX C i OU o gt u 2 gt Pu gt gt gt gt gt gt gt gt gt gt gt gt oou gt UUU gt gt gt gt gt U0U PPDDDDO P gt r gt gt gt gt gt gt gt gt r gt gt gt gt gt gt gt D gt gt umwUo gt Uu gt gt gt gt 00Q Tubing S T D D D A B D D B A B A D D D D B A B A D A A A A B A B B A A D D D B
23. gt gt gt gt UUWUW gt U gt p P O V i B gt gt OoO gt gt u gt gt v o EL 2 25 5UUO02 UO gt O gt JU gt gt OO Head Material PC PPS D A A A E C A C A A A A A B A D A D A D A A A D A D A A A A A D A A A O 0 UO0 0 U0umocooog 60 LE MEER APPENDIX N t A safe and desirable alternative to using the Autodilution system with a nonaqueous solvent is to convert the Nicomp to disposable cell operation This is easily carried out by removing the flow through cell from the cell holder by unscrewing the four thumb screws moving it to one side away from the scattered light detection path reassembling the cell holder and dropping into the latter the black anodized adapter for the disposable 6 mm cylindrical glass tubes or 1 cm glass cuvette Following this procedure the Nicomp is now compatible with virtually any solvent that is compatible with glass A second requirement when using a solvent other than water concerns the accuracy of the measurement The same principles of light scattering diffusion and autocorrelation hold true regardless of the solvent used However substitution of new values into the Input menu must be made for the physical parameters of the solvent viscosity n and index of refraction n In general the former is highly temperature dependent so that the value entered must correspond to the running temperature chosen Often the reverse i
24. indicating that almost no adjustment in the value of B was needed to obtain the lowest value of Chi Squared Hence this particular sample contained a negligible concentration of large particle aggregates or other contaminants The value for Data 1047 K represents the contents of channel 1 of the autocorrelation function C t Such a relatively large value exceeding one million 1000 K usually indicates a high degree of statistical accuracy in C t At this point the results of the Gaussian Analysis should have become stable with relatively little change with additional run time In the example summarized above 1047 K was achieved in channel 1 after a run time of 14 mm 48 sec The relationship between the value of Data and the run time will in general depend on the average Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 28 scattered light intensity and the channel width At DLS THEORY Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 29 T mir AM DLS THEORY Effects of weighting in the Gaussian Analysis The problem with an intensity weighted particle size distribution is that it is generally not very useful in the real world Typically one wishes to obtain a size distribution that is either weighted by particle number a trivial weighting or by particle volume or mass i e weighted by diameter cubed The DLS Module uses the rules of light scattering including corrections
25. possibly a baseline adjust Hence we should NEVER be lulled into believing the actual heights of each of the slices that comprise the distribution curve These values have originated from an idealized mathematical table notfrom a measurement procedure which counts particles one at a time Because the DLS technology is unable to measure particles individually it yields only smoothed idealized estimates of the actual particle size distribution curves Nevertheless the DLS Module will usually be found to be very sensitive to small changes in the shape of the underlying particle size distribution Importance of acquiring data of sufficient accuracy A central repeated theme of this section will be the importance of acquiring autocorrelation data of high statistical accuracy If this is achieved the resulting computed particle size distributions will be found to be stable in time As we shall see the level of statistical accuracy required for a reliable analysis depends greatly on the analysis method chosen Because the Gaussian Analysis is only a 2 parameter fit with the possible addition of a baseline adjustment it will settle to a reliable result relatively quickly certainly faster than the NJCOMP Distribution Analysis discussed in the next section This simply reflects the fact that the best quadratic fit to the reduced autocorrelation data is relatively insensitive to small variations in individual data points given a total of 64 poin
26. s DK with K given by Equation 9 Using these new definitions we can recast Equation 14 in its integral form in which there are an infinite number of different particle diameters Equation 14 therefore becomes H t ro exp st ds 15 where s or D ranges from zero corresponding to an arbitrarily large particle to infinity corresponding to a particle of diminishing small diameter The above expression defines the integral Laplace transform of H t which is the desired weighting function f s Returning to Equation 14 we must use an ILT technique to obtain the best possible estimates for the weighting coefficients fi The details of the mathematical procedure used to perform the discrete Laplace transform inversion of the autocorrelation data in the NICOMP Distribution Analysis are beyond the scope of this manual and in any case are proprietary Suffice to say that the procedure represents a variation on other ILT methods discussed in the light scattering literature most especially that developed by S Provencher The ways in which the NICOMP procedure differs from this and other methods relate mainly to the manner and extent of smoothing carried out within the mathematical algorithm Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 46 ME DLS THEORY m l It is most important to appreciate the practical limitations inherent in any solution of Equation 14 using an ILT algorithm In principle one would
27. 380 Manual U gt U gt gt gt U0O22200 gt 900 0OPU gt P gt gt gt gt gt gt 0n gt gt gt gt 0n gt U gt gt gt gt gt 2 O n QUO UOUUUUUUUUUUUUUO UOUUUUUUUUULDSUUUUU Tubing S T B D D B A B B D D D D A A C B A A A A B A C A C A A B C A A A A A D D D B D B B D D D D A A A D D B B B A A D A D B B B D D D D A A B A D B A D D OOUUO 0 U 0 00900990g U gt UO gt gt WVUDuUU gt gt U UDU gt gt uwD D O lt O PP PUU DD DUO Orrrrrroy gt UU gt gt UO0D PDI gt DUP gt PU gt gt gt gt gt gt U O OD gt T Head Material PC PPS SS A A A A A A C C A B D A A D A A A A A B A A A A A B D A B A A B A C D A A A A A A B A A B D A A A A D A A D D A D D D D D A A B C A B D A B A A B D D D A A C A A D A B A B A B A A A A D A B PSS 380Nicomp 030806 06 06 Page C 3 APPENDIX C Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page C 4 Fluid Tubing Head Material PC PPS S v gt DPP OPDP P gt gt gt gt gt gt 00 U U0 gt PU U U gt U 9 0 500090 o Chlorine dry Chlorine wet Chloroacetic Acid Chlorobenzene Chlorobromomethane Chloroform Chlorosulfonic Acid Chromic Acid 30 Chromium Salts Copper Salts Cresol Cyclohexane Cyclohexaone Diacetone A
28. 40 8 Baseline Adjust 0 00 X 30608 4 K Scale Parameters MIN DIAM 10 PLOT SIZE 60 SMOOTHING 3 RANGE 100 RUN TIME 8 Hours 9 Mins 37 Secs AVG COUNT RATE 297 5 kHz 19 0 23 DEGREES CENT 0 9325 CENTIPOISE 1 333 1000 K COUNTS 10 CHANNEL WIDTH a OF PRINTOUTS Figure 22d Printout of volume weighted Distribution Analysis result for the 3 1 91 261 bimodal sample after 8 hrs 10 min Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 73 MM hi DLS THEORY What is the lesson behind these printouts should samples be run for 8 hours Certainly not Rather the point is that if one wishes to optimize the analytical capabilities of the instrument they should make it a habit to obtain several intermediate printouts within the first 20 or 30 minutes of a run to verify the settling and stability of the computed distribution Only in this way will confidence be instilled in the user that the Distribution Analysis results are close to the best that can be obtained by the DLS Module given practical constraints on the total amount of the allotted time Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 74 INITIAL HARDWARE SETUP INITIAL HARDWARE SETUP 1 Connect the 25 pin male connector of the cable provided to the port identified on the back of the Nicomp instrument 2 Connect the 9 pin female end of the connector to a serial port on the PC controller Nico
29. 80 Chi Squared 0 56 60 t F Auto B Adj i 0 08 40 Ch 1 Data i x1000 20 i i 1854 Mean 0 i 1 i i i 1 Diff Coeff 20 cm2 sec Diam nm gt 1 7e 008 PQ260 300 Mean Diam nm X Coeff of Var n Stnd Dev nm 261 4 0 059 15 420 It will often be the case that the three weightings do not match in magnitude of mean diameter This is the representative of applying the different weightings to the original data set Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 48 Intensity and Volume Weighted Gaussian 80 60 40 20 0 20 Diam nm gt Intensity Weighting Mean Diameter 263 0 nm Stnd Deviation 15 5 nm 5 9 25 96 of distribution lt 235 1 nm 50 of distribution lt 244 6 nm 75 of distribution lt 254 6 nm 90 of distribution lt 260 7 nm 99 of distribution lt 294 0 nm REL INT WT GAUSSIAN DISTRIBUTION Solid Particle Cumulative Result Int We Vol Wt lt 235 1 nm lt 244 6 nm lt 254 6 nm lt 260 7 nm lt 294 0 nm NICOMP SOFTWARE REL VOL WT GAUSSIAN DISTRIBUTION Diam nm gt Solid Particle Volume Weighting Mean Diameter 263 1 nm Stnd Deviation 15 5 nm 5 9 Coeff of Yar n 0 059 ChiSq 0 56 Baseline Adj 0 08 Run Time 0 10 3 It is often most useful to observe both the intensity and volume weightings on the screen simultaneously Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 49 NICOMP
30. B vs t for data of Figures 8a and 8b Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 19 T mir AM DLS THEORY Using Equations 7 9 we can now calculate the particle diameter predicted by the semilog behavior shown in Figure 8c From Equation 7 we have log C t B 10geA t x I0a logeA 2DK t 10b The slope in Figure 8c is 2DK where K 3 489 X 10 cm 2 we obtain the particle diffusivity D 5 08 X 10 cm s From Equation 9 with T 23 C and n 0 932 X 10 poise we obtain R 4 57 X 10 cm or 45 7 nm 457 Angstroms This diameter of 91 4 nm agrees very well with the nominal size of this particular Dow polystyrene latex standard Its mean volume averaged diameter is generally agreed to lie in the range 89 90 nary Unfortunately the simple straightforward analysis just discussed has only limited usefulness As must be obvious to all but the most casual observers most samples of practical interest differ appreciably from the uniform monodisperse case discussed in the previous section Real samples usually contain a range of sizes often of substantial width and are said to be polydisperse Such a particle size distribution might be conceptually simple consisting of a smooth single peak unimodal population of well defined mean diameter and width Or the distribution might be qualitatively more complex resembling two discrete peaks a bimodal distribution o
31. Equation 2 The constants f are the weighting coefficients which mix together the individual exponentially decaying functions exp D K t according to the relative amounts of the different particle diameters present in the sample solution of this more later In the trivial case of a single uniform particle size Equation 14 reduces to C t A f exp 2D K t B which is exactly the same as Equation 7 or lOa and b provided that the constant A f replaces the previous prefactor A Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 45 T mir AM DLS THEORY The goal of the ILT technique is to solve Equation 14 The quantity which is known is the left hand side the measured autocorrelation function C t where t is given by the discrete channels t At 2A 64 At The unknowns are the M individual weighting coefficients fi buried in the summation on the right hand side of Equation 14 This multi variable equation must therefore be inverted in order to yield the answer which is the set of weighting coefficients fit Equation 14 is in fact the discrete representation of a more general integral equation that defines the Laplace transform To appreciate this we can define a new reduced autocorrelation function H t obtained from the original C t H t C t ByAy We also define a new variable s which is simply proportional to the diffusion coefficient variable D
32. Menu File Used to access complete sets of parameters that were previously saved for the Control Menu F3 CNTL F7 Save Menu File Used to save or update an existing default TBL file which displays in the File Name window F10 Start measurement The instrument starts taking measurements once this option is initiated Alt_F10 Start Auto Save Print Used to initiate the automatic saving and printing of particle size results Please refer to the Tool Bar section of this manual for more detail Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 67 rity T qp it SAMPLE ANALYSIS ar The following are simple step by step instructions for successful use of the Nicomp Particle Sizer for the first time SAMPLE ANALYSIS MATERIALS Uniform latex particle reference material with a coefficient of variation less than 15 standard deviation as stated by the manufacturer Distilled filtered 0 2 micron water for dilution of reference material and cleaning glassware Autodilution 1 sterile 10cc syringe for each latex standard used 1 sterile 50 ml beaker 1 sterile pipet Drop in Cell 1 micropipet capable of delivering 15 ul solution 1 sterile 50 ml beaker 1 sterile pipet 1 6x 50 mm glass culture tube Important Reference material stated size is not an absolute Proper storage and handling of reference material over time is essential to reduce the possibility of aggregation and contaminat
33. SOFTWARE Nicomp Used to review the Nicomp distributions of an analyzed sample that had a chi squared value of 3 or higher The warning on the screen will indicate which of the two data fits is most appropriate based on assessment of the Chi Squared value INTENS WT NICOMP DISTRIBUTION solid Panicle Run Time 100 hr min sec 0 10 3 80 Fit Error 2 91 60 1 1 H 1 1 1 H f Residual 0 75 40 Gaussian Chi Squared 20 0 56 Ch 1 Data 0 i i i x1000 5 10 20 50 100 200 500 1854 Diam nm gt CHI Squared Small Use Gaussian Analysis PQ260 300 Diameter nm 1 269 5 nm S Dev nmp4 31 2 nm 11 6 Percent 100 0 6 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 50 Cumulative NICOMP SOFTWARE The coefficients of the Gaussian analysis will display as a cumulative sum starting at the lowest diameter and will increase towards the larger diameter Intensity Weighted Cumulative Result Gaussian INTENS WT CUMULATIVE DISTRIBUTION 100 60 60 40 20 0 1 50 100 200 500 1K 2K Diam nm gt PQ260 300 Mean Diam nm X Coeff of Var n Stnd Dev nm 263 0 0 059 15 518 Solid Particle Run Time hr min sec 0 10 3 Chi Squared 0 56 Auto B Adj 0 08 Ch 1 Data x1000 1854 Mean Diff Coeff cm2 sec 1 7e 008 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 51 NICOMP SOFTWARE Corr Data Used to view the block of raw data col
34. USE DISTRIBUTION ANALYSIS 81 5 23 X VOLUME MEAN DIAMETER STD DEVIATION WEIGHTING Figure 12b Volume weighted Gaussian Analysis Gaussian Analysis Solid Particles Chi Squared 1 3 Baseline adjust 0 00 X Data 454 5 K Mean Diffusion 5 03E 8 cm2 sec irte eene emm 4 gt Diameter log 10 100 92 2 9 INTENSITY MEAN DIAMETER STD DEVIATION WEIGHTING Figure 12c Intensity weighted Gaussian Analysis Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 41 T MM m hi DLS THEORY Gaussian Analysis Solid Particles Chi Squared 1 3 Baseline adjust Data 454 5 K Mean Diffusion 5 16E 8 cm2 sec 1 y 1 1 1 D t i D t t D t t t t i 4 i D 4 t t n 1 i t a gna ae tia dos eat gt Diameter log 10 100 90 0 9x VOLUME MEAN DIAMETER STO DEVIATION WEIGHTING Figure 12d Volume weighted Gaussian Analysis results for 91 nm latex after more Data 455K What appear to be large fluctuations in the Standard Deviation e g from 2 to 12 while the Mean Diameter with whatever weighting remains more or less constant While this factor of six variation in the Standard Deviation appears at first glance to be very large it really is not In fact all of the values within that range are effectively nearly equal Why Because they all describe autocorrelation functions which are very close to ideal single decaying exponentials Using Equation
35. a given MIN DIAM and Range In general this over resolution of the distribution can be rectified by decreasing the Plot Size which does not affect the MIN DIAM or Range Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 38 NICOMP SOFTWARE Read Menu File Access to complete sets of parameters that were previously saved for the Control Menu and the Auto Print Save Menu options can be gained using this option 1 Position the highlight bar over the Read Menu File option and click once the following window will display Look in CX Cw388 155 amp Zw388 157 amp Al ex a Cw388 tbl File name Cw388 t Cw388 tbl Files of type eb y Cancel IV Open as read only 2 Position the highlight bar over the table file of interest and click The selected table file with preset parameters will be retrieved and loaded from memory Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 39 NICOMP SOFTWARE Save Menu File This option is used to save or update an existing table of default values in a TBL file The name for the table file displays in the File Name window 1 Position the highlight bar over the table file name and click the mouse once The following window will display Save Menu File RR Save in CJ version 1 68 y amp e Edy E Cw388 tbl E test tbl ww EE Save as type tbl y Cancel 2 Either type the new file name that the table file parameters sho
36. and hence of R using Equation 2 A cursory examination of the three fluctuating scattering signals in Figure 4a b c suggests that extraction of the diffusion coefficient from the noise is not a straightforward matter Signal b clearly fluctuates faster than does c but is slower than a hence its particle size must lie between the values associated with a and c However obtaining quantitative information from these kinds of scattering signals is another matter altogether What comes to our rescue is the mathematical operation known as autocorrelation Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 7 RE AM DLS THEORY Autocorrelation function definition and motivation Let us consider the autocorrelation function of the net scattered light intensity Is t which fluctuates in time as shown in Figures 4a b and c The autocorrelation function which we denote by C t is used to study the correlation or similarity between the value of I at a given time t and the value of at a given time t and the value of at an earlier time t t This comparison is then made for many different values of t in order to obtain a good statistical average for C t i e averaged over many wiggles of the fluctuating intensity Is C t is evaluated according to C t lt Is t Is t 8 gt 3 The bracket symbols lt gt are shorthand for a summation over many values of t That is one calculates a runnin
37. be drawn into the instrument do not force all of the contents The system will dilute the concentrated sample injected When the proper concentration is reached the system will automatically stop the pump and begin taking data The system will automatically switch to a data acquisition screen After a few minutes of equilibration and set up time the system will begin displaying the differential intensity weighted distribution on the screen Press T to switch the screen to the time series plot Allow time for the data to stabilize where the mean diameter is not changing for at least 3 minutes as a function of time A flat straight line Press F to go back to the distribution plot in Intensity weighted mode Verify the size standard to the reported size by following the section Interpretation of Data Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 6 4 SAMPLE ANALYSIS R Drop in Cell 1 2 Ensure that parameters are set according to the below Auto Print Save Menu Menu File CAPSS Software cw388 version 1 68 Cw388 tbl Data Directory Es Browse File Name test 0 Printout IB 261 nm latex standard Auto Operation Options No Print Save Cycles EA Using Run Time 5 mn C Using Fit Error lt Ea I with Chi Squared gt PS Clear Autocorrelator Print Result Printout Option Automatic Choice of Distrib Gauss vs NICOMP IV Store Data on Disk Overwrite Old File Cancel
38. been created in the same data directory Ctrl_F Turn On Flow Pump Press ESC to Stop Used to flush the sample out of the system G Toggle Gaussian Nicomp Distribution Used to toggle between the Gaussian and Nicomp interpretations of the data CTRL G Reset Fiber Angle to 90 deg for Multi angle system Aligns the fiber at the 90 mark for data acquisition This option is used in conjunction with the Multi angle option Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 62 l K L M P rity Til iT jT NICOMP SOFTWARE JAIL Edit NICOMP Input Menu This function will change the fit of the Nicomp distribution to the new parameters specified The input parameters min diam plot size smoothing and range along with their identifying labels may be changed using this option Display Autocorrelation Function Curve and Intensity weighted Gaussian Summary Used to observe the autocorrelation function produced by the scattered intensity data The value of Decays lies in the approximate range 1 7 to 2 7 This is the number of exponential decays spanned by the 64 channels of the autocorrelation function Change Distribution for Line Graph The format in which data displays can be changed from bar mode default to line mode Edit Correlator Control Menu Allows for the edit of te Control Menu Please refer to the Particle Sizing section of this manual for additional information concerning t
39. change in elasticity or color etc then it is probably safe to subject the fluidics system to the solvent However in any case operate conservatively When the measurement is completed the new solvent should be flushed out of the system first by running the pump Ctrl F on air allowing most of the solvent to exit via the drain line and then replaced with water Run sufficient water through the system to insure complete replacement by dilution of any remaining solvent Additional replacement tubing or tubing of a different composition can be obtained directly from Cole Parmer The tubing is size 16 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page C 1 APPENDIX C Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page C 2 Fluid Acetaldehyde Acetate LHW Acetic Acid gt 5 Acetic Acid lt 5 Acetic Anhydride Acetone Air Aliphatic Hydrocarbons Aluminum Chloride Aluminum Sulfate Alums Ammonia gas liquid Ammonium Acetate Ammonium Carbonate Ammonium Chloride Ammonium Hydroxide Ammonium Nitrate Ammonium Phosphate Ammonium Sulfate Amyl Acetate Amyl Alcohol Amyl Chloride Aniline Aniline Hydrochloride Aqua Regina 80 HC1 20 H Aromatic Hydrocarbons Arsenic Salts Barium Salts Benzaldehyde Benzenesulfonic Acid Bleaching Liquors Boric Acid Bromine Butane Butanol Butyl Alcohol Butylacetate Butyric Acid Calcium Oxide Calcium Salts Carbon Bisulfide Carbon Dioxide Carbon Tetrachloride Nicomp
40. cloudy Draw approximately 1 mL of this turbid suspension into a sterile 3cc or 10cc syringe Fill the glass culture tube with latex solution and discard solution twice to clean tube Fill a glass culture tube within 5mm of the opening Clean the tube with lint free tissue and place the tube in the sample holder of the Nicomp A wait time of 5 minutes will allow the sample to temperature equilibrate Check the concentration of the latex solution by closing the port on top of the Nicomp and reading the intensity value on the front panel of the instrument Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 6 6 n Tum Pt SAMPLE ANALYSIS Rub 15 Click the increase and decrease neutral density filter setting buttons on the tool bar until the intensity hovers at approximately 300 kHz 16 Click on the Green G icon to start sample measurement and follow software instructions on the screen Important The instrument will adjust the channel width to the optimal value The system will automatically switch to a data acquisition screen 17 Press T to switch the screen to the time series plot Allow time for the data to stabilize where the mean diameter is not changing for at least 3 minutes as a function oftime A flat straight line 18 Press F to go back to the distribution plot in intensity weighted mode 19 Verify the size standard to the reported size by following the section Interpretation of Data Nicomp 380
41. for Chi Squared Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 32 rity Til iT jT NICOMP SOFTWARE JAIL 1 Click on the square box located to left of the selection to include the Chi Squared parameter The lower limit for Chi Squared will appear on the same line and to the right 2 Enterthe desired Chi Squared value Clear Autocorrelator The decision to Clear the autocorrelator at the end of each Auto Operation Cycle is made by setting this parameter Clearing of the Autocorrelator can performed in preparation to run another sample Clear Autocorrelator Click on the square box located to the left of the selection and a check will display Each cycle then will consist of a new independent measurement No Clearing of the Autocorrelator Do not click on the box so light scattering data will continue to be acquired at the End of each cycle thereby allowing the statistical accuracy of the autocorrelation function to improve over time This is the preferred mode of operation Print Save Options As indicated in the previous section after completion of an automatic Operation Cycle one or more Print Save options will be implemented Each parameter may be set using this option A brief description of each follows Print Result Click on the square located to the left of the selection and a check mark will display in the box A printout will result at the end of each Auto Operation Cycle If a relatively large nu
42. for Mie scattering when intraparticle interference effects become important for diameters greater than about 150 nm in order to obtain number weighted and volume weighted diameter distributions from the starting result which is an intensity weighted plot Note Volume weighting and mass weighting are equivalent terms The volume weighted and number weighted video displays corresponding to the example discussed above are shown in Figure IOb and lOc respectively The three display plots Figure lOa b and c reveal an important property that is generally observed when using the Gaussian Analysis That is the value of the mean diameter can vary significantly with the choice of weighting depending on the width of the Gaussian like distribution In general the wider the distribution i e the larger the percentage standard deviation the greater the differences in mean diameter for the various weightings In the present example we see that the mean diameter of the volume weighted peak 209 1 nm is shifted almost 10 below that of the intensity weighted result 226 1 nm The number weighted mean diameter is reduced even more dramatically to 144 7 nm Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 30 MM TQ at DLS THEORY m Gaussian Analysis Solid Particies Chi Squared 1 1 Baseline adjust 0 01 X Data 1047 0 K Mean Diffusion 2 22E 8 cm2 sec dme ee Wm TUTO ts gt Diameter log 22 30 VOLUME MEAN
43. for the autocorrelation function for a particular choice of t C t one must obtain many products I t I t t using many different values of t for each value of t Only in this way will one average the value of C t over sufficiently many bumps and wiggles in the fluctuating signal to obtain a statistically meaningful value of the autocorrelation function Then one must repeat this process for sufficiently many values of t so as to obtain a well defined smooth representation of C t as a function of t It is useful to have an idea at this point of the kinds of numbers that are involved when we use the word many For a typical particle size measurement of duration 5 minutes on 0 2 micron 200 nm particles the DLS Module performs approximately 15 million multiplications in order to obtain C t for one value of t e g t 20 microseconds for channel 1 The instrument makes 64 such sets of calculations simultaneously in order to obtain C t for 64 different values oft The essential point about the autocorrelation function is that it serves as a useful probe of the characteristic lifetime or duration of the fluctuations in t That is once the interval t between two sampled intensities exceeds the average width of a major bump or fluctuation in Is t the two sampled intensity values will cease on average to be correlated At this point the value of C t will have fallen substantially Nicomp 380 Ma
44. from different regions of the particle interfere at the distant point of detection The resulting total scattered intensity is therefore diminished relative to the values given by Equations la and b which assume that all of the effective scattering mass is packed into a very small particle size The expressions in Equations la and 1b can be repaired to include the effects of interference by multiplying them by a so called Mie form factor this quantity has a limiting value of 1 0 i e no effect in the Rayleigh region but falls below unity in a non monotonic way as the particle size grows Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 2 ME Toot at DLS THEORY m Using Equation la or Ib one can in principle determine either the molecular weight or the volume of the particles from a measurement of the scattered intensity using known calibration standards together with empirical determinations of f ny n and g Nnp ns This forms the basis for the technique of classical light scattering The newer DLS method however departs radically from this traditional approach to light scattering The quantity of interest is no longer the magnitude per se of the scattered light intensity Rather DLS concerns itself with the time behavior of the fluctuations in the scattered intensity Dynamic scattering the effects of diffusion To understand why the scattered intensity fluctuates in time we must appreciate that it is the
45. in cell CE MARK The CE mark officially CE marking is a mandatory marking on certain products which is required if they are placed on the market in the European Economic Area EEA By affixing the CE marking the manufacturer or his representative or the importer assures that that the item meets all the essential requirements of all applicable EU directives The CE mark is a mandatory European marking for certain product groups to indicate conformity with the essential health and safety requirements set out in European Directives To permit the use of a CE mark on a product proof that the item meets the relevant requirements must be documented This has been achieved using an external test house which evaluates our particle size analyzers and its documentation CE originally stood for Communaut Europ enne or Conformit Europ enne French for European Conformity The following label is affixed to the back panel of the AccuSizer SIS to indicate that the instrument has passed CE mark testing and conforms to the European Union Directives for Electromagnetic Compatibility EU EMC CE Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 1 3 TAE DLS THEORY m i DYNAMIC LIGHT SCATTERING THEORY In recent years the technique of dynamic light scattering DLS also called quasi elastic light scattering QELS or photon correlation spectroscopy PCS has proven to be an invaluable analytical tool for characterizing the siz
46. is one of the ways to ensure a measurement is progressing in the proper manner Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 45 NICOMP SOFTWARE Gaussian Provides the ability to view the analysis in Gaussian format Intensity Weighted Gaussian 100 80 60 40 20 0 20 Diam nm gt PQ260 300 Mean Diam nm X Coeff of Var n Stnd Dev nm 263 0 0 059 15 518 INTENS WT GAUSSIAN DISTRIBUTION Solid Particle Run Time hr min sec 0 10 3 Chi Squared 0 56 Auto B Adj 0 08 Ch 1 Data x1000 1854 Mean Diff Coeff cm2 sec 1 7e 008 When displaying the data scroll through the different weightings of the data set by clicking on the Weighting pull down menu or by pressing W on the key board Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 46 TT 3 NICOMP SOFTWARE Volume Weighted Gaussian VOLUME WT GAUSSIAN DISTRIBUTION oma ane Run Time hr min sec 0 10 3 80 Chi Squared 0 56 60 t t t t T Auto B Adj 0 08 40 Ch 1 Data x1000 20 i 1854 Mean 0 N i i i i N Diff Coeff 20 cm2 sec Diam nm gt 1 7e 008 PQ260 300 Mean Diam nm X Coeff of Var n Stnd Dev nm 253 1 0 059 15 520 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 47 NICOMP SOFTWARE Number Weighted Gaussian NUMBER WT GAUSSIAN DISTRIBUTION sella Paride Run Time 100 hr min sec i 0 10 3
47. like to be able to obtain with reasonable accuracy all values f there are M of them thereby obtaining a faithful representation of the true particle size distribution In practice however one falls far short of this goal Typically it is possible to obtain a stable distribution containing one two or even three peaks In the case of two peaks we obtain two values of peak diffusion coefficients D4 and D and one relative strength parameter f f for a total of three independent parameters in addition to the baseline adjust Note in all of the distribution results of the DLS Module the maximum height of any distribution is always normalized to 100 hence in the case of two peaks only the ratio of their heights is an independent parameter In the case of three peaks one will obtain three peak D values D D and D and two relative strength parameters f2 f and fy f Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 47 RE AM DLS THEORY The Nicomp allows one to establish the set of diameter values 1 to M over which Equation 14 is defined The total number M is given by the PLOT SIZE the values vary from MIN DIAM i to RANGE MIN DIAM i M In most cases it is desirable to use the logarithmic diameter scale in which the individual diameter slices are spaced logarithmically Each of the diameter values is converted to a diffusion coefficient Di the resulting individual decaying exponential fun
48. of the concentrated sample in the syringe Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 6 3 Tl ir 14 15 16 17 18 19 T An SAMPLE ANALYSIS RUN NOTE Prior to introducing the sample into the Nicomp the flow through system must be flushed with fresh filtered diluent to an intensity of less than 10 kHz as displayed on the front panel screen This can be accomplished by following the below a Click on Flow Pump icon The system pump will begin running An internal timer will allow the pump to run for two minutes and then stop automatically Important Press ESC to stop the pump prior to the two minute setting b Check to see that the following two conditions are being met c Water should be exiting from the outlet tube in a continuous flow without bubbles d The light scattering intensity indicated by the digital LED display on the front panel of the Nicomp should be below 10kHz indicating a relatively well flushed clean scattering cell If either of these conditions is not met an additional flush cycle should be performed Click on the Autodilution icon or the Green G The system will prompt to inject the sample Turn the valve on the front panel 90 counter clockwise and inject approximately 1 mL of concentrated material Turn the valve on the front panel 90 clockwise to its original position after approximately 1mL of concentrated solution has been drawn into the instrument Sample will
49. particles or macromolecules suspended in solution The word estimate is emphasized because of the fact that the measurement of molecular weight using the technique of dynamic light scattering DLS is not as accurate as the determination of particle size for which all DLS based instruments are primarily designed The fundamental quantity measured in a DLS based instrument is the particle diffusivity or diffusion coefficient D There is a simple empirical formula that can be used to relate the molecular weight M of a suspended particle or macromolecule to its diffusivity D D2 MM E 1 The pre factor constant is related to the specific composition of both the diffusing particles and the surrounding solvent Constant B in the exponent is related to the shape configuration of the particles and macromolecules which may also be a function of particle and solvent composition as in the case of polymers The form of Equation E 1 can be motivated by considering the simple case of solid spherical particles of uniform radius R How is it related to M for such ideal particles Using the Stokes Einstein relation D can be related to R D kT 6Mn 1 R E 2 which can be re written D C R E 3 where C is a constant C4 kT erln For particles for a given density molecular weight My is proportional to the particle volume My C 4 3 NR E 4 where C is a constant of proportionality related to the mass density of the particles
50. run vs diameter The value of the number weighted particle size distribution is also calculated assuming that the particles are spheres of uniform density which scatter light according to classical Mie Theory Intensity weighted The result first displays from either of the autocorrelation functions Displays the relative intensity of scattered light vs diameter for a sample run X Start Automatic Channel Width Adjustment Control of setting the optimum Channel Width for the sample being measured is left to the instrument Z Zero NICOMP Distribution Channels Used to zero the three smallest diameter bins on both the computer display and printout so that they do not dominate the remaining portion of the size distribution plot These zeroed bins are still included in the Distribution Analysis fit but will not be included in the calculation of the Mean Diameter which appears on the printout These bins will appear as zeroes 000 on the printout but their actual contributions will be printed along side the other values next to the particle size histogram the relative numbers for the Zero bins may exceed 100 This option does not change the computed fit to the data only the presentation of the results Decrease sensitivity Used to lower the sensitivity to achieve the optimum Photopulse Rate Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 65 T rh roman AM N NICOMP SOFTWARE Alt_T F1 F2
51. size and width of the resulting droplet distribution are usually sensitive functions of the stoichiometry of the starting compounds and the duration and detailed nature of the preparation technique employed In Figure 9a we show the autocorrelation function for a fat emulsion used for intravenous IV feeding The channel width used here was 21 usec Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 21 T MM m hi DLS THEORY ix 1047006 xk Correlation Function xx as x zkt xxXx zxz XXXXXE ARA xxkkxxk XZ3 SXXXXE x 877427 4 1 1 i 4 1 1 1 V L L i 4 t 4 LI L i 3 i t i t I 3 i Li 1 U 847726 une 4 4 42 4 denn 4 Intensity Weighted Gaussian Summary DIA 226 1 NM STD DEV 0 29 DECAYS 2 0 CHI SQ 1 10 BASE ADJ 0 01 X Figure 9a Autocorrelation function for an IV fat emulsion Let us make a visual comparison between Figure 9a and 8a obtained for the narrow 91 nm latex standard The shapes of the two decaying curves appear to be quite similar which is somewhat surprising given the differences between the two samples Qualitatively we conclude that the average or characteristic particle diameter associated with Figure 9a must be roughly twice that associated with Figure 8a The reason both curves possess about the same number of decays in falling to the 64th channel and the channel width for the latter s
52. sufficiently small choice of t Next let us consider a larger value for t equal to t as shown in Figure 5 In this case t7 has been chosen to be large enough relative to the time scale of the fluctuating signal that the two sampled values of I Is t and I t tz are now somewhat different In this case the two sampled intensities are less well correlated However there still remains some relationship between these two intensities If t has been chosen so that I t is near a minimum in the intensity then I t t2 will still be a relatively low value Similarly if t lies near a maximum then it is apparent from Figure 5 that Is t t2 must also be at a relatively high value or certainly not near a minimum given the fact that t is not a very large time interval relative to the characteristic time scale of the intensity signal shown in Figure 5 Finally we consider a very large time interval t3 as seen in Figure 5 Here we see that t5 is so large that has undergone two large fluctuations between the two sampling times t and t tz It is clear here that the two sampled intensities will in general be almost completely uncorrelated for such a large choice of t The two values could easily be both high both low one high and the other low or any other intermediate possibility We have carried out these examples assuming a single choice for time t and three different values of t In order to obtain a meaningful value
53. t l l 1 i i f l RUN TIME x Hours 14 Mins 48 Secs AVG COUNT RATE 298 0 kHz CHANNEL WIDTH 21 0 USEC TEMPERATURE a 23 DEGREES CENT VISCOSITY 0 933 CENTIPOISE INDEX OF REFRACTION 1 333 COUNT RATE SETPOINT 300 ALPHA Molec Wt 1 0 BETA Molec Wt 0 50 Figure 11a Printout of volume weighted Gaussian Analysis result for fat emulsion See Figure lOa Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 33 DLS THEORY Gaussian Analysis Solid Particles 1 Fat emulsion test sample SIZE nanometers VOLUME REL Cu T 0 Hours 14 Mins 48 Secs RUN TIME AVG COUNT RATE 298 0 kHz CHANNEL WIDTH TEMPERATURE VISCOSITY 0 933 INDEX OF REFRACTION 1 333 COUNT RATE SETPOINT 300 ALPHA Molec Wt 1 0 BETA Molec Wt 0 50 21 0 23 USEC DEGREES CENT CENTIPOISE Figure 11b Printout of volume weighted Gaussian Analysis result for fat emulsion See Figure IOb Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 34 PERCENT RESULTS Gaussian Analysis Volume Weighting Mean Diameter 209 0 nm Standard Deviation S rm 29 9 x Chi Squared 1 1 Baseline Adjust 0 01 Mean Diffusion 2 88E 8 cm sec Data 1047 0 K Cumulative Results Diam s 151 0 50 184 8 75 227 0 99 376 0 Percent 2 DLS THEORY Gaussian Analysis Solid Particles i Fat emulsion test sample ae nanometers
54. technique is to determine the diffusion coefficient D of the particles assumed uniform here from the raw data i e the fluctuating light scattering signal as represented in Figure 4a b c From D we can easily calculate the particle radius R using the well known Stokes Einstein relation D kT 6xnR 2 where k is Boltzmann s constant 1 38 X 10 erg K T the temperature K C 273 and n the shear viscosity of the solvent e g n 1 002 X 10 poise for water at 20 C Thus we see that the rate at which the particles jitter about in the suspension as measured by D is inversely related to the particle radius R From Equation 2 we see that in general the diffusion coefficient D of particles of a given size increases with increasing temperature T This is due primarily to the T dependence of the solvent viscosity n The fact that T is the numerator in Equation 2 is less small in percentage when expressed in deg Kelvin For example n for pure water falls to 0 890 X 10 poise at 25 C i e more than a 10 change from the value at 20 C Clearly the less viscous the solvent the more rapid will be the random walk diffusion of the particles and the faster the resulting intensity fluctuations Hence changes in T are completely indistinguishable from changes in particle radius R as they affect D For this reason the sample temperature MUST be constant and accurately known in order to obtain a meaningful measurement of D
55. to more difficult samples for which the results of the Gaussian Analysis show larger fluctuations and require a longer time to settle Next we examine another sequence of results obtained from the Gaussian Analysis as a function of accumulating data this time returning to the narrow nearly monodisperse 91nm latex standard It is useful to compare these results summarized in Table 2 with those just discussed for the broad fat emulsion Again as was found for the broad fat emulsion the value of Chi Squared is a poor indication of the extent of settling of the Gaussian Analysis results at the beginning of a run Data 64K when insufficient data has been collected The abnormally large value of the Standard Deviation 23 the result again of insufficient data causes the volume weighted Mean Diameter to be pushed well below the intensity weighted value However after additional time has elapsed the Standard Deviation settles down to 8 or 9 At this point there is much less discrepancy between the two Mean Diameter values based on the different weightings Data Int Wt Std Dev Vol Wt Chi Sq Base Ch 1 MeanDiam Mean MeanDian Adj 64K 93 7 nm 23 81 5 nn 115 93 1 12 89 4 166 92 6 90 6 218 92 4 1 3 92 4 91 1 92 2 90 0 Table 2 Sequence of Gaussian Analysis results obtained for the narrow 91 nm latex standard Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 39 T mir Toot ait AM N DLS THEORY
56. true particle size distribution may be The statistical accuracy that can be realistically achieved in the underlying autocorrelation data simply cannot support an analysis which reliably yields any more detail in the distribution However it usually turns out that the resolution and accuracy of the NICOMP algorithm are sufficient to yield a meaningful representation of the true particle size distribution for many systems of practical significance A simple test of Distribution Analysis uniform particle size latex In order to investigate the capability of the NICOMP Distribution Analysis we must apply it to a polydisperse sample which differs qualitatively from those investigated earlier i e those consisting of simple unimodal populations which could be well characterized using the Gaussian Analysis First however we should check the performance of the new algorithm using a narrow nearly monodisperse polystyrene latex standard For such a sample it must be able to determine the mean particle diameter with high accuracy In Figure 13 we see the video display of the volume weighted distribution obtained for the 91nm latex standard The corresponding result obtained using the Gaussian Analysis was shown earlier in Figure 12d We find a mean diameter of 88 8 nm for Figure 13 compared Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 49 T mir AM DLS THEORY Gaussian Analysis Solid Particles chi Squared 0 2 Baseline
57. weight of the particles reliable estimates of constants o and R must be available These quantities are related to the Mark Houwink equation for the intrinsic viscosity n of dilute solutions containing suspended particles or macromolecules given by n CuAMw E 9 where constants C and a are respectively a prefactor and exponent Equation E 9 provides one operational method for obtaining and amp for a given particle solvent system Suppose one has available several samples of the same composition but different known Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page E 2 ME hmi APPENDIX E m molecular weights One can then measure and plot the experimental value of n as a function of the known My values the slope of a plot of log n as a function of the known My values the slope of a plot of log n vs log Mw yields the exponent a The desired constant amp can then be obtained from exponent a using the equation R a 1 3 E 10 Constant can be obtained for a measurement of the diffusivity D for one of the known samples using Equation E 1 As an alternative to measuring the intrinsic viscosity one could simply measure D for related homologous series of particles of macromolecules of known My From Equation E 1 we have log D log R logMw E 11 A plot of D vs log M should yield a straight line the slope is equal to negative R and the intercept yields log from which is obtaine
58. 00 2305 3459 Fit Error 18 03 38 52 61 48 7 349 INT 288 9nm 52 1nm 3 86 0 00 2308 3453 Residual 18 03 42 91 57 09 0 000 NUM 263 8 nm 47 6nm 3 86 0 00 2301 3453 18 0396 47 72 52 28 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 9 APPENDIX A GAUSSIAN NICOMP ALL WEIGHTED Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm7 3 DataFile CProgramFilesiParticle Siang SystemsiNicomp388Wwersion 1 68 Bimodal 300 GAUSSIAN NICOMP DISTRIBUTION Analysis Solid Particle Fit Error ChiSquared RunTime CountRate Channel 1 ChannelWidth 7 35 Residual 0 00 Baseline Adj 0 00 96 0 Hr 6 Min 8 Sec Wavelength 632 8 nm 0 KHz Temperature 23 degC 6126 K Viscosity 0 933 cp 35 0 uSec Index ofRef 1 333 Ww co oa GAUSSIAN DISTRIBUTION Intensity Weighting MeanDiameter 283 9nm 100 E StndDeviation 52 1 nm 18 03 96 80 Volume Weighting on MeanDiameter 295 5nm pe h StndDeviation 53 3 nm 18 0396 20 Number Weighting MeanDiameter 263 8nm StndDeviation 247 6 nm 18 03 96 0 20 50 100 200 500 Diam nm gt intensity Weighting Peak 1 Peak2Peak3 MeanDiam nm 230 8 3453 Percent 96 42 91 57 09 Volume Weighting Peak1Peak2Peak3 MeanDiam nm 230 5 3459 Percent 38 52 6148 Number Weighting Peak 1 Peak2Peak3 MeanDiam nm 230 1 3453 Percent 36 47 72 5228 Intensity Wt Volume W
59. 06 06 Page D 4 TEMP C 20 25 40 20 20 23 25 40 15 30 20 15 30 20 40 20 25 30 40 15 30 15 25 15 30 15 20 25 20 30 40 VISCOSITY cpoise 0 409 0 386 0 341 3 45 0 326 0 3068 0 294 0 271 4 703 2 876 0 223 2 86 1 77 0 381 0 320 0 597 0 547 0 510 0 456 0 423 0 365 0 360 0 328 0 449 0 393 2 24 2 03 0 620 2 37 1 91 1 63 INDEX REFRACTION 1 388 1 388 1 388 1 433 1 375 1 375 1 375 1 375 1 397 1 397 1 355 1 385 1 385 1 380 1 380 1 326 1 326 1 326 1 326 1 379 1 379 1 346 1 346 1 424 1 424 1 550 1 550 1 380 1 547 1 547 1 547 TT Ti iT y v APPENDIX D Au SOLVENT TEMP C VISCOSITY cpoise INDEX REFRACTION m Nitrotoluene 20 2 33 1 545 30 1 77 1 545 40 1 60 1 545 p Nitrotoluene 60 1 204 1 533 n Octane 20 0 542 1 395 40 0 433 1 395 Pentane 20 0 240 1 357 Propyl acetate 20 0 59 1 382 40 0 44 1 382 n Propyl alcohol 20 2 256 1 385 30 1 72 1 385 40 1 405 1 385 1 1 2 2 15 1 844 1 494 Tetrachloroethan Toluene 20 0 590 1 494 30 0 526 1 494 40 0 471 1 494 Trichlorethane 20 1 20 1 438 o Xylene 16 0 876 1 506 20 0 810 1 506 40 0 627 1 506 m Xylene 15 0 650 1 495 20 0 620 1 495 40 0 497 1 495 p Xylene 16 0 696 1 493 20 0 648 1 493 40 0 513 1 493 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page D 5 ME Tat iT APPENDIX E AA ESTIMATING MOLECULAR WEIGHT It is possible to use the Nicomp to estimate the average molecular weight M of
60. 0Nicomp 030806 06 06 Page 5 59 NICOMP SOFTWARE HELP MENU Index Using Help About CW388 Index The index is a listing of the older keystroke commands that have been replaced by icons buttons and menu choices Using Help At this time the default Window help is available About CW388 This window will provide the user with the version number of the software in use About CW388 Cw388 Application Version 1 68 Nicomp Copyright 2001 NICOMP Particle Sizing Systems Santa Barbara California USA Tel 805 968 1497 Fax 805 968 0361 ES Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 60 rity Til iT jT NICOMP SOFTWARE JAIL COMMAND KEYS The following set of commands will display any time that ALT H is accessed while operating the C380 software A Display Gaussian NICOMP Intensity Volume Number Weighted Distribution Both the Gaussian and NICOMP distributions along with the Intensity Volume and Number weighted information will display when this key is used ALT A Toggle Number Weighted Area Weighted Distribution Both the Number and Area weighted distributions for a sample analysis may be reviewed using this option B Change Distribution to Bar Graph The format in which data is displays can be changed from line mode to bar mode default It is helpful to change to line mode when reviewing overlays of the distributions C Clear correlator Cle
61. 1 905 0 716 0 009 0 389 0 281 1 597 2 136 2 465 2 373 0 835 1 159 0 200 0 627 1 799 1 333 1 464 1 565 0 341 1 855 2 963 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 53 NICOMP SOFTWARE Time History Displays the time history plot where the stability of the data in terms of mean diameter is shown to be changing as a function of time File View Setup Particle Sizing Display Weighting Help zem ejelolwjilw mj 7 0 al Sala eae Diam nm GAUSSIAN CALCULATION HISTORY Run Time T hr min sec 0 6 8 au exl Count Rate xi000 0 250 esu 186 A A E 0 4 8 12 16 Runtime min gt Intensity Wt E Volume Wt a Number Wt Printout ID 350nm 220nm 7 3 For Help press F1 Read File Bimodal 300 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 54 NICOMP SOFTWARE The Gaussian time history shows the intensity volume and number weighted means on a single screen while the Nicomp time history shows different peaks In order to view the Nicomp time history plot select Nicomp from the Display pull doen menu Diam nm INTENS WT NICOMP CALCULATION HISTORY 350 300 250 200 150 100 50 0 1 Runtime min gt Peak1 Peak 2 Peak3 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 55 NICOMP SOFTWARE Summary Result Displays the Volume Intensity and Number weighted data for the Gau
62. 11 we are saying that the curvature represented by coefficient a is nearly zero for the above mentioned range of Standard Deviations There are small changes in the least squares quadratic fit due to tiny fluctuations in the correlation data which are responsible for these seemingly large changes in the Standard Deviation Indeed these small fluctuations may be caused by normal Poisson statistics particle aggregates dirt particles stray scattered light etc By contrast broad distributions result in substantial curvature in the quadratic fit i e relatively large values of a and much less sensitivity to small fluctuations in the correlation data Hence in this case the Gaussian Analysis results are observed to settle relatively more quickly to a final reliable answer Typically it is necessary to start with very clean well Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page2 42 ME DLS THEORY m l dispersed samples and acquire data for a relatively long time e g 30 to 60 minutes to obtain reliable Standard Deviations smaller than 8 to 1096 for a sample which is known to be nearly monodisperse Under such controlled ideal conditions the DLS Module is certainly capable of yielding single peak distributions of exceptionally small Standard Deviation Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 43 T mir AM DLS THEORY NICOMP DISTRIBUTION ANALYSIS Rationale for use of the inverse Laplace tr
63. 380 Manual PSS 380Nicomp 030806 06 06 Page 1 2 T T T Tr f T GENERAL INFORMATION An Access to the sample cell holder necessary for inserting or removing a sample cell is provided by a square opening at the front left corner of the top cover of the instrument A rectangular dust cover with handle and three thumb screws are provided to keep the scattering cell and internal optical components free of excessive amounts of dust when the unit is not in use for extended periods of time and to prevent the laser light from scattering outside the unit during operation During normal operation this cover can be secured with one screw and swung to one side to provide easy access to the cell holder It can be swung shut during operation to keep out stray room light and keep in beam light being scattered by the particles During operation of the NICOMP Autodilute Submicron Particle Sizer the Top Cover of the unit Must Remain Closed i e attached to the cabinet by means of the 3 screws provided The Warning label on the cover warns of the possible exposure to the laser beam a minimum of 5 milliwatts 632 8 nm wavelength if the top cover is removed for any reason while power is applied to the unit Important Any attempt to remove the front panel while the instrument is in operation may result in possible Direct Exposure to Dangerous Laser Radiation Also power must be off to the unit if the Autodilution cell is being replaced by the drop
64. 5 43 NICOMP SOFTWARE DISPLAY The following provides a brief summary of the Display options that are used during the use of the Windows CW388 Software Access of the Display options 1 Position the highlight over the DISPLAY option and click once using the mouse The following window of File options displays Corr Function K Gaussian ALT G NICOMP ALT N Cumulative Q Corr Data D Channel Error Ctrl E Time_History x Summary Result ALT S GAUSS NICOMP A Show Distribution F Time Plot Scale Ctrl B 2 Position the highlight bar over the desired selection and click once Following is a description of each of the options offered Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 44 NICOMP SOFTWARE Corr Function This option is used to observe the autocorrelation function produced by the scattered intensity data The value of Decays should lie sin the appropriate range 1 7 to 2 7 This is the number of exponential decays spanned by the 64 channels of the autocorrelation function counts X1000 AUTOCORRELATION FUNCTION Solid Particle eue Run Time 1800 hr min sec 0 10 3 1750 1700 Ch 1 Data 1650 x1000 1854 1600 1550 Count Rate 1500 x1000 377 1450 1371 34 900 1400 uSec gt Channel Width 34 0 uSec INTENSITY WEIGHTED GAUSSIAN SUMMARY Mean Diam nm Coeff of Var n Chi Sq Decays Auto Baseline Adj 263 023 0 059 0 563 2 512 0 082 Viewing the correlation function
65. 696 0 66 0 493 0 920 0 853 0 84 0 80 INDEX REFRACTION 1 372 1 372 1 400 1 400 1 400 1 628 1 628 1 459 1 459 1 459 1 459 1 523 1 523 1 523 1 444 1 444 1 444 1 426 1 426 1 456 1 456 1 450 1 450 1 445 1 445 1 404 1 409 1 409 1 427 1 427 TT REI y T APPENDIX D AA SOLVENT TEMP C VISCOSITY cpoise INDEX REFRACTION Dimehtylaniline 20 1 41 1 558 30 1 17 1 558 40 1 04 1 558 n Dodecane 25 1 35 1 415 Ethyl Acetate 15 0 473 1 380 20 0 455 1 380 25 0 441 1 380 30 0 400 1 380 Ethyl alcohol 20 1 200 1 359 Ethanol 30 1 003 1 359 40 0 834 1 359 Ethyl benzene 15 0 697 1 495 30 0 581 1 495 Ethyl bromide 15 0 418 1 424 20 0 402 1 424 30 0 348 1 424 Ethyl ether 20 0 233 1 352 25 0 222 1 352 40 0 197 1 325 Ethyl formate 15 0 419 1 361 30 0 358 1 361 Ethylene bromide 17 1 950 1 538 20 1 721 1 538 Ethylene dichloride 15 0 887 1 445 30 0 730 1 445 Ethylene glycol 20 19 90 1 431 30 13 35 1 431 40 9 13 1 431 60 4 95 1 431 Formamide 25 3 30 1 446 Formic acid 20 1 804 1 371 30 1 465 1 371 40 1 219 1 371 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page D 3 T mir AN y APPENDIX D SOLVENT n Heptane n Hexadecane n Hexane Isobutyl alcohol Isopentane Isopropyl alcohol Methyl acetate Methyl alcohol Methanol Methyl ethyl ketone M EK Methyl formate Methylene dichloride Nitrobenzene Nitromethane o Nitrotoluene Nicomp 380 Manual PSS 380Nicomp 030806
66. 88 v Oe E version 1 68 sj Bimodal 300 Files oftype t 7 Cancel v Open as read only 1 Position the highlight bar over the data file of interest and click the mouse once Prior to the selected data file being retrieved from memory the following window will display 2 Click on Yes the data will be re calculated No the window will disappear and the volume weighted distribution of the file selected will display Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 3 NICOMP SOFTWARE Read New A data file that has been stored following a measurement can be retrieved to display the resulting particle size distribution PSD with the desired weighting When this option is selected a list of data files will display in the Read Data File window such as in the example below Read New Data File Look in B Nicomp 388 y O ex Ely version 1 68 E Bimodal 300 wm EHE 000000 Files of type lc v Cancel v Open as read only 1 Position the highlight bar over the data file of interest and click the mouse 2 Click on the OK button The selected data file will be retrieved from memory Once again the CW388 Window for re calculating the data will re display Please refer to the Read section of this manual for additional information Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 4 NICOMP SOFTWARE Save This option is used throughout the CW388 software whenev
67. C t 2 2 3 0 1 3 4 1 etc In the following section we shall generalize our discussion to include polydisperse systems those that contain a mixture of particle sizes We shall discuss in some detail the methods by which the DLS Module is able to deal effectively with these more complex systems These are the so called algorithms for analysis of the autocorrelation function which yield estimates of the true particle size distribution The next section will be heavily weighted toward results i e actual pictures of screen displays and printouts and less concerned with theoretical details Hence if you have survived the proceeding discussion and equations you have our congratulations and assurance that you should experience clear sailing in the section ahead In any case you are STRONGLY URGED to read Section C carefully It should help you appreciate the power of the DLS technique when applied to difficult particle size distributions like those that are frequently encountered Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 15 DLS THEORY THE SIMPLEST APPROACH TO SIZE DISTRIBUTIONS GAUSSIAN ANALYSIS Uniform particle size trivial analysis In Figure 8a we see a video display of the 64 channel autocorrelation function obtained using DLS for a 90 nm 0 090 micron polystyrene latex particle size standard counts X1000 AUTOCORRELATION FUNCTION Solid Particle Run Time hrminisec 0 15 21
68. DIAMETER STD DEVIATION WEIGHTING Figure IOb Volume weighted Gaussian Analysis corresponding to Figure IOa and data of Figures 9a and b This behavior can be easily understood qualitatively by reviewing the relationship between particle size and scattering intensity discussed in the previous section For particle sizes sufficiently small to permit neglect of the Mie form factor arising from intraparticle interference of individual scattered waves we see from Equation Ib that the contribution to C t of a particular decaying exponential corresponding to a given diameter d should be weighted by the factor N V where N represents the number of particles having diameter di and Vi is their individual volume which is proportional to di Hence each diameter slice of the starting intensity weighted distribution has associated with it this factor of N Vi Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 31 T mir AM DLS THEORY Gaussian Analysis Solid Particles Chi Squared 1 1 Baseline adjust 0 01 Data 1047 0 K Mean Diffusion 3 21E 8 cm2 sec Fangen a Ferne gt Diameter 109 22 30 x NUMBER MEAN DIAMETER STD DEVIATION WEIGHTING Figure lOc Number weighted Gaussian Analysis corresponding to Figure lOa and data of Figure 9a and b The constituent diameters of the volume weighted distribution should each have the weighting factor N V i e the number of particles having a particu
69. E Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 52 ME Toot at DLS THEORY m Figure 14b Volume weighted Gaussian Analysis result for 261 nm latex standard See Figure 14a It is most important for one to appreciate the fact that reliable estimates of the widths of peaks in the particle size distribution CANNOT be obtained using the Distribution Analysis The peak widths obtained from this procedure depend on details of smoothing and basis vector coupling in the ILT algorithm as well as the operator s choice for the SMOOTHING parameter On the other hand it should now be clear that the Gaussian Analysis does indeed yield a reliable estimate of the peak width as in Figure 14b provided Chi Squared is low indicating a good fit A harder test for Distribution Analysis a bimodal distribution latex We now consider the first difficult test of the NICOMP ILT procedure analysis of a bimodal distribution Figure 15 shows the autocorrelation function after 23 minutes for a bimodal sample prepared using the 91 and 261 nm latex standards investigated earlier This test bimodal was made using a 3 1 ratio by mass or volume in favor of the 91 nm latex so as to enhance the relative scattering contribution of the smaller size component t more later Figure 16 shows a plot of the logarithm of the reduced autocorrelation data C t B vs t channel with a straight line drawn for reference purposes 1735954 au Cor
70. E een de 9 GAUSSIAN NICOMP ALL WEIGHTED u ana ee 10 AUTOGORRELATION FUNC TION en ae a ee 11 AUTOGORRELAHON DATA Zen es en een 12 TIME HISTORY PEOT ae sde 13 Nicomp 380 User Manual PSS 380Nicomp 030806 11 06 Page iii mir TQ i y TABLE OF CONTENTS CHANNEL ERROR PLOT ee sinne 14 APPENDIX B NICOMP PARTS LIST aa Taa Taaa aaa aaa ma Aa aaa Aaaa a diran 1 APPENDIX C NONAQUEOUS SOLVENTS FOR THE NICOMP ooocccccnocccccccocnccccnnnnnnccnnnnnnac cnn nennen nannte nnn 1 APPENDIX D SOLVENT TEMPERATURE VISCOSITY amp INDEX REFRACTION TABLE 1 APPENDIX E ESTIMATING MOLECULAR WElGHT ooocccccnnccccccccocaccccnnonaccnononanonononanacononnnna conc n nn nara nann cnn n naaa 1 Nicomp 380 User Manual PSS 380Nicomp 030806 11 06 Page iv TT Yi iT Ti qm t TABLE OF CONTENTS m LIST OF FIGURES Figure 1 Simplified block diagram NICOMP DLS Instrument sseseee 1 Figure 2 Simplified scattering model two diffusing particles uunssesssssssnsnnnnnnnnnennnnnnnnnnnnnnn 4 Figure 3 Typical intensity vs time for two diffusing particles ooonnoonocccnnnnnnnccnccccccnncnnnnnnnnnannnn 5 Figure 4 a b c Representative intensity vs time for small a medium b and large c size partici pm P EE 6 Figure 5 Computation of autocorrelation function C t see 8 Figure 6 Autocorrelation function C t for diffusion of uniform particles exponential
71. ERE FOREN EIERELR 12 Display Help for Current Task or Command sse 16 Start Measurement ssssssssssssssssseeeeneneeneeene RR RR essen sse eser ns enn sen sse n enne 16 Status Bales ee T C P 18 ice ML ERU TEE LT EM 18 SETUP docete a Bed e vcf Fee ve p e ev B ru cei Pe t Co fees 19 Select Setia POM EE 19 M ltAndglS OPONE at Ar eek eae ia 19 Interrupter Angle ta bU m IR iaa 20 Change Laser Wavelength coi ana 21 APD Overload Protection ocoocccccccccccnnncnnnconnnnnnnnnnnnnncnnnnnnnnnonnnonnnnnnnnnnnnnnnnnnnnonnnnnnnnnnnns 21 Intensity OvershobEb FabtOE eei ie cocotte ee 22 NICOMP Intens Wt Threshold esses nennen enemies 22 Enable Intensity MONITO scio eet A E cede y bie A 22 Dual Particle Sizing DLS Detector 2 2 keit 22 PARTICLE SIZING vis itcr ttt erc ne Cras Fede E cir o Po rtr rea de Fiera LP br deri d FR Tr s 23 Gontrol Mehl ohio aa a oir Seaton a a aai aeia 24 Nicomp Input Men il A Rn un EE 36 o PETEA FEE A E A ATT 36 Read Menu Fill caco cosa 2 tet A e EN GE EN iti 39 Save Mente TEILT 40 Change Graph CIO tit A AA AAA E a AAA AS 41 Control BUIN N eek te Serie ed cede Yr DER en es 42 Initialize ND Filter coo a Ri Se Ad nie 43 GorrsEOhcetlOn 4c 22055 E IE 45 GauSSianiivetiicii EE 46 NC 1 1 det ET A E 50 IGNI data A HR HIT 51 COM Data ct td nu Es 52 Channel EO ciutat tas ara ee OO ieee eee ene ied 53 A ES 54 parc RM E EN 56 E CET To1o a 0
72. LS THEORY There are 6 pieces of quantitative information contained in the summary display of the Gaussian Analysis Figure 0a Mean Diameter 226 1 nm Standard Deviation 30 Chi Squared 1 1 Baseline Adjust 0 01 Data 1047 0K Mean Diffusion 2 05 X 10 cm sec The value 30 for the standard deviation means that AR R 0 30 A critical piece of information contained in Figure lOa is the value of Chi Squared Any value close to or below 1 0 indicates an exceptionally good fit ofthe quadratic function to the reduced data Assuming that sufficient statistics have been collected in the autocorrelation function to make the value of Chi Squared meaningful which is NOT true early into a run a low value means that the Gaussian representation of the particle size distribution is a good assumption i e that no other distribution shape can offer a better fit to the data given the limitations implied by Poisson statistics Indeed the test of a good Gaussian Analysis result is whether Chi Squared remains small i e below 2 or 3 over the course of time as additional intensity values are collected and incorporated into autocorrelation function C t The DLS Module makes this judgment automatically and will provide an appropriate warning message if the value of Chi Squared exceeds 3 suggesting that the Gaussian Analysis result is inappropriate The value for Baseline Adjust shown above 0 01 is very close to the ideal value of zero
73. LT This rather sophisticated technique has also been used to analyze a variety of problems in other scientific areas unrelated to light scattering The specific mathematical procedure is a more sophisticated version of the least squares calculation used in the Gaussian cumulants Analysis it is referred to as nonlinear least squares NLLS analysis We can appreciate the problem which must be solved by the ILT technique by considering the most general expression for the autocorrelation function C t corresponding to an arbitrary distribution of particle sizes We recall for the trivial example of a single particle size that C t is simply an exponentially decaying function given by Equations 7 and 8 or 10a and b This is now replaced by a more general expression containing a weighted sum of individual exponentially decaying functions each of which corresponds to a different particle diameter C t AD f APD K 1 14 Here again the baseline of the autocorrelation function t o is given by B and constant A simply relates to the total amount of data acquired in channel 0 t 0 of C t such that C O A B In Equation 14 above we have assumed a discrete distribution of particle sizes containing M different diameters each labeled by index running from 1 to M The diffusion coefficient for each particle diameter is given by Di which is obtained from the radius R using the Stokes Einstein relation
74. MP DISTRIBUTION 100 Diam nm gt Bimodal 300 M ean Diameter 304 8 nm Fit Error 7 349 Residual 0 000 NICOMP SCALE PARAMETERS Min Diam 100 nm Plot Size 50 Smoothing 2 Plot Range 10 GAUSSIAN SUMMARY Mean Diameter 295 5 nm Variance P 0 033 Stnd Deviation 53 3 nm 18 096 Chi Squared 3 861 Norm Stnd Dev 0 180 Baseline Adj 0 000 Coeff of Var n Z Avg Diff Coeff 1 61E 008 cm2 s Run Time 0 Hr 6 Min 8 Sec W avelength 632 8 nm Count Rate 0 KHz Temperature 23 deg C Channel 1 z612 6 K Viscosity 0 933 cp Channel Width 35 0 uSec Index of Ref 1 333 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 5 APPENDIX A INTENSITY WEIGHTED NICOMP Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm 7 3 Data File C Program Files Particle Sizing Systems Nicomp 388 version 1 68 Bimodal 300 INTEN SITY Weighted NICOMP DISTRIBUTION Analysis Solid Particle NICOMP SUMMARY Peak 1 Mean Diam 230 8 nm S Dev 7 4 nm 3 22 Intens 42 91 96 Peak 2 Mean Diam 345 3 nm S Dev 13 8 nm 4 01 Intens 57 09 INTENS WT NICOMP DISTRIBUTION 100 Diam nm gt Bimodal 300 Mean Diameter 299 5 nm Fit Error 7 349 Residual 0 000 NICOMP SCALE PARAMETERS Min Diam 100 nm Plot Size 50 Smoothing 2 Plot Range 10 GAUSSIAN SUMMARY Mean Diameter 288 9 nm Variance P 1 0 033 Stnd Deviation 52 1 nm 18 0
75. Manual PSS 380Nicomp 030806 06 06 Page6 7 T Y Tl irn T An SAMPLE ANALYSIS RUN Interpretation of Data 1 Referencing data the following conditions should be met a Gaussian Int weighted Mean Diameter within 15 of stated diameter b Std Dev lt 15 0 c ChiSq lt 3 00 d Base Adj lt 0 05 2 The Nicomp is working within the manufacturer s specifications when all of these conditions are met 3 Starting with a fresh sample repeat measurement one time if any of these conditions are not met If on the second measurement any of these conditions are still not met then measure a second latex standard using the same procedures Important It may not be necessary to measure both latex standards If one works that means the instrument is operational Review of Completed Sample Results 1 Position the highlight bar over the File option and click once 2 Position the highlight bar over the Read option and click once The Read Data File Window will display 3 Position the highlight bar over the data file to be viewed 4 Click on OK A second pop up window displays prompting to re calculate the results 5 Press Y The distribution will display in Number wt mode 6 Press W to view alternate data forms Volume wt Number wt Intensity wt Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 6 8 ME Tum Pt SAMPLE ANALYSIS Rub Print Sample Results 1 Position the highlight bar over the Print icon and click o
76. NUMBER REL PERCENT 42 6 50 0 8 6 68 7 80 5 RESULTS Gaussian Analysis Number Weighting Mean Diameter 144 8 nm Standard Deviation 43 3 nm 29 9 x Ur DI Cl GJ OO PO A O DJ i e A Chi Squared 1 1 Baseline Adjust 0 01 l I I t I I i t f Mean Diffusion 3 20E 8 cm2 sec Data 1047 0 K Cumulative Results Percent Diam 25 104 0 127 8 156 9 RUN TIME O Hours 14 Mins 48 Secs AVG COUNT RATE 298 0 kHz CHANNEL WIDTH 21 0 USEC TEMPERATURE e3 DEGREES CENT VISCOSITY 0 933 CENTIPOISE INDEX OF REFRACTION COUNT RATE SETPOINT kHz ALPHA Molec Wt E 4 BETA Molec Wt Figure 11c Printout of number weighted Gaussian Analysis result for fat emulsion See Figure lOc Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 35 T mir AM DLS THEORY It MUST be re emphasized that the distribution shapes shown in Figure lDa b and c and Figure 11a b c are obtained from the tabulations for the Gaussian or normal curve found in standard mathematical references Their shape is an idealization and should NEVER be construed to offer more detailed information than can legitimately be expected from this fitting procedure The peak shown in Figure 11b for example contains 13 discrete diameter slices each evidently possessing a well defined height or relative volume In a way this is misleading this shape has emerged from what is fundamentally just a 2 parameter fit plus
77. O such breakpoint is evident in Figure 16 Indeed the curvature in the reduced data in Figure 16 qualitatively resembles that seen for the broad unimodal emulsion in Figure 9b Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 56 MM T T T DLS THEORY AAL Suppose we were to analyze this new system as we did the emulsion using the Gaussian Analysis From the substantial curvature evident in Figure 16 we would expect the unimodal generated by the Gaussian Analysis to possess a large standard deviation i e to be very broad This is indeed the case as seen in Figure 17 Gaussian Analysis Solid Particles Chi Squared 24 7 Baseline adjust 0 00 Data 1736 8 K Mean Diffusion 3 34E 8 cm2 sec I 1 i 1 i 1 n D D t D r 3 t 1 PA SR pr ER gt Diameter log 19 100 1000 CHI SQUARED LARGE USE DISTRIBUTION ANALYSIS 139 0 45 X VOLUME MEAN DIAMETER STD DEVIATION WEIGHTING Figure 17 The volume weighted Gaussian Analysis result corresponding to Figure 15 and Figure 16 The volume weighted mean diameter is 139 nm with a standard deviation of 4596 We also obtain the expected result that the volume weighted mean diameter is pulled down substantially below the intensity weighted value 198 nm from Figure 15 owing to the large standard deviation Figure 17 would have represented the state of the art of DL S based particle sizing technolo
78. PMT detector Technically the pulses which comprise the PMT photocurrent vary substantially in height as well as rate of Occurrence owing to the statistical nature of the secondary electron multiplication mechanism in the PMT 1 t t Figure 7 A typical photopulse sequence representing I t divided into intervals of equal time width t However a discriminator with a low reference level is used to convert this signal to a train of pulses of uniform height suitable for manipulation by standard integrated logic circuits in the autocorrelator The procedure for computing the digital representation of C t should now be conceptually clear The train of photopulses from the PMT detector is divided into intervals of equal time or channel width At Running sums of the products t I t t are then produced for many values of t 64 in the case of the DLS Module The separation times t are quantized in multiples of At At 2At 3At 64 At In addition a long delay baseline value is obtained t 64 1024 At Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 14 ME DLS THEORY m l Using the photopulse sequence shown in Figure 7 we now show how to compute the values of C t for the first few channels in t t AU C t 2 8 3 1 1 4 4 2 270 0 3 3 1 t 2At C t 2 1 3 4 1 2 440 273 0 1 t 3At C t 2 4 3 2 l 0 4 8 271 t AAt
79. Paper Orientation Size Letter 8 1 2x 11 inch v Portrait Source Auto E C Landscape ied Printer Default Printer The printer that is used for the majority of the printing when using the computer controller In some cases this may be a black and white printer Specific Printer Allows for the selection of another printer type such as a color printer for printing out color distributions Paper Size The paper sizes available depend on the model printer that is being used By clicking on the down arrow located to the right of the Size window a listing of the available paper sizes for the computer being used displays The default is set to Letter 8 1 2 x 11 since most printers accommodate for this size Source The default for this option is set to Portable Sheet Feeder however some printers have two paper trays for printing The source of the paper feed depends on the model of printer being used Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 10 TT AU T Orientation y NICOMP SOFTWARE AAA Portrait 8 1 2 x 11 in The data distributions will display vertically on the paper selected Landscape 11 x 8 1 2 in The data distributions will display horizontally on the paper selected Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 11 NICOMP SOFTWARE VIEw MENU Click on View in the Main Window in order to pull down the View Menu In the default condition both the Tool bar containing t
80. SS 380Nicomp 030806 06 06 Page 5 37 T ri Ts Toot at AM N NICOMP SOFTWARE There are two simple criteria which govern the selection of optimal values of MIN DIAM Range and Plot Size First it is always necessary to choose values of MIN DIAM and Range so that no substantial amount of the particle size population in Intensity weighting appears at either edge of the plot Ifthe MIN DIAM is chosen to be too large the small diameter end of the distribution will be pressed against the left hand edge of the diameter axis In this case the Fit Error will be seen to increase dramatically the more MIN DIAM exceeds the smallest size in the distribution Alternatively if the Range is chosen to be too small the large diameter end of the distribution will be pushed into the right hand edge of the diameter axis Here again the Fit Error may increase substantially In general it is desirable to choose some value of MIN DIAM such that there remains at least i e 5 or more empty bins at the left hand edge of the diameter axis In this way the resolution of the computed distribution would have to be maximized for a given value of Plot Size The correctness of choices of MIN DIAM and Range can be ascertained by examining the Intensity weighted plot in the Distribution Analysis What is of consequence is whether there is an appreciable intensity contribution i e more than 1 or 2 units compared to 100 units at the peak of the distribution
81. TION Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 12 TIME HISTORY PLOT Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm7 3 DataFile CProgramFilesiParticle Sizing SystemsWWicomp388Wersion 1 68 Bimodal 300 Diam nm VOLUME WT NICOMP CALCULATION HISTORY Runtime min gt Peakt Peak2 Peak3 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 13 APPENDIX A CHANNEL ERROR PLOT Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm 7 3 DataFile CProgramFilesiParticle Sizing Systems Nicomp388 wersion 1 68 Bimodal 300 Channel Error CHANNEL ERROR Channel Error unused channel 9033 6246 5804 1 743 1 327 1 941 3 184 4 178 8 336 6 225 7 592 9 761 10693 9 920 10504 6 083 4925 02359 2745 5339 6089 8260 9288 12 948 9818 13 999 12 240 11 856 11 329 8202 6330 2467 0663 3357 2823 6415 7 007 8 980 8 255 9 796 5510 amp 905 0817 1 869 0 193 2 908 2554 0715 5041 1416 Q088 8469 5676 7 466 2078 3 579 5309 2986 7 185 18 317 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 14 ME n LH APPENDIX BAM y PART DESCRIPTION PART NUMBER NICOMP PARTS LIST ASSEMBLIES PCB DSP Corr Assembly HV Control Board Pre Amp Board 380 Pulse Rate Board 380 Static Mixer Assembly Static Mixer Support Temperature Control Board Temperature Control Block 8 SS Terminal Block Assem
82. ZE 60 RANGE 100 A 6 Mins 48 Secs 9020 kHz 19 0 e3 0 9325 333 1000 10 USEC DEGREES CENT CENTIPOISE K COUNTS Figure 22a Printout of volume weighted Distribution Analysis result for the 3 1 91 261 bimodal sample after 7 min Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 70 DLS THEORY NICOMP Distribution Analysis Solid Particles SIZE nanometers log scale REL VOLUME 65 0 RESULTS Distribution Analysis Peak 1 1 4 100 Diameter 90 3 9 Volume 74 1 Peak 2 Diameter 270 0 Volume 25 9 Fit Error 1 6 Residual Mean Diameter 141 9 nm Standard Deviation nm I i I l l l i t l 1 l l i l i I l l SERSIBZW NSEIIEITZDZ Gaussian Analysis Volume Weighting Mean Diameter 127 7 nm Standard Deviation 59 1 nm Chi Squared 7 4 Baseline Adjust 0 01 Data 570 9 K Scale Parameters MIN DIAM 10 PLOT SIZE 60 SMOOTHING 3 RANGE 100 RUN TIME OQ Hours 39 Mins 46 Secs AVG COUNT RATE 287 5 kHz CHANNEL WIDTH 19 0 23 USEC DEGREES CENT eraser CENTIPOISE 1 333 000 OF PRINTOUTS 10 K COUNTS Figure 22b Printout of volume weighted Distribution Analysis result for the 3 1 91 261 bimodal sample after 10 min Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 71 DLS THEORY NICOMP Distribution Analysis Solid Particles SIZE nanometers log scale REL VOLUME
83. a clean single peak distribution For more complex distributions it is usually advisable to achieve a Fit Error below 1 5 or even approaching 1 0 to Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 50 rity Til iT jT DLS THEORY AAL obtain the most accurate reproducible results As a point of reference one can typically achieve a Fit Error of 0 1 or 0 2 after 8 to 12 hours of data collection Again this parameter only characterizes the stability of the resulting answer and NOT its appropriateness vis a vis the simpler Gaussian Analysis result In Figure 13 we find an ideal value for the Residual 0 0 We therefore conclude that there is a negligible concentration of aggregates or other large particle contaminants in the sample suspension In the event that the value for the Residual is significant e g 10 or larger one of two possibilities will occur with increased data acquisition 1 The Residual will simply remain high with little change in the composition of the size distribution plot 2 The Residual will fall to zero resulting either in a shift in the position s of the peak s usually to higher diameter s or in the appearance of a new peak at the high end of the diameter scale The latter explicitly reveals the large size component previously signaled in 1 by the large value of the Residual In the early stages of data acquisition these two possibilities frequently alternate in seemingly random fashion be
84. a A ui RIA OR BR A na DLR anes s ase Hinin MIRA TELE LODDO T HERI EEUU LET PME musti HANHI AAN ri itt td UULUDILUHLUUEU UM HUELLA DD APERUIT EEUU HL DA 1050000001 KLEE MILI HRLENLLI CIERRE ODA UU LITER DONO NE ELLE FERE HARE LEHRER LER MEI RII MH NUM UTR A PH VA RUNE mnm en he m y nr Areh AAA AA TEE A TA Lin A v ERRARE ULM DNGDDEDUODERGREDRONGUOUGEAGAOLADGABOANROREGULOEIEON UAC TLE ALA EDUC EAT HH ELLE TULERIT Unt HEEL HELLE LU LLL ELI LLL FLUE STAA PNREN ESUSSSESSEREEDO ROME ERODE MEGA LH LLL NE AG EN Stes scent saces ERRA AMARAS wanevas sevanonwas Pan os senon sonan Sannn annen sunsn va we MIA RARO DEEDE A 29998 9 CREA THES AAA AA sans E Laue AE Figure 16 Log C t B vs t for data of Figure 15 It is perhaps surprising to see that autocorrelation curve in Figure 15 does not the share of the Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 54 ME DLS THEORY m l appear to differ appreciably from that observed previously for the broad unimodal fat emulsion of Figure 9a Here we have particle sizes which differ by almost a diameter We therefore would expect to see evidence of a breakpoint or knee in the logarithmic plot of the reduced Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 55 RE hi DLS THEORY autocorrelation data Figure 16 given the admixture Equation 14 of two exponential functions whose decay time constants differ by a factor of three However N
85. a time sequence of volume weighted printouts is shown in Figure 22a d corresponding to run times of approximately 7 min 10 min 42 min and 8 hours 10 min Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 67 po e see Mi I n E 3 RE LN kt iw EEE DAS A ree Cite er ie IDEAS NONAS poer man res er FATSA pese eunt MT FIN ESERE FAR E E E RON ad E EHRE PETES GUHSELERENEREEL 90004 RESTA ACTA LEIOA URSUA EEREU LAGEN DOES DEERE COON 00 decl decies IDAS BEER CLES ose PROA HEEL DER GA LUN HH DTGA CANON GB FOTOS WA TORA Ano SI DAUEN RAC HI VAATE PAAA OU PB PRADA SM CONST ERNO NONE EDEL 00 LI OS DEREN E CLOS ER ESEEN EA BAITEDEN ES RDA HUSO NAO SU ALEDO OOG SOT EC FU LAESA E LE IM EB EEE BE a ER FG ERBEN CH o SETE ll SUE ERBE DOS EIERN HEUER LEE OS HERE FAM US E FORERO PETRA DOGS HEISE L1 Hl fetes At EE ACI ERR E PT AAA do E PE Ad de tes di ASEO DEA ALA O FEE RE UO RET LR LL uL LU rail Uu nu i LL Figure 20 Log C t B vs t for a widely separated bimodal latex sample 3 1 vol ratio 91 and 1091 nm Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 68 ME T T T DLS THEORY AAL NICOMP Distribution Analysis Solid Particles 1 Fit Error 2 50 Residual 0 0 Gaussian Chi Squared 12 0 Data 341 8 K gt Diameter log VOLUME Figure 21 The volume weighted Distribution Analysis res
86. adjust Data 916 4 K Mean Diffusion 1 73E 8 cm2 sec L L t Li t Li t L y t L 1 3 t t M LI L I t 4 i a AA aA 26 100 1000 268 4 7X VOLUME MEAN DIAMETER STO DEVIATION WEIGHTING Figure 13 Volume weighted Distribution Analysis result for 91 nm latex standard to 90 0 nm for the Gaussian Analysis result This discrepancy only 1 is excellent considering the fact that two radically different mathematical procedures were used to obtain the two results It was pointed out earlier that the true diameter of this standard lies closer to 88 89 nm rather than the nominal 91 nm value In Figure 13 we see that the Fit Error for this analysis fell to 4 60 with 480K of Data Ch 1 This value should continue to decrease with increasing data acquisition It is IMPORTANT to appreciate that the Fit Error only provides an indication of how relatively stable or settled the Distribution Analysis results should be but NOT on whether it is the preferred result compared to the Gaussian Analysis This judgment of course is provided by the value of Chi Squared As the warning message clearly states in this case the Gaussian Analysis must be used One will typically find that narrow ideal distributions like this one will settle relatively quickly i e with values of Fit Error of 3 or 4 or higher However in general one should run long enough to achieve a value below 2 0 in the case of
87. aks 71 7 and 28 3 are very close to the actual values of 75 and 25 for a 3 1 volume ratio NICOMP Distribution Analysis Solid Particles 1 Fit Error 2 88 Residual 0 0 Gaussian Chi Squared 5 9 Data 346 6 K i 1 1 1 1 1 t n D 1 y r D D 3 t 4 4 i 1 1 1 1 1 1 D t gt Diameter log 1000 Diam nm VOLUME Figure 18a The volume weighted Distribution Analysis result for the 3 1 91 261 nm test bimodal after Data 347K The distribution obtained after more than 10 minutes of running Data 840K is shown in Figure 18b The peak locations and relative volume percentages have changed somewhat relative to the results of Figure 18a By this time the Gaussian Chi Squared has risen further to 8 2 and the Fit Error has fallen to 1 84 The non zero value of the Residual suggests that additional data acquisition would be desirable Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 58 ME T T T DLS THEORY AAL NICOMP Distribution Analysis Solid Particles 1 Fit Error 1 84 Residual 5 0 Gaussian Chi Squared 8 2 Data 840 0 K gt Diameter log 000 VOLUME Figure 18b The volume weighted Distribution Analysis result for the test bimodal after Data 840K 10 mm Figure 18c shows the volume weighted Distribution Analysis result for the same test bimodal after 23 minutes elapsed time Now the Fit Error has fallen close to 1 0 and the Resid
88. ample is twice that of the former We can acquire a better appreciation of the subtlety of the analysis task which faces us by looking at the reduced data C t B on a semilog plot in Figure 9b and comparing this with the previous reduced data for the narrow 91 nm latex standard Figure 8c Indeed we must look closely to find any qualitative difference between the two plots However on closer examination we see that the reduced data points in Figure 9b possess a slight curvature a displaced straight line has been drawn to enable this curvature to be seen As we shall see in a moment this fat emulsion sample does indeed possess a substantial width or range of sizes in its particle size distribution Therefore what may seem surprising and perhaps intimidating is how relatively little deviation there is from the single exponential behavior of C t in Figure 9b given the apparently large qualitative difference in distribution shapes between the sharp latex standard and the broad emulsion sample Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 22 ME DLS THEORY m l The example above illustrates the inherent difficulty which all DLS based particle sizing instruments face what distinguishes the shape of one computed particle size distribution from another is often a relatively subtle deviation of C t from single exponential behavior Hence we MUST learn to appreciate the importance of acquiring data of high st
89. ansform In the proceeding section we saw that the Gaussian Analysis has the useful attribute that the resulting distribution settles quickly to a stable reproducible result with increasing acquisition of light scattering data As was pointed out this is due to the fact that the least squares quadratic cumulants fit to the reduced data is essentially just a two parameter fit apart from a possible baseline adjustment Obviously this smooth fit to the data is relatively insensitive to subtle changes in the autocorrelation constitutes the good news Analysis function This insensitivity associated with the Gaussian Analysis The bad news of course is that this method is inherently limited to describing only simple particle size distributions symmetric single peak unimodal populations More complex distributions such as highly skewed unimodals or bimodals are completely misinterpreted by the Gaussian Analysis the degree of error will be either quantitative or qualitative depending on the precise shape of the true distribution The only indication of the existence of a more complicated distribution is the occurrence of a large and rising value of Chi Squared as described previously This warning flag informs the operator in unambiguous terms that the Gaussian Analysis result is inappropriate In this case another approach for analyzing the data is clearly needed The NICOMP Distribution Analysis provides the need
90. ars the contents of the autocorrelator channels This is part of the initialization process For all subsequent runs it is necessary to clear the data whenever the Channel Width is changed in the input menu or the sample is changed This option may also be used to restart the autocorrelator after the sample temperature has settled to its stable set point or whenever a new sample has been introduced into the scattering cell This option may only be used in display mode ALT C Clock The internal clock of the computer controller can be viewed when this option is accessed The clock window can be removed by pressing the same keys used to access the time D Display autocorrelator data Used to view the block of raw data collected from a sample run when in display mode Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 61 mir That at AM i NICOMP SOFTWARE E Edit Caption Used to enter or edit a caption for a printout Follow these steps to enter the caption a Press E b Type in the desired caption A maximum string of eighty alpha numeric characters is allowed C Press ENTER The string of characters entered will be saved Ctrl E Channel Error Used to compare the statistical discrepancy between the actual data and the ability to curve fit the data F Display Gaussian or Nicomp Distributions Returns to the Gaussian Analysis Display Will also display a list of all of the data files which have previously
91. ash of the hard drive Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 4 1 NICOMP SOFTWARE FILE The following provides a brief summary of the File options that are used during the use of the Windows CW388 Software Access of the File options 1 Position the highlight over the FILE option and click once using the mouse The following window of File options displays CW388 Version 1 68 NICOMP Particle Sizing Systems 8 131 Fer oip press t Ti start 7 mn Seen 2 Position the highlight bar over the desired selection and click once Following is a description of each of the options offered Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 1 NICOMP SOFTWARE Restore Ctri R Read F6 Read New SHF F6 Print Setup Exit Restore Used to restore collecting data once the autocorrelator has been halted Click on Yes the screen will clear and data will be collected No the warning screen will disappear and the autocorrelator will not start taking data Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 2 NICOMP SOFTWARE Read A data file that has been stored following a measurement can be retrieved to display the resulting particle size distribution PSD with the desired weighting When this option is selected a list of data files will display in the Read Data File window such as in the example below Read Data File RR Look in Nicomp 3
92. at either edge of the size scale If this is the case either the MIN DIAM must be decreased or the Range increased or some combination of the two Otherwise two things will happen the Fit Error will rise and most important the remaining portions of the size distribution histogram will become increasingly distorted and unreliable It is vital to understand that the choices of MIN DIAM Range Plot Size and Smoothing interact in a complicated way in the calculation If the rules governing intensity contributions at the edges of the size scale are ignored the choices for these parameter values may significantly and adversely affect the computed particle size distribution results The second criterion relates to the optimal selection of the Plot Size One would like of course to maintain the highest possible resolution of the size histogram plot at all times by setting the Plot Size equal to 60 However it turns out in practice that in the case of broad highly polydisperse distributions whether unimodal or bimodal this choice may result in over resolution leading to an unstable fit This condition could be recognized by the fact that the distribution breaks up leaving a number of holes where there is no apparent volume surrounded on both sides by large relative amounts of particle volume For example broad unimodal populations can be transformed into false bimodals given too large a Plot Size and or too low a smoothing value for
93. atistical accuracy Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 23 A AAA 30000 ERES 00900 MEET MOMS IERI ITEDUEINOS DUDAS LADOS DENOU VI BD 10000 090000 caste 0000 Ll DOBA BEDAE PASA DEOD E EM ndo RENA E rags ate rem EPEHESIT HESS mr LU if inated in alba DRE e RATE SADA pep LA AT A HN HI i MT NIHIL TUI E ALT MIT Tae E PEN SERERE rt nim RE A ERERGUBBEL 15203 MOGDO ESOO LEROS 10590 00048 LEERE MIN A OT unt ELATI ER PLUME AME ren ILL CIL SL EL en u REN Pe ud TRI MERE RU EE IH HH REA GT GG A ROD EEL MM UBL NM LN ttem emp ESERSCSERAER I HRLRABE ETE ELA BAAGA EHE RERO RR TERT GERRA AGE ERI AAA Ed S9 EDEN SUE 1900419095 3034814853 i ese a LL TH east HERE ETHER ERA HA RR RT IT At AL PB D LLL LIII RR ILIA HER ipM ANHI EHE BR RHEIN RT UL HH FITLIFTERSELTITTLITIITELET EN reee HH Hu mio NHIEU EHLEPELEHLHLH HEU LE LUE LU HEU LUE LI ORAE EEUU CLANS ESHPS LEER EDTSICSESUESOLSUSEEINSHOLGAODOEQUUSNONSI INES RIEN EHE UL AENA ODO OIL HMM IHE ELLE REL AR ha este etree ee en eee eee ee rbd ALA Lid HATE qu both LLLI ELO MITA HILL ILL ILL IL xri i2 LE TE RE IEREISRBEGELGLISEESDSRDESSUEITERTIHUHHNSNNSE E 117 7 COTO cen A torefamesssaatsed ee rds Sed REECE I LUE EISE eet MEE Figure 9b Log C t B vs t for data of Figure 9a This requirement is intimately related to the run time and the efficacy of sample preparation Not surprisingly it turns out that there are NO short cuts to obtaining good raw data a
94. bly CABLES DB 25 Pin to 9 Pin 4 Cable PSS FANS Fan 12V 4 6 4710NL o4W B20 NMB FILTERS 0 2 um Millipore Hydrophobic filter KTGRO4NP3 Neutral Density Filter Only available for HPL and VHPL lasers FUSES Fuse fast 3A 312003 Littlefuse Fuse Holder 342014A Littlefuse Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page B 1 RE v APPENDIX B PART DESCRIPTION INJECTION PORT VALVE Hamilton Valve 86907 HVX 2 Fitting Leured 35071 Fitting Barber 35072 Nut Valve 35121 LASER Laser 5 mW 5mW High Power Laser 35mW Very High Laser Air cooled 75mW POWER SUPPLIES PMT Power Supply Power Cord 115V Power Supply PUMPS 380 Pump Assembly 60 rpm 115V Masterflex Pump Head MasterFlex SAMPLE CELL Flow thru Cell Assembly Sample Cell Holder Flow Sample Cell Bottom Plate Sample Cell Block M A Sample Cell Back Plate Glass pieces Autodilutor TUBING SIL Perox 96400 16 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page B 2 PART NUMBER Hamilton Hamilton Hamilton Hamilton Uniphase Melles Griot UniPhase PMT 20CN 3 Bertan 174075 Belden MAP80 4DO2 PowerOne T 7543 60 T 7016 20 121 114 OS Hellma Cole Parmer LE MEA APPENDIX N t NONAQUEOUS SOLVENTS FOR THE NICOMP The Nicomp can be used to measure the size of particles that are suspended in solvents other than pure water For example certain dry powders such as ceramic compounds are best dispersed
95. bution 80 of distribution Run Time Count Rate Channel 1 Channel Width Nicomp 380 Manual APPENDIX Particle Sizing Systems Inc Santa Barbara Calif USA 295 5 nm 53 3 nm 18 0 0 180 VOLUME Weighted GAUSSIAN DISTRIBUTION Analysis Solid Particle Variance P Chi Squared Baseline Adj Z Avg Diff Coeff VOLUME WT GAU SSIAN DISTRIBUTION 257 3nm 290 6 nm 328 2 nm 366 1 nm 442 0nm 338 2nm A A A AAA 0 Hr 6 Min 8 Sec 0 KHz 612 6 K 35 0 uSec W avelength Temperature Viscosity Index of Ref Data File C Program Files Particle Sizing Systems Nicomp 388 version 1 68 Bimodal 300 0 033 3 861 0 000 1 61E 008 cm2 s 632 8 nm 23 deg C 0 933 cp 1 333 PSS 380Nicomp 030806 06 06 Page A 1 APPENDIX A INTENSITY WEIGHTED GAUSSIAN Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm 7 3 Data File C Program Files Particle Sizing Systems Nicomp 388Wwersion 1 68 Bimodal 300 INTEN SITY Weighted GAU SSIAN DISTRIBUTION Analysis Solid Particle GAU SSIAN SUMMARY Mean Diameter 288 9 nm Variance P 0 033 Stnd Deviation 52 1 nm 18 0 Chi Squared 3 861 Norm Stnd Dev 0 180 Baseline Adj 0 000 Coeff of Var n Z Avg Diff Coeff 1 61E 008 cm2 s INTENS WT GAUSSIAN DISTRIBUTION 20 Diam nm gt Bimodal 300 Cumulative Result 25 of distribution lt 251 6 nm 50 of distributi
96. cause of inadequate statistical accuracy in the autocorrelation function Obviously it is desirable to achieve result 2 above a zero Residual One can often hasten the transition from 1 to 2 by increasing the maximum value of the diameter scale equal to MIN DIAM RANGE This is accomplished by increasing the value of RANGE e g to 500 or 1000 if necessary Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 51 DLS THEORY In Figure 14a we show a typical result obtained using Distribution Analysis for a larger size polystyrene latex standard 261 nm Here we obtain a volume weighted mean diameter of 269 9 nm which agrees very well with the value found using Gaussian Analysis 268 4 nm shown in Figure 14b with the same Data NICOMP Distribution Analysis Solid Particles 1 Fit Error 3 55 Residual 2 4 Gaussian Chi Squared 0 2 Data 916 4 K 4 4 1 4 H i t 1 t t t n t 1 1 t 1 ET RD AA Na e gt Diameter log 20 100 1000 CHI SQUARED SMALL USE GAUSSIAN ANALYSIS Peak 1 t Diam nm 269 9 i VOLUME 100 0 Figure 14a Volume weighted Distribution Analysis result for 261 nm latex standard Gaussian Analysis Solid Particles chi Squared 0 2 Baseline adjust 0 14 Data 916 4 K Mean Diffusion 1 73E 8 cm2 sec A eae DD 4 Diameter log 1000 7 VOLUME y MEAN DIAMETER STD DEVIATION WEIGHT ING
97. ctions in Equation 14 exp DiK2t constitute the starting basis vectors for the NICOMP ILT algorithm The latter systematically varies the weighting coefficients fi in Equation 14 in order to determine the sensitivity of the resulting computed fit to C t to each coefficient In this way many if not most of the coefficients are set equal to zero The non zero values of fi which survive constitute the solution to Equation 14 As additional data are acquired in C t this solution will change either a little or a lot depending on the current statistical accuracy of C t There is an additional observation concerning the ILT fitting procedure and Equation 14 that is VERY IMPORTANT The weighting coefficients f represent the relative scattering intensities contributed by particles of size type i That is the individual decaying exponential functions corresponding to each particle diffusivity D are added together in Equation 14 with weighting factors fi each of which is proportional to the total scattered intensity produced by all of the particles of that size From the discussions in the previous sections we have fi Ni V GY 16 where N is the total number of particles of size type i and V is the particle volume giving rise to scattering In the case of a solid particle V di for a thin walled vesicle Vi di See the following section The extra factor for the additional effects of intra particl
98. d If a series of related samples of known molecular weight is not available consult the physical chemical literature to locate values of and for the particle solvent system under investigation Values for a few representative polymer solvent systems are listed below POLYMER SOLVENT T C oc B Poly Acrylamide Water 20 8 46 x 10 0 69 Poly acrylonitrile DMF 25 32 0 63 DMF 35 2 19 0 58 poly vinyl acetate MEK 20 78 0 63 poly styrene MEK 25 Sch 0 53 poly isoprene Chloroform 20 35 0 42 poly methyl Ethyl Acetate 20 1 60 0 48 methacrylate poly acronitrile poly vinyl alcohol Water 20 5 5 x 10 0 68 poly 1 hexane Acetone 20 3 0 x 10 sulfone Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page E 3
99. diffusion of the particles there is a well defined characteristic lifetime of the fluctuations which is inversely proportional to the particle diffusivity We compute the autocorrelation function of the fluctuating intensity obtaining a decaying exponential curve in time From the decay time constant t we obtain the particle diffusivity D Using the Stokes Einstein relation Equation 2 we finally compute the particle radius R assuming a sphere Photon counting and digital autocorrelation functions We now consider the practical application of the theory discussed above in an actual DLS particle sizing instrument The first step is computation of the autocorrelation function C t from the scattered light intensity I t as prescribed in Equation 3 It should be apparent that the fundamental operation of multiplication is most easily accomplished if both t and I t t are expressed as digital quantities Fortunately it turns out that this is already the case In our discussion thus far we have represented as an analog signal which varies continuously in magnitude as a function of time e g Figure 3 and 4a b and c However in reality this is not correct The scattering signal actually consists of a series of individual photopulses produced by the PMT detector Figure 1 That is the particle suspension is sufficiently dilute that the average scattering intensity at the PMT photocathode is extremely low resulting in a photocurr
100. display Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 57 NICOMP SOFTWARE Time Plot Scale Is used to manipulate the Time Plot to zoom in out on collected data The Time History Plot must be displayed in order to use this option Change Scale Minimum Diameter 1186 Maximum Diameter 1339 Starting Time Ending Time v Auto_scale Minimum Diameter Maximum Diameter Enter the minimum and maximum diameter to isolate the range for which the time history plot is to display Starting Time Ending Time Enter the start and end time for the range that is being isolated The time history plot s scale is then updated to reflect the information entered Auto_scale Automatically sets the min max of the time plot scale based on the value of the graph Show Intensity Shows the intensity weighted values of the time plot Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 58 TE Tht at NICOMP SOFTWARE m Please refer to the Display section of this manual for examples of these distributions WEIGHTING INTENSITY VOLUME NUMBER INTENS VOL Intensity Displays the relative intensity of scattered light vs diameter for a sample run Volume Displays the relative particle volume vs diameter Number Displays the relative number of particles in a sample run vs diameter Intens Vol Displays the overlays of the intensity and volume weighted sample runs Nicomp 380 Manual PSS 38
101. e G accounts interference important when the particle diameter is no longer negligible compared to the laser wavelength Mie region In the small diameter Rayleigh region diameters smaller than about 50 nm G is essentially unity for all particle shapes and sizes and can be ignored in Equation 16 We have explicitly written the expression for the weighting coefficients f in Equation 16 to help motivate Equation 14 which is the basic starting point for Distribution Analysis We must Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 48 rity Til iT jT DLS THEORY AAL remember of course that the coefficients f are the unknowns they are computed by inverting Equation 14 The answer produced by Distribution Analysis is the intensity weighted particle size distribution a plot of f vs d diameter The d scale is determined by the choices of MIN DIAM PLOT SIZE and RANGE as discussed earlier The volume and number weighted distributions are obtained from the intensity weighted plot by dividing each value of f by V G and V G respectively for each diameter slice It is for this reason that Equation 16 is needed to permit the various weightings to be obtained from the raw intensity weighted distribution of fi values which emerges from the ILT algorithm To conclude this Section remember that the actual plots obtained from the Distribution Analysis represent very simplified versions of whatever the
102. e distribution of diffusion coefficients D is approximately equal to a Gaussian or normal shape This is a bell shaped distribution that requires only two parameters for its full description not including the peak magnitude which is arbitrarily set equal to 100 for Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 25 T mir AM DLS THEORY all of our relative particle size distributions These are the mean diffusivity D and the half width AD of diffusivity values Strictly speaking the cumulants fit results in an approximately Gaussian distribution of intensity weighted diffusivities This point will be discussed later The connection between coefficients a and a obtained from the best quadratic fit to the logarithm of the reduced data and the parameters D and AD of the Gaussian distribution of diffusivities is given by a DK 12a or D a K 12b and AD D 2a a 13 Equation 13 gives the normalized standard deviation or coefficient of variation of the diffusivity distribution i e standard deviation AD divided by the mean diffusivity D Naturally the width parameter AD is related to coefficient a2 which describes the extent of curvature in the reduced data For distributions that are very narrow nearly monodisperse we expect az to be very small so that the quadratic function effectively reduces to being a straight line Ultimately one wishes to have the result expressed in terms of a dist
103. e distribution of particles suspended in a solvent usually water The useful size range for the DLS technique is quite large from below 5 am 0 005 micron to several microns The power of the technology is most apparent when applied to the difficult Particularly for diameters below 300 nm submicron size range where most competing measurement techniques lose their effectiveness or fail altogether Consequently DLS based sizing instruments have been used extensively to characterize a wide range of particulate systems including synthetic polymers e g latexes PVCs etc oil in water and water in oil emulsions vesicles micelles biological macromolecules pigments dyes silicas metallic sols ceramics and numerous other colloidal suspensions and dispersions PRINCIPLES OF DLS A QUALITATIVE REVIEW Classical light scattering intensity vs volume A simplified schematic diagram of the DLS module is shown below Light from a laser is focused into a glass tube containing a dilute suspension of particles The temperature of this scattering cell is held constant for reasons which will soon become apparent Each of the particles illuminated by the incident laser beam scatters light in all directions The intensity of light scattered by a single isolated particle depends on its molecular weight and overall size and shape and also on the difference in refractive indices of the particle and the surrounding solvent The incident light wave can b
104. e square of the average of the values because of the existence of correlations This is referred to as the baseline of the autocorrelation function In practice it can be effectively determined by evaluating Equation 3 using a sufficiently large value for t Hence we can say with certainty that the function C t for our situation of diffusing particles must fall from the value lt I t gt at t 0 to the baseline value lt l t gt at very large t The problem remains what is the shape of C t between these two extreme values Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 10 DLS THEORY Ideal case uniform particle size It turns out that for random diffusion of non interacting particles the autocorrelation function C t of the fluctuating scattered light intensity I t is an exponentially decaying function of time t as shown symbolically in Figure 6 This is described by the expression C t A exp t 1 B 7 where A lt I t gt lt I t gt and B lt l t gt counts X1000 AUTOCORRELATION FUNCTION 310 300 290 280 270 260 250 240 230 350 550 Channel Width 12 0 uSec Figure 6 Autocorrelation function C t for diffusion of uniform particles exponential decay Variable t is the characteristic decay time constant of the exponential function t characterizes quantitatively the speed with which the autocorrelation function C t decays toward the long t limi
105. e stepper motor arm provided the cell is highly cylindrical and well aligned i e centered on the shaft of the stepper motor Multi Angle Model 170 Designed to be used with the Nicomp 170 Computing Autocorrelator Any value for the actual scattering angle independent of the type of scattering cell used may be entered Interrupter Angle The interrupter angle parameter is only used for the Nicomp units which include the multi angle option The interrupter angle is the reference angle for the moveable arm on the stepper motor which carries the pinhole optical fiber receiver When power is first applied to the Nicomp the internal computer causes the stepper motor arm to rotate until it intersects an optical interrupter The latter defines the reference point or interrupter angle which is approximately 122 4 degrees with respect to the forward direction of the laser beam which defines the zero angle The stepper motor than advances in the opposite direction at 0 9 degrees step until the arm reaches 90 degrees in angle The number of steps depends on the value of the interrupter angle Any subsequent changes in angle are made from th 90 degree resting angle If the moveable arm becomes misaligned the resting angle may differ from 90 degrees The resulting error can easily be eliminated by resetting the value of the interrupter angle If the resting angle is too small e g 88 degrees the interrupter angle must be decreased by the ap
106. e thought of as consisting of a very rapidly oscillating electric field of amplitude E frequency approx 10 Hz Detector Diode Deconvolution Autocorrelator Figure 1 Simplified block diagram NICOMP DLS Instrument Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 1 T mir AM DLS THEORY The arrival of this alternating field in the vicinity of a particle causes all of the electrons which are free to be influenced the so called polarizable electrons to oscillate at the same frequency These oscillating electrons in turn give rise to a new oscillating electric field which radiates in all directions the scattered light wave The quantity of interest in a scattering measurement is the intensity of the scattered wave ls rather than its amplitude Es The intensity is given simply by the square of the amplitude I Es The dependence of the scattered light intensity lg on the molecular weight MV or volume V of the particle is particularly simple when the particle diameter is much smaller than the laser wavelength the so called Rayleigh region In this case all of the polarizable electrons within a particle oscillate together in phase because at any given time they all experience the same incident electric field Hence the scattered wave amplitude E is simply proportional to the number of polarizable electrons times the incident wave amplitude E The former quantity is essentially proportional t
107. e user should be warned that this analysis is a difficult one Failure to disperse adequately the latex particles or to achieve a dirt free suspension will in general cause the quality of the Distribution Analysis results to deteriorate markedly Most of the commercially available DLS particle sizing instruments find it more difficult to achieve results on a routine basis like those seen in Figures 18a d Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 61 T mir Toot ait AM N DLS THEORY The number weighted distribution corresponding to the above plots is shown in Figure 18e It is obtained from the volume weighted result as described earlier using Equation 16 NICOMP Distribution Analysis Solid Particles 1 Fit Error 1 19 Residual 0 0 Gaussian Chi Squared 24 7 Data 1735 8 K a 3 a 4 a a L s a e e e a J a 4 a i n e a H e gt Diameter log 000 Figure 18e The number weighted Distribution Analysis result for the test bimodal corresponding to Figures 18c d see in Figure 18e that the mean peak diameters are very close to the nominally correct values for this simplest of weightings However we can also appreciate why this choice of weighting is often avoided and volume weighting chosen in its place For any highly polydisperse distribution such as the present one there is usually too large a disparity between the numbers of smallest and largest par
108. ed alternative method for analyzing the autocorrelation function In contrast to the Gaussian Analysis it makes NO a priori assumption of the shape of the final distribution Rather than relying on only two parameters plus possibly a baseline adjustment the Distribution Analysis typically yields as many as three or even four independent parameters as well as a baseline adjustment Because the amount of information i e the number of parameters potentially provided by the Distribution Analysis exceeds that yielded by the Gaussian Analysis considerably more data is needed in general in the autocorrelation function to obtain a reliable result This is simply a requirement of better statistical accuracy or signal to noise ratio in the autocorrelation function In this game which is inherently mathematical a simple rule applies you cannot obtain something for nothing Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 44 rity Til iT jT DLS THEORY AA Obtaining more information relating to a more complex distribution requires that more time be expended acquiring more scattered intensity data While this same rule was implicit in our discussion of the settling of the Gaussian Analysis results we shall see that it is much more critical in the case of the Distribution Analysis The general mathematical procedure that is utilized in the proprietary NICOMP Distribution Analysis is referred to as inversion of the Laplace transform I
109. ent which consists of discrete pulses separated by zero baseline current corresponding to individual photons which comprise the weak scattering signal Hence the DLS instrument is said to operate in the photon counting regime Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 12 ME Poma DLS THEORY m If I consists of a train of discrete pulses rather than an analog signal what is the quantitative meaning of the intensity at time t Clearly the intensity must be represented by the number of photopulses per unit of time the larger the number of pulses occurring in that time unit the larger the intensity For example in typical operation the DLS Module might show a photopulse rate of say 300 kHz This value is updated every one second and represents the number of photopulses detected in the proceeding one second interval The sequence of values might resemble the series 302 297 299 304 296 etc We would therefore say that the average intensity is approximately 300 000 meaning pulses per one second interval However it would be equally valid to express the average intensity as 150 000 meaning per 0 5 second interval or as 30 000 meaning per 0 1 second interval That is any unit of time is as valid as any other for the purpose of defining the average value of the scattered intensity depending on the length of time which one wishes to use to define that average value Earlier we saw that it is typicall
110. er a data file is to be saved To access this option 1 Position the highlight bar over the File option and click the mouse once 2 Position the highlight bar over the Save As option and click the mouse once The following screen will display Save Data File Ae Save in Co Nicomp 388 3j ek Edy E version 1 68 E Bimodal 300 Filename E Save astype v Cancel Save ASCII Use this option to save the data collected for a particular sample to an ASCII file format The data can then be imported to a spreadsheet program for presentation To create new files in standard ASCII format to export data files into other software packages e g spreadsheets for manipulation of the original data follow these steps 1 Select a data file 2 Position the highlight bar over the File option and click once 3 Position the highlight bar over the Save ASCII File option using the mouse and click once the following window displays Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 5 NICOMP SOFTWARE Save ASCII File E Save in Nicomp 388 x amp ex E3 version 1 68 ferme ESO Save as type t asc 7 Cancel 1 Type in the desired file name A new file in ASCII format will be created and stored in the Data Directory with the file extension asc If the same file name already exists the following message will appear 2 Click on Yes the existing ASCII file having the same fi
111. falls to approximately the Intensity Setpoint which is set in the Conrol Menu the pump in the Nicomp will halt thereby stopping the flow of fresh diluent The Intensity Overshoot Factor compensates for the proper stopping of the pump NICOMP Intens Wt Threshold Sizing is not performed until this specified level of intensity is achieved Enable Intensity Monitor Provides intensity as a function of time Dual Particle Sizing DLS Detector PMT Output standard APD Output High gain detector offers 7 time the gain of a standard PMT for sizing small nanoparticles 0 1 10 nm or low concentration colloidal solutions Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 22 NICOMP SOFTWARE PARTICLE SIZING The following provides a brief summary of the Particle Sizing options that are used during the use of the Nicomp Software Access of the Particle Sizing options 1 Position the highlight over the Particle Sizing option and click once using the mouse The following window displays Control Menu F3 AutoPrint SaveMenu F2 NICOMP Input Menu I Read Menu File F7 Save Menu File Ctri F7 Control Buttons F8 Change Graph Color Alt Ctrl B Initialize ND Filter Alt Ctrl I 2 Position the highlight bar over the desired selection and click once Following is a description of each of the options offered Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 23 NICOMP SOFTWARE Control Menu The Con
112. fat emulsion See Figure IDC 2 2e 35 Figure 12a Intensity weighted Gaussian Analysis uuusssesssssnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn nenn 40 Figure 12b Volume weighted Gaussian Analysis oooccococococcccccccconanaoaanncncnnnnnnnnnnnnn cnn e 41 Figure 12c Intensity weighted Gaussian Analysis sssssesee eem 41 Figure 13 Volume weighted Distribution Analysis result for 91 nm latex standard 50 Figure 14a Volume weighted Distribution Analysis result for 261 nm latex standard 52 Figure 14b Volume weighted Gaussian Analysis result for 261 nm latex standard 53 Ke AS UIC LISMM EE 53 Figure 15 Autocorrelation function for a test bimodal 3 1 vol ratio 52 91 and 261 nm latex particles 93 Figure 16 Loge C t B vs for data of Figure UD ooconniocococcccnnonannnnnnnnencnnerennnnnnnnernnnnnnnncnnnnanaos 54 Figure 17 The volume weighted Gaussian Analysis result corresponding to 57 to Figure 15 and Figure 16 d ringe 57 Figure 18a The volume weighted Distribution Analysis result for the 3 1 91 261 nm 58 test bimodal after Data 347K eee tate ade teat head te ee ated Enuunan een 58 Figure 18b The volume weighted Distribution Analysis result for the test bimodal 59 after Dette 840K 10 Mm ae d 59 Figure 18c The volume weighted Distribution Analysis result for the
113. g sum of many products I t I t t all having the same separation in time t for many different values of t The ability of C t to extract useful information from the fluctuating scattering intensity I t can best be understood by considering a portion of a typical signal l t shown in Figure 5 We arbitrarily choose a particular time t and record the value of ls at that time I t We next consider a very small value of t equal to t and evaluate at this slightly earlier time t t t t Because t is presumed to be small t t must be very similar to I t The reason for this of course is that the particles have not been able to change their positions significantly i e compared to X under diffusion in the presumed short time interval t In Figure 5 I t t is shown to be slightly larger than I t 1 1 43 I t t3 1 t t 1 4 Figure 5 Computation of autocorrelation function C t Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 8 rity Til iT jT DLS THEORY MAAL However if t had been chosen differently in Figure 5 the order of the two values might have been reversed In any case what matters is that the two intensity values that become multiplied in Equation 3 are nearly the same They are said to be highly correlated Clearly the choice of t is irrelevant for any value of t I t and I t t must be highly correlated i e nearly the same for a
114. gy only 10 years ago prior to the commercial introduction of ILT algorithms for analysis of highly polydisperse distributions Of course we recognize that the Gaussian Analysis result is probably seriously flawed given the large value of Chi Squared 24 7 and the fact that it was observed to increase continually and dramatically with increasing data acquisition The appropriate warning message is prominently displayed in Figure 17 Notwithstanding the apparent qualitative resemblance of the autocorrelation data for the broad unimodal emulsion and the bimodal latex suspension the NICOMP Distribution Analysis has little difficulty revealing the true nature of the latter sample Figure 18a clearly shows a bimodal Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 57 T mir Toot ait AM N DLS THEORY distribution which is surprisingly close to the correct result given the relatively small amount of data acquired Data 374K It yields volume weighted peak mean diameters of 92 9 and 284 3 nm which are close to the nominal values of 91 and 261 nm Of course more data need to be acquired before one can be confident that the Distribution Analysis result has reached a stable solution the Fit Error value of 2 88 is still too high providing a cautionary signal Nevertheless with this small amount of data the value of Chi Squared for the alternative Gaussian Analysis has already climbed to 5 9 The values of volume found for the two pe
115. he icons near the top of the Main Window and the Status Bar located at the bottom of the Window are activated checked They should remain activated v Tool Bar v Status Bar Clock Tool Bar The Tool Bar is the bar that displays all of the CW388 icons that may be used during the use of the CW388 Windows software Following is a summary of each of the icons displayed on the Tool Bar A template has also been provided in this manual for ease of use Read Data File gt A data file that has been stored following a measurement can be retrieved to display the resulting particle size distribution PSD with the desired weighting When this option is selected a list of data files will display in a the Read Data File window The right most icon will overlay the PSD associated with the new retrieved file onto existing PSDs that are already displayed belonging to files that have already been retrieved and displayed Additional distributions corresponding to other data files can be accessed using this option Each distribution curve on the screen can be identified by matching its color with that of the data file name displayed at the bottom of the screen A maximum of eight data files can be overlaid Save Data File Use this option to manually save the data collected for a particular sample run Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 12 TE Total NICOMP SOFTWARE A m Run Autocorrelator The u
116. he two peaks Figure 20 shows a logarithmic plot of the reduced autocorrelation data for a 3 1 mixture by mass or volume of 91 and 1091 nm latex standards Here for the first time we see a very substantial curvature in the reduced data with the initial slope at t 0 significantly greater than the final slope at t 64At This difference notwithstanding it is still surprising that there is no obvious breakpoint in the plot despite the factor of 12 difference in the two constituent particle sizes However this sample is handled with relative ease by the NICOMP algorithm The Distribution Analysis result video display is shown in Figure 21 After only 7 minutes of data acquisition the analysis finds two peaks at 84 and 1041 nm quite close to the correct values Even the relative volumes are accurate 77 23 compared to the ideal values of 75 25 In conclusion it is worth returning to the issue of statistical accuracy of the autocorrelation function and the stability of the results produced by the Distribution Analysis Here it turns out to be even more important than for the Gaussian Analysis that adequate data be acquired so that the ILT algorithm can settle to a reliable result In general the more complex the distribution the longer the time needed to obtain a stable result This principle is illustrated nicely by a sequence of results obtained from the 91 261 nm test bimodal discussed earlier This sample was run again and
117. his option Print output The printer will begin printing the Volume weighted Gaussian analysis plot together with a summary of the relevant parameters and a listing of the input menu The instrument will automatically print the results when preset values of elapsed time or Fit Error are reached CNTL P Preview Printout A preview of the distribution printout will display The following options are available to click on after the distribution is reviewed Print A printout of the distribution being previewed can be obtained Zoom In Clicking on this button magnifies the preview so that it is readable Close Closes the preview of the distribution and the original distribution which displayed prior to accessing this options redisplays Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 63 TT A ay MEA m NICOMP SOFTWARE Q Display Cumulative Distribution The coefficients of the Gaussian analysis will display as a cumulative sum starting at the lowest diameter and will increase towards the larger diameter R Run Autocorrelator Used to exit from the input menu and begin data acquisition in the autocorrelator It is presumed that the Channel Width listed in the input menu is appropriate for the particular sample being analyzed Ctrl RJ Resume Running Autocorrelator Taking Data from Sample S Restores taking data once the system is halted from collected data from a sample run Stop Correlator Used to st
118. ifferent distribution weightings population number wt area wt and volume wt 1 Click on the Print tool The Printout Option window will display with print options Please refer to the FILE section of this manual for available PRINT options 2 Position the highlight bar over the desired option Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 15 NICOMP SOFTWARE 3 Click on the box located to the left of the selection A check mark will display to the left ofthe option 4 Continue steps b and c until all print desired print options are selected If a print option is not desired and a check mark is displayed in the box to the left of the option position the cursor on the check mark and click once 5 Click on OK when the desired combination of printout options has been selected in the above manner This will initiate communication with the printer displaying the printer setup window 6 Click on OK to start printing Please refer to the Distribution section of this manual to view the different distribution printouts that are available Display Help for Current Task or Command Provides quick on line help for a particular operation Flow Pump On mm This icon is used to start the flow pump and is used in flushing the system if autodilution is present Start Measurement This icon is used to START the sequence of particle size measurements Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5
119. in organic fluids There are two issues that must be considered when using any nonaqueous solvent for the suspending medium First if the Autodilution option is to be used in the 380 be certain that the silicone tubing in the unit is compatible with the solvent in question All of the other components in the 380 which come into contact with the diluting solvent are highly resistant to most fluids These include the teflon injection valve stainless steel dilution chamber and glass flow through scattering cell A copy of the MasterFlex tubing compatibility chart obtained from Cole Parmer Instrument Company located in Chicago Illinois is shown in table B 1 The relevant column which must be considered is the one labeled S for silicone tubing Important When considering whether to subject the fluidics system to a fluid other than water there is a good rule to follow When in doubt don t Only those fluids which are clearly marked by an X satisfactory in column S are safe to use in the Autodilution system of the Nicomp If Table B does not provide adequate information for a particular solvent a simple experiment can be performed to test the compatibility of the tubing to the solvent A short length of silicone tubing e g the last couple of inches of the output drain line can be immersed overnight in the solvent in question If the physical characteristics of the tubing remain unchanged i e if there is no swelling or shrinkage
120. ion resulting in a high standard deviation and baseline adjust reading Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 6 1 SAMPLE ANALYSIS RUN Hardware 1 Apply power to the system in the following order Computer Nicomp power switch is located on the back right hand panel of the instrument The Nicomp must be powered up for a minimum of twenty minutes prior to making a measurement 2 Access the Nicomp software Procedure Autodilution 1 Click on the Setup menu from the title bar and ensure that flow cell is checked on 2 Press F2 or choose Auto Print Save Menu from the Particle Sizing pull down menu 3 Ensure that parameters are set according to the below Auto Print Save Menu Menu File CAPSS Software cw388 version 1 68 Cw388 tbl Data Directory EN B rowse test File Name Printout ID 261 nm latex standard Auto Operation Options No Print Save Cycles D Using Run Time 5 min C Using Fit Error lt n2 with Chi Squared gt D Clear Autocorrelator Print Result Printout Option Automatic Choice of Distrib Gauss vs NICOMP IV Store Data on Disk Overwrite Old File Cancel Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page6 2 NOTE The menu file location may be different for each computer and is unimportant for this exercise 4 Press F3 or choose Control Menu from the Particle Sizing pull down menu 5 Ensure that parameters are set according
121. isition The reason for this that the Standard Deviation has not yet settled to a constant value Clearly a higher degree of statistical accuracy i e signal noise ratio is required in the autocorrelation function to establish the value of the curvature coefficient a in the least squares quadratic fit Equation 11 than is needed to fix the value of the linear coefficient a Consequently early into the run we observe a 20 variation in the Standard Deviation coming from az as opposed to less than a 2 fluctuation in the intensity weighted Mean Diameter from a4 This affects the results in two ways First the relatively large values of the Standard Deviation 30 to 35 of the Mean Diameter serve to push the volume weighted Mean Diameter fully 10 below the intensity weighted value to approximately 209 nm Second because of the substantial fluctuation in the computed Standard Deviation in the early stages of data acquisition the volume weighted Mean Diameter fluctuates considerably more than does the intensity weighted value However regarding these fluctuations a simple rule applies allow more time Obviously the sample represented in Table 1 is rather well behaved requiring no Baseline Adjust and yielding good results very early into the run close to the final settled Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 38 ME Toot at DLS THEORY m values Nevertheless the comments above apply equally well
122. l only about 0 6 micron The connection between the diffusion of particles and the resulting fluctuations in scattered intensity is perhaps more easily understood by considering a simplified situation in which there are only two particles in suspension shown in Figure 2 The net intensity at the detector located far from the scattering cell with a pinhole aperture is a result of the superposition of only two scattered waves In Figure 2 we have defined the two optical path lengths L ha Ip and L2 I24 l More precisely the optical path length is the distance corrected by the index of refraction but for simplicity we assume an index of 1 0 and show L and L to be simple distances in Figure 2 When the positions of the two particles are such that the difference in optical path lengths AL L Lz becomes equal to an integral multiple of the laser wavelength A then the two scattered waves will arrive in phase at the detector This is called total constructive interference and produces the largest possible intensity at the detector Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 3 DLS THEORY 1 incident lightwave incident wavefront pinhole aperture PMT photopulse signal Detector to autocorrelator Figure 2 Simplified scattering model two diffusing particles At the other extreme it is possible for the two particles to find themselves at positions such that AL equals an odd nu
123. l run time all other Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 26 28 P T Tj i DLS THEORY AAA variables being equal The result of the Gaussian Analysis when applied to the autocorrelation data of Figs IV 9a b is shown in Figure lOa Gaussian Analysis Solid Particies Chi Squared 1 1 Figure lOa Intensity weighted Gaussian Analysis corresponding to the data of Figure 9a and b This is the summary under the decaying curve C t which is updated approximately every 30 seconds on the video display It carries the label Intensity Weighting because it represents the immediate result of the cumulants calculation before any specific type of particle weighting is taken into consideration That is the underlying autocorrelation function C t is constructed from the original scattered intensity values as a function of time Hence the quadratic fit and the corresponding Gaussian like representation of the distribution of particle diffusivities and ultimately diameters reflect the fact that the D contributions or R contributions are weighted by their corresponding scattering intensities Again the peak shown in Figure lOa is approximately a Gaussian shape with respect to the log diameter scale i e it is approximately a log normal shape provided the standard deviation is not excessive 25 or so of the mean value Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 27 T mir AM D
124. lar diameter multiplied by the particle volume or mass for that diameter These new factors are obtained from the previous intensity weighted factors by dividing each by Vi or dif The resulting approximate distribution is shown above in Figure lOb To obtain the number weighted result we perform one additional division of the weighting factors by V or d to obtain a final weighting factor of just Ni shown in Figure lOc above In the case in which the intraparticle form factor cannot be ignored it must first be divided out of the original intensity weighted factors The printouts that correspond to the three weightings discussed above are shown in Figure 11a b and c The various distribution plots are labeled with two sets of numbers The REL set is normalized such that the peak value is always 100 The PERCENT set is normalized such that the sum of all of the numbers equals 100 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page2 32 DLS THEORY JAA Gaussian Analysis Solid Particles i Fat emulsion test sample SIZE nanometers INTENSITY REL PERCENT 50 0 RESULTS Gaussian Analysis Intens Weighting Mean Diameter nm 0 0 1 4 10 1 Standard Deviation ei 67 5 nmi 19 5 2 0 0 Chi Squared 1 1 a LI P 1 5 7 9 6 7 8 4 5 8 8 Baseline Adjust 0 01 Mean Diffusion 2 05E 8 cm2 sec Data 1047 0 K Cumulative Results percent Diam i i i i
125. lcohol Dimethyl Formamide Essential Oils Ethers Ethyl Acetate Ethyl Alcohol Ethyl Bromide Ethyl Chloride Ethylamine Ethylene Chlorhydrin Ethylene Dichloride Ethylene Glycol Ethylene Oxide Fatty Acids Ferric Chloride Ferric Sulfate Ferrous Chloride Ferrous Sulfate Flouboric Acid Flouroborate Salts Fluosilicic Acid Formaldehyde Formic Acid Freon TMS Gasoline high aromatic Gasoline non aromatic Glucose Glue P V A Glycerin DDUU gt wo gt gt O 0O UU gt PPP gt gt U gt gt gt 0 UO gt gt COLO 20 gt 01 PWPUOPDP B PU U UPDP Pu gt gt gt um gt 100000 O DOJO UDOUOU OP DD m UUUUUUUUUU OO 00O gt BP gt gt gt gt 000 U UUOOOOOOUO OUO OOO O0OUO OoZ2 gt gt OUU gt W gt gt UUUOW gt gt W gt gt gt gt gt WWWWW gt W gt WUOO WOUUWWWWUUUWWWN gt gt gt U0 00 gt O00OWOUPUOOUOWODVO 00 gt 00000 UU PPPUUUWWP gt gt gt gt gt gt PWWUUUUUWUUWUUHUUUPTWUUUUn gt gt gt wWwWUwWwu gt gt gt gt gt gt gt gt uUWwWuU uU Do uUUWIOUWUOUIUOUOUT gt gt gt UWV U UI UUU gt M m gt i gt gt gt Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page C 5 APPENDIX C Fluid Tubing Head Material PSF PC PPS SS o lt Hydriodic Acid Hydrobromic Acid 30 Hydrochloric Acid 100 conc Hydrochloric Acid Hydrochloric Acid med Hydrocyanic Acid
126. le name will be erased and the new one stored in its place No no new file will be stored Add Data Subtract Data Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 6 Print NICOMP SOFTWARE Printouts of the sample distributions that display on the computer monitor can be achieved using this option Please refer to Appendix A for printout samples 1 Click on the File Window option and position the highlight bar over the Print option and click The following Printout Window will display PRINTOUT OPTION Gaussian Distribution Volume Weighted Intensity Weighted Number Weighted Int Vol Weighted Summary Result Autocorrelation Function Autocorrelation Data Time History Plot Channel Error Plot Gaussian NICOMP All Weighted NICOMP Distribution F Volume Weighted Intensity Weighted Number Weighted Int Vol wW eighted Cancel 2 Click on the square located to the left of the print selection A black check mark will display in the box selected 3 Click on OK to start printing the distributions and or plots The following Print window will display Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 7 NICOMP SOFTWARE Print 2 x Printer Name KONICA MINOLTA C350 PCL5c Properties Status Ready Type KONICA MINOLTA C350 PCL5c Where IP 192 168 0 199 Comment Print to file Print range Copies e All Number of copies 1 z e
127. lected from a sample run when in display mode The data will display on the screen in the following format counts X1000 AUTOCORRELATION FUNCTION Solid Rania m Run Time hr min sec 1700 0 10 3 1600 Ch 1 Data 1500 1000 1854 1371 Count Rate uSec gt Channel Width 34 0 uSec x1000 Channel Data xi 1853952 1836313 1819471 1803168 1787453 1772468 1757910 1744052 1730653 1717803 1705383 1693483 1682026 1670905 1660244 1649975 1640201 1630695 1621584 1612930 1604508 1596515 1588812 1581370 1574229 1567343 1560768 1554388 1548307 1542408 1536755 1531252 1526034 1521001 1516098 1511498 1507019 1502785 1498654 1494708 1490869 1487238 1483653 1480286 1477037 1473887 1470881 1468004 1465220 1462538 1459950 1457396 1454998 1452725 1450496 1448345 1446306 1444389 1442439 1440638 1438923 1437254 1435602 1434026 Baseline 1393856 Total Counts 225460 K Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 52 NICOMP SOFTWARE Channel Error Used to compare the statistical discrepancy between the actual data and the ability to curve fit the data CHANNEL ERROR Solid Particle Run Time hr min sec 0 10 3 Channel gt Channel Error unused channel 6 526 4 953 4 256 1 211 1 304 1 347 2 376 4 018 4 162 5 166 5 734 3 825 2 608 0 894 1 570 0 523 1 890 0 555 1 921 0 505 0 429 0 330 0 613 0 305 1 372 0 664 1 583 0 867 0 765 1 582 1 439 1 835 4 449 2 992 3 537 1 389 1 465 0 562 1 383 0 278
128. licable requirements of 21 CFR Subchapter J 1040 10 and 1040 11 Radiation Control for Health and Safety Act of 1968 42 U S C 263f As presently constructed this instrument is designated by the Bureau of Radiological Health Class product Exposure to negligible levels of Laser Radiation during normal operation results The two labels below are affixed to the back panel of the Nicomp 380 Autodilute They attest to the above Safety Certification and also establish the place and date of manufacture of the unit THIS EQUIPMENT CONFORMS TO PROVISIONS OF US 21 CFR 1040 10 AND 1040 11 75 Aero Camino Suite B Santa Barbara CA 93117 Tel 805 968 1497 FAX 805 968 0361 MODEL SERIAL NO VOLTS MANUFACTURED MONTH YEAR MADE IN UNITED STATES OF AMERICA Important Read carefully before attempting to operate the Nicomp If the Nicomp is to be used with the Autodilution option then all liquid samples will be introduced into the system by means of a syringe or tube connected to the manual sampling valve that is located on the front panel of the instrument In this case NO entry into the sample holder space will be required Alternatively if the Nicomp is to be used without the autodilution option then all liquid samples will be introduced into the light scattering cell using 6 mm disposable glass culture tubes or standard 1 cm cuvettes In this case entry into the sample cell holder space will be required Nicomp
129. ltiple scattering Therefore the viscosity which is needed in the Control Menu is only that of the pure diluent in which the sample particles are suspended The value of 0 933 cP shown in the Control Menu example above is the viscosity of water at 23 C The viscosity of a simple solvent can easily be determined from reference books However it must be remembered that the viscosity of many solvents including water is highly dependent on temperature Viscosity values for various common organic solvents can be found in Appendix C of this manual Liquid Index of Refraction This parameter establishes the index of refraction of the solvent in which the particles are suspended assuming a dilute suspension The value of 1 333 shown in the example above is the index of refraction of water Unlike the viscosity the index of refraction has very little dependence on temperature Values for various common organic solvents can be found in Appendix D of this manual Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 25 T ri Ts Toot at AM N NICOMP SOFTWARE Intensity Setpoint The average scattered intensity or photopulse rate expressed in kHz which is desired for a measurement can be established by setting this parameter The default value is set to 300 kHz This value is typically recommended for most samples which scatter adequately Itis designed to optimize the efficiency of the autocorrelation process and thereby minimize the ti
130. manually setting the parameters for sample analysis Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 28 rity Til iT jT NICOMP SOFTWARE JAIL Auto Baseline Adj Adjustment Adjustment of the autocorrelation baseline function can be made automatically in order to obtain the best theoretical fit by setting this parameter If this option is selected automatic adjustment of the baseline will be made during each calculation cycle 30 sec for both the Gaussian Analysis and the NICOMP Distribution Analysis The baseline will be permitted to rise in very small increments in order to find the level at which the goodness of fit is optimized In general baseline adjustment is required for samples which contain a long tail of aggregates or other large off scale particles If this option is not selected the baseline adjustment is expressed as a percentage of the original measured baseline before adjustment The value can be considered small and therefore unimportant if it consistently remains below 0 03 Values larger than 0 1096 provide a useful indication of the presence of significant amounts of large particles in the sample In the NICOMP Distribution Analysis the automatic baseline adjustment is called the Residual and is expressed as a pure number A value of 200 which would be considered extremely large indicates a doubling of the actual measured baseline before adjustment Residual values smaller than 2 or 3 ca
131. mber of Auto Operation Cycles is required it would be beneficial to request automatic data storage but not printouts for each cycle It may be more convenient to produce the printouts later after they have been reviewed If this option is not selected no printout will result However the data for the sample run will be saved if the parameter to do so has been set The data saved under the directory and file name specified in the Auto Print Save Menu can then be reviewed and printed at a later date Printout Option Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 33 Nicomp PSS 380Nicomp 030806 06 06 Page 5 34 NICOMP SOFTWARE A pre selection of the type s of printout s desired after the completion of each Auto Operation Cycle must be made using this option Each analysis type and weighting which is desired can be chosen by following these steps PRINTOUT OPTION x Gaussian Distribution NICOMP Distribution Volume Weighted Volume Weighted Intensity Weighted Intensity Weighted Number Weighted Number Weighted Int ol Weighted Int Vol Weighted 0 M Summary Result Gaussian NICOMP All Weighted Cancel 1 Click on the square box located to the left of the desired print selection A black check mark will display in the box 2 Click on OK after all print selections has been made Please refer to the DISPLAY section of this manual for examples of all possible types of printouts Please
132. mber of half wavelengths 4 2 In this case the two scattered waves arrive at the detector totally out of phase with each other This is total destructive interference resulting in zero net intensity Over time diffusion of the particles will cause the net intensity at the detector to fluctuate in random fashion like a typical noise signal between these two extreme values A representative total intensity signal is shown in Figure 3 The intensity varies between the maximum value and the minimum value zero when the optical path length difference changes i e increases or decreases by A 2 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 4 ME DLS THEORY m l MAX Figure 3 Typical intensity vs time for two diffusing particles The key physical concept that underlies the DLS particle sizing measurement is the fact that the time scale of the fluctuations shown in Figure 3 depends on the size of the particles For simplicity at this point in the discussion we assume the particles to be uniform in size with a single well defined diffusion coefficient Small particles will jitter about in solution relatively rapidly resulting in a rapidly fluctuating intensity signal by contrast larger ones will diffuse more slowly resulting in a more slowly varying intensity At this point we make the simplifying assumption that the temperature of the particle suspension is held constant We shall see that the temperature play
133. me needed to obtain reliable accurate results for most samples The intensity setpoint is relevant for both modes of sample measurement i e using either Autodilution or Drop in Cell Autodilution mode The scattering intensity will increase to a relatively high value as the sample concentration initially increases due to the flow of fresh sample and diluent through the mixing chamber and into the flow through sample cell in the Nicomp It will then reach a maximum and then decrease as the sample concentration falls When the intensity falls to approximately the Intensity Setpoint the pump in the Nicomp will halt thereby stopping the flow of fresh diluent into the flow through sample cell The scattering intensity will then stabilize because the particle concentration will no longer change Drop in Cell mode The sensitivity will be changed automatically in Automode operation so that the final intensity will approximate the value stored for the Intensity Setpoint This will be the case provided the sample concentration is not too large or small thereby putting the initial scattering intensity outside the accessible range of the automatic system The sensitivity can be adjusted manually at any time one of two ways 1 Alter the concentration of the sample 2 Increase or decrease the incident laser intensity by adjusting the neutral density filter First Channel Used This parameter is used to establish the channel number in the aut
134. mp 380 Manual PSS 380Nicomp 030806 06 06 Page 3 1 INITIAL HARDWARE SETUP 3 Plug the power cable provided into the back of the unit 4 If the unit has the Autodilution feature place the drain line into a waste bucket 5 Apply power to the PC controller 6 Install the Nicomp software package Review the Software section of this manual 7 Apply power to the Nicomp Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 3 2 rity Til iT jT SOFTWARE INSTALLATION JAMAL SOFTWARE INSTALLATION The software program that is responsible for controlling the NICOMP Submicron Particle Sizer and also the NICOMP Computing Autocorrelator is named CW388 The version can be identified when the program CW380 is run it is displayed in the upper right hand corner of the default display screen which is the Main Menu The software diskette that is included with the NICOMP 380 will contain four software files CW388 EXE the largest of the four files It is the executable program which controls the Instrument and initiates the running of the software CW388 CFG contains the configuration of the 380 system The configuration includes the identification of the serial port for communication with the 380 instrument and the parallel port for printing as well as specification of available options such as the flow pump for Autodilution use of an external laser and the multi angle accessory In some cases the CW380 CD Rom will be mis
135. n e UA AS Gees 1 Verify the printer type selected If changes need to be made follow these steps 2 Click on the Down arrow A window of all of the printer brands and types will display Selection of the correct printer driver software depends on the setup of this option 3 Position the highlight bar over the Printer type and model that is currently hooked up to the computer being used 4 Click the mouse once 5 Click on the OK button The Print windows will re display with the printer and type and model selected 6 Position the cursor over the print range desired All will print all data pertinent to the distribution being reviewed Pages will print the range of pages desired Selection will only print those pages desired 7 Position the cursor in the Copies option and type a number for the number of copied desired for the printout The default is set to one 8 Click on the OK button to start printing Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 8 Print Preview NICOMP SOFTWARE Allows the printout to be viewed prior to being sent to the printer Following is an example of the window that will display A data file must be accessed prior to using the Print Preview option Print will print the distribution that is being previewed Next Page will advance to the next page of the preview Two Pages will preview two pages of the same file side by side Zoom In provides
136. n be considered negligible while values in excess of 10 indicate the presence of significant amounts of off scale particles These larger values usually have the effect of pushing to smaller diameters any peaks in the size distribution particularly in the case of bimodals After all of the choices have been made and reviewed in this window exit to the Main Menu by clicking on OK Cancellation of all changes made may be accomplished by clicking on CANCEL Cum Set Pt Autodilution ND Position Sets the neutral density so it mimics a standard 5mW HeNe laser with a PMT Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 29 NICOMP SOFTWARE Auto Print save Menu Access to the Edit Auto Save Print Menu is gained using this option This is the second of the two menus used to define the basic operation of the Nicomp Position the highlight bar over the Auto Print Save Menu and click once The Auto Print Save Menu will display Auto Print Save Menu x Menu File CAPSS Software cw388 version 1 58Cw388 tbl Data Directory Bi g rowse test 0 File Name Printout ID 350nm 220nm 7 3 Auto Operation Options No Print Save Cycles E Using Run Time 5 o min C Using Fit Error lt n2 M with Chi Squared gt D Clear Autocorrelator Print Result Printout Option V Automatic Choice of Distrib Gauss vs NICOMP IV Store Data on Disk M Overwrite Old File Cancel Press TAB to advance to each
137. nce The Printout option window will display Click on the square box next to the desired selection A black check mark will display Click on OK The Print Window will display Click on OK to start printing Post Measurement System Flush Press Ctrl F to draw clean de ionized water into the system via the injection port Pay close attention to the front display panel to see that the counts start to lower Keep injecting fresh diluent until the counts fall below 10 kHz It is important to remember to clean the small injection valve on top of the system between sample runs to avoid cross contamination This can be accomplished by following these steps a Filla 10cc syringe with clean diluent b Inject it into the system by turning the valve mounted on the front panel of the instrument 90 counter clockwise The system will draw in all of the diluent in the syringe c Rotate the valve 90 clockwise to its original position when approximately 5 cc s has been pulled into the system Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 6 9 MAE MEL SYSTEM MAINTENANCE The absolute accuracy of the particle sizing technique used in the Nicomp depends on the frequency of a crystal controlled clock and the wavelength of a laser neither of which can drift over time The only other relevant variable is the scattering angle The optics of the Nicomp have been permanently aligned at Particle Sizing Systems factory and should req
138. nd hence good analyses Suffice to say there is more than meets the eye in the successful extraction of particle size distributions using the DLS technique More evidence will unfold below supporting the claim that this can be a difficult business Nonetheless let us take courage and press on Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 24 rity Til iT jT DLS THEORY MAAL What is needed clearly is a method for dealing with the simple kind of polydispersity in the particle size distribution illustrated by the IV emulsion example above The word simple is used to emphasize the fact that we have gone from a sharp population consisting of essentially one size to one which represents a smooth not too wide range of sizes centered about some average In the case of a sharp distribution it is a simple matter to obtain the best straight line fit to the logarithm of the reduced data loge C t B vs t using the well known method of least squares One simply adjusts the slope and intercept of the straight line to minimize the sum of the squares of the deviations or errors between the reduced data points and the values implied by the theory i e by the straight line The needed generalization which can deal effectively with non exponential behavior of C t B brought about by smooth Gaussian like distributions of particle diameters is provided by the methods of cumulants This procedure first int
139. ng to be measured on a repeat basis Use of the same channel width for each sample measurement may improve the reproducibility of the results Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 24 rity Til iT jT NICOMP SOFTWARE JAIL The default value for the channel width in the table file CW388 TBL is 10 usec which is appropriate for a mean particle diameter of approximately 100 nanometers nm or 0 1 micron If much smaller particles such as small proteins or surfactant micelles with mean diameters between 5 and 10 nm were to be measured a value of about 1 usec would be more appropriate for the channel width Temperature The temperature of the sample cell which is regulated to within 0 2 C by a Peltier thermoelectric element and feedback circuit can be moderated using this option The value entered will be transmitted to the temperature regulator when the Control Menu is closed The software includes a lower limit of 4 C and an upper limit of 60 C Important Sufficient time must be allowed for the temperature of the sample to reach that of the cell holder It is the cell holder that is regulated in the Nicomp a minimum of 5 minutes should be allowed Liquid Viscosity The viscosity of the sample suspension is expressed in units of centipoise cP The particle suspension must be very dilute for measurements based on dynamic light scattering DLS in order to avoid errors due to interparticle interactions and or mu
140. ng value suitable for a broad unimodal distribution The following table provides some suggested criteria for choosing a value for Smoothing STD DEV CHI SQR Choice of Smoothing Standard Deviation Chi Squared lt 15 lt 2 Narrow single peak Use low Smoothing 1 3 gt 20 lt 2 Broad unimodal distribution Use high Smoothing 5 6 gt 20 gt 3 Probably a bimodal distribution Use low Smoothing 1 3 Table 3 Smoothing Table It is important to realize that a relatively high degree of smoothing in the Distribution analysis calculation is especially useful when investigating a broad single peak distribution may be achieved by properly manipulating the parameters Plot Size and Range If a low value for the Plot Size is 30 and a relatively large value for Range i e for a given value of IN DIAM These choices will usually improve the performance of the instrument when analyzing broad particle size distributions such as are frequently encountered with dry milled powders dyes pigments abrasives etc and emulsions produced by homogenization or other procedures Plot Range Determines the maximum diameter of the size scale as a multiple of the MIN diameter The plot range can be any number from 10 to 1000 The default value is set at 50 which sets the maximum diameter to be 50 times the value of the minimum diameter Click on the OK button to save the parameters to memory Nicomp 380 Manual P
141. nual PSS 380Nicomp 030806 06 06 Page A 7 APPENDIX A INT VOL WEIGHTED Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm 7 3 Data File C Program Files Particle Sizing Systems Nicomp 388Wwersion 1 68 Bimodal 300 INT VOL Weighted NICOMP DISTRIBUTION Analysis Solid Particle Fit Error 735 Residual 0 00 Chi Squared 386 Baseline Adj 0 00 96 Run Time 0 Hr 6 Min 8 Sec Wavelength 26328 nm Count Rate 0 KH Temperature 23 degC Channel 1 6126 K Viscosity 0 933 cp Channel Width 35 0 uSec Index of Ref 1 333 INT WT NICOMP DISTRIBUTION Intensity Weighting Peak 1Peak2 Peak 3 Mean Diam nm 230 8 345 3 Stnd Dev nm 7 4 138 C V 3 22 4 01 e Percent 42 91 57 09 Min Diam 100 Plot Size 50 Smoothing 2 Plot Range 10 Volume Weighting Peak 1 Peak 2 Peak 3 Mean Diam nm 230 5 345 9 Stnd Dev nm T5 13 4 e C V 3 26 3 89 e Percent 38 52 61 48 e Diam nm gt Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 8 APPENDIX SUMMARY RESULT Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm 7 3 DataFile C ProgramFiles P article Siang Systems Nicomp388 version 1 68 Bimodal 300 Run time Data Ch 1 Avg Intensity Sensitivity OHr 6Min 8Sec 612 6K 0 0kHz 150 Gaussian Analysis Solid Particle NICOMP Analysis Mean Dia Std Dev Chi Sq Base Adj Peak 1 Peak2 Peak3 VOL 295 5nm 53 3nm 3 86 0
142. nual PSS 380Nicomp 030806 06 06 Page 2 9 T mir AM DLS THEORY What can we say about the shape of C t as a function of the sampling separation t Without knowing anything about the physics of diffusion and its effect on I t we can nevertheless say something useful about C t in two limiting extreme cases t gt 0 and t gt In the limit in which t approaches zero the two sampled intensities are essentially identical because there is no time for the particles to rearrange their positions Hence C O t 4 That is the value of C t for t gt 0 is simply the sum over many values of t of the square of the scattering intensity In the opposite limit in which the sampling interval t becomes very large approaching infinity we have already seen Figure 5 that there should be no correlation between the pair of sampled intensities Hence Equation 3 reduces to the square of the average scattering intensity t i e the normalized sum of I t values taken over many values of t C o0 lt Is t gt 5 It is known and easily demonstrated that for any fluctuating quantity the average of the squares of that quantity is always larger than the square of the average lt I4 t gt gt lt I t gt 6 The quantity on the right hand side of Equation 6 is the lowest value possible for the correlation function all other values of C t for finite values of t must in principle be larger than th
143. number of cycles specified in the AutoPrint Save Menu A separate file is saved for each cycle and the numerical extension is incremented by 1 each time Using Fit Error with Chi Squared The other way in which an Operation Cycle can be defined is through the Fit Error Parameter alone or a combination of the Fit Error and the Chi Squared parameters Fit Error 1 Click on the circle located to the left of the option and click once The upper limit for the Fit Error will appear on the same line and to the right 2 Enter the desired upper limit for the Fit Error The end of the first Operation Cycle will occur when the Fit Error falls below this value causing the pre selected Operating Functions to be implemented Fit Error and Chi Squared Distributions which are relatively complex e g bimodals and trimodals or asymmetric unimodals having a tail due to large aggregates the NICOMP Distribution Analysis must be used to obtain a reasonable result By contrast the simple 2 parameter Gaussian Analysis will be completely inadequate to characterize such complex distributions as generally indicated by a relatively large value for the Chi Squared goodness of fit parameter In general Chi Squared will remain close to one only when the true distribution is close to a simple unimodal Consequently when it is expected to obtain non Gaussian results it is useful to combine the requirements of a low value for the Fit Error with a high value
144. o the overall molecular weight of the particle MW or its volume V for a given particle density The constants of proportionality that connect these various physical quantities depend on the indices of refraction of the particle n and solvent n That is how well a given particle scatters light depends not only on MW or V but also on the polarizability of the particle related to n relative to that of the solvent related to n For the very small particles in the Rayleigh region we arrive at simple expressions for the scattered intensity ls l f nsn MW I 1a or r ls g np ns V lo 1b where is the incident laser intensity and f n ns and g n n are functions of the indices of refraction of the particle and solvent which are fixed for a given system composition e g latex particles in water For these small particles in the Rayleigh region i e diameters lt approx 0 1 micron or 100 nm there is negligible angular dependence in the scattered intensity The simple expressions above must be modified when the characteristic particle dimension i e the diameter in the case of spheres is no longer negligible compared to the wavelength of the incident light beam In this so called Mie Scattering region Equations la and 1b must be altered to take account of intra particle interference With a larger particle the oscillating electrons no longer oscillate together in phase the individual scattered waves originating
145. ocorrelation function which is chosen to be the starting channel for all calculations of the particle size distribution The default value is 2 which is the preferred value for most measurements The value of 2 means that the first channel is effectively discarded this will eliminate artifacts in the results caused by high frequency noise principally after pulsing in the PMT detector Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 26 TAE Tiam at NICOMP SOFTWARE m In principle it will be useful to increase the value of this parameter to 4 the maximum value in cases where the channel width is very small i e 0 5 microsec This will reduce artifacts in the analysis results caused by PMT after pulsing which influences the initial 1 to 1 5 microsec of the autocorrelation function Laser Wavelength The laser wavelength parameter is normally not accessible The default value is 632 8 nm which is the wavelength for the small HeNe laser used in the basic Nicomp system Please refer to the Setup menu section of this manual for all wavelength settings If the system includes a laser that is not HeNe it is necessary to change this parameter in the Setup Window External Fiber Angle Scattering Angle The external angle of the stepper motor arm and the resulting actual scattering angle are only accessible when the multi angle option is used with the system The default value for both parameters is 90 0 degrees which is
146. of the parameters on the screen Menu File Specifies the file name under which this table of parameters will be saved Data Directory File Name The data directory is used to specify the path drive directory where the raw data obtained from a measurement using either the automatic or manual save data mode will be stored in memory The path includes the disk drive allocation as well as any subdirectory path that is desired Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 30 TT Ti 3T Ti qm t NICOMP SOFTWARE Y Example To store data blocks on disk drive C using the directory CW388 and the sub directory data 1 Double click on Browse to locate the directory in which to save the data files This directory must be created before entering the Nicomp software 2 Position the cursor in the File Name window and type the desired file name followed by a numerical extension Extension numbers are automatically incremented after each analysis unless the user intervenes and renames the run Note File names are a maximum of 8 alphanumeric positions Printout ID The printout caption will display on all data screens and on all printouts for a particular data file sample run This caption will display on all printouts until it is manually changed 1 Edit Printout ID 2 Position the cursor in the Printout ID window 3 Typein or edit the existing caption in the window that displays Use the Backspace key to delete any un
147. on lt 284 1 nm 75 of distribution lt 320 9 nm 90 of distribution lt 358 0 nm 99 96 of distribution lt 432 2 nm 80 96 of distribution 330 7 nm Run Time 0 Hr 6 Min 8 Sec W avelength 632 8 nm Count Rate 0 KHz Temperature 23 deg C Channel 1 6126 K Viscosity 0 933 cp Channel Width 35 0 uSec Index of Ref 1 333 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 2 APPENDIX NUMBER WEIGHTED GAUSSIAN Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm 7 3 Data File C Program FilesiP article Sizing Systems Nicomp 388 version 1 68 Bimodal 300 NUMBER Weighted GAUSSIAN DISTRIBUTION Analysis Solid Particle GAU SSIAN SUMMARY Mean Diameter 263 8 nm Variance P I 0 033 Stnd Deviation 47 6 nm 18 0 Chi Squared 3 861 Norm Stnd Dev 0 180 Baseline Adj 0 000 Coeff of Var n Z Avg Diff Coeff 1 61E 008 cm2 s NUMBER WT GAU SSIAN DISTRIBUTION 0 20 Diam nm gt Bimodal 300 Run Time 0 Hr 6 Min 8 Sec W avelength 632 8 nm Count Rate 0 KHz Temperature 23 deg C Channel 1 2512 6 K Viscosity 20 933 cp Channel Width 35 0 uSec index of Ref 4 333 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 3 APPENDIX A INT VOLUME WEIGHTED GAUSSIAN Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm 7 3 Data File C Program Files P article Sizing Systems Nicomp 388 version 1 68 Bimodal 300
148. op the collection of data in the autocorrelator Once this option is exercised the following message will display ALT S Summary Result T V The window of the distribution divides in two and a summary of the data displays showing the Volume Number and Intensity weighted information Display Time Plot Displays the time series plot calculation history where the stability of the data in terms of mean diameter is shown not to be changing for at least three minutes as a function of time a flat straight line Toggle Vesicle Solid Particle Calculation The instrument defaults to solid particle weighting when power is first applied The weighting may be changed using this option The weighting will change when the results of a new analysis is displayed at which time the title will also change to Gaussian Analysis vesicles The original weighting can be re displayed by using this option again Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 64 rity Til iT jT NICOMP SOFTWARE JAIL W Weighting select Three weightings may be viewed for a particle size distribution using this option Volume weighted Displays the relative particle volume vs diameter The value of the volume weighted particle size distribution is calculated assuming that the particle are spheres of uniform density which scatter light according to classical Mie Theory Number weighted Displays the relative number of particles in a sample
149. p 030806 06 06 Page C 12 TT REI y v APPENDIX D AU SOLVENT TEMP C VISCOSITY cpoise INDEX REFRACTION Acetaldehyde 10 0 256 1 332 20 0 220 1 332 Acetic Acid 15 1 31 1 380 25 1 16 1 380 41 1 00 1 380 59 0 70 1 380 Acetone 15 0 337 1 357 25 0 316 1 357 41 0 280 1 357 Acetonitrile 15 0 375 1 346 25 0 345 1 346 30 0 325 1 346 n Amyl acetate 11 1 58 1 400 45 0 805 1 400 n Amyl alcohol 15 4 65 1 410 30 2 99 1 410 n Amyl ether 15 1 188 1 410 Aniline 15 5 31 1 583 25 3 71 1 583 35 2 71 1 583 Benzaldehyde 25 1 39 1 544 Benzene 20 0 652 1 498 30 0 564 1 498 40 0 503 1 498 Benzonitrile 15 1 45 1 526 25 1 24 1 526 30 1 11 1 526 Benzyl Alcohol 20 5 80 1 538 30 4 65 1 538 Benzyl amine 25 1 59 1 540 Bromoform 15 2 152 1 587 25 1 89 1 587 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page D 1 RE nami m y APPENDIX D SOLVENT n Butyl acetate n Butyl alcohol Carbon disulfide Carbon tetrachloride Chlorobenzene Chloroform Cyclohexane Cyclohexanol Cyclohexanone Cyclohexene Cyclopentane n Decane N N Dimehtylformamide DM F Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page D 2 TEMP C 20 40 20 30 40 20 40 15 20 30 40 15 20 40 20 30 39 15 30 20 30 15 30 13 5 20 13 5 20 25 20 25 VISCOSITY cpoise 732 563 2 948 2 3 1 782 0 363 0 330 1 038 0 969 0 843 0 739 0 900 0 799 0 631 0 58 0 514 0 500 1 06 0 82 68 0 41 1 2 45 1 80 0
150. propriate amount Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 20 ME T T T NICOMP SOFTWARE JAIL Flow Pump The flow pump parameter indicates whether the Nicomp contains a flow pump which is required for the Autodilution option The flow pump may be operated manually or by automatic computer control in Autodilution mode Deactivate the flow pump by selecting this option Drop in Cells Use of the flow pump must be suspended when using a drop in cell to take a measurement If it is not flooding in the unit will occur causing major damage to the instrument Change Laser Wavelength The appropriate laser wavelength for the type of external laser being used Is entered using this option The default wavelength is 632 8 nm which is required for the basic Nicomp with internal 5 mW HeNe laser LASER WAVELENGTH RLD 5 MW HENE 632 5 nm RLD 12 MW HENE 635 nm RLD 35 MW HENE 639 nm RLD 50 MW HENE 664 nm RLD 100 MW HENE 664 nm GLD 20 MW HENE 532 nm GLD 50 MW HENE 532 nm GLD 100 MW HENE 532 nm APD Overload Protection The overload protection option is selected to when trying to control the amount of scattering when too much light enters the detector Maximum Count Rate Defines the maximum amount of light that can enter the detector Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 21 mir That at AM i NICOMP SOFTWARE Intensity Overshoot Factor When the intensity
151. r an even more complicated shape We shall see that two very different mathematical procedures or algorithms have been developed to analyze the autocorrelation raw data C t depending on the nature of the underlying particle size distribution The software automatically selects the more appropriate of the two analysis procedures and provides the user with a running measure of the accuracy or goodness of fit of the computed distribution resulting from the particular analysis chosen Nevertheless we feel it essential to gain an appreciation of the rationale behind each of the analysis methods and to become comfortable with some typical results obtained for actual particle systems The latter can be studied in a controlled accurate way using polystyrene latexes oil in water emulsions and other well characterized materials Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 20 rity Til iT jT DLS THEORY AAL Broad unimodal distribution Gaussian Analysis Following the discussion in the previous section it is now obvious that a mixture of particle sizes must give rise to an autocorrelation function C t which decaying exponential function is no longer a simple i e having a single well defined decay time constant t as shown in Figure 8c The existence of more than one rate of diffusion must necessarily give rise to a mixture of decaying exponential functions each of which has a different time decay constant 7
152. refer to Appendix A for sample printouts Automatic Choice of Distrib Gaussian vs NICOMP Automatic selection of the Gaussian Analysis vs the NICOMP Distribution Analysis for the printout at the end of one of the Auto Operation Cycles can be obtained using this option Click on the square box next to the selection and a check mark will display in the box The Nicomp will make this selection automatically based on the value of Chi Squared If Chi Squared is less than 3 0 the Volume weighted Gaussian Analysis PSD will be printed at the end of each Auto Operation Cycle If Chi Squared exceeds 3 0 the Volume weighted NICOMP Distribution Analysis PSD will be printed 380 Manual rity Til iT jT NICOMP SOFTWARE JAIL Store Data on Disk Raw data can automatically be stored at the end of each Auto Operation Cycle Click on the square located next to the selection and a black check mark will display Data will then be stored in Data Directory and with the file name which has been specified in the Auto Print Save Menu Decline saving data collected for a sample run by not making this selection Important It is strongly advised to choose this Print Save option since it is impossible to reanalyze the results of a previous measurement or obtain a different printout if the raw data was never saved in memory Overwrite Old File Overwriting old files enables the saving of new data files over old no longer needed data files and con
153. relation Function a ss aeacesse SELTEREREN ssssx 1429751 t 4 4 i i 4 t L t L t 1 1 y y t t LI a Li r t 4 4 L 1 t L V 4 1382606 enm s a e me m amm deae um m cp ay tb am ap tm rm deem mao o mm mm e Intensity Weighted Gaussian Summary DIA 198 1 NM STD DEV 0 48 DECAYS 2 3 CHI SQ 24 71 BASE ADJ 0 00 X Figure 15 Autocorrelation function for a test bimodal 3 1 vol ratio 91 and 261 nm Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 53 DLS THEORY latex particles Hetero HE BRE DENTE PRESS EUIS ERDOOCAANALASCOREOAA DETAL A dd AAA rt OT VONTGUEL REGU GREECE LIAE LABOR IG Adee ET a eiie GEESE DIEGA DRIN ENT LO ENGEL GHG REIT AMR RUN SSA DAS DDR ELE USEVORURONLERTERUORGEEEUEBUDUUEBGUFRRREINERLEBERE IDO a pepper ter rb i3 EIER EE EEREEHET ESTER n asad o eos et Re nee e pretend e rre HIT MEUS FREED MA KEMANA le GEERT IMHUHTISUN MA EN NS 1 tt Unb Ate eae ee ete O OLE Mtis HIHCOHLUSHLIUULUUUUULIULUMETUUUHLULRELEOLLAULUER ER DSUESUUSLS BSUEUETAEREUESESUGSELEELS FOTOS DPEUSEINS MEERE ANOTA gt USARA OME RU LAUDO LEUR ERU VEL EB ERA EA MA LL ODER OLAS AO Ba SR URN EE GR IA LESA OA RH A DURARON EA ES SA BRA SEE ER RM RH RA A WIHIFBUHUR ORAU DES GEAR MM FOUR PROA DODGE 10100 BEN PD COLORIDO ELE FORT LATTES GERNE PIAR LR LODO ANGREN ppt MIU HS COLOCO NO E LLEILLLLU LEUTE CROLAERERS LEE LLL UA edi eene ATEOS er
154. result of the coherent addition or superposition of many individual scattered waves each of which originates from a different particle located in the illuminated detected volume This is the physical phenomenon known as interference Each individual scattered wave arriving at the detector bears a phase relationship with respect to the incident laser wave which depends on the precise position of the suspended particle from which it originates All of these waves mix together or interfere at a distant slit on the face of a photomultiplier detector PMT in Figure 1 which measures the resulting net scattering intensity at a particular scattering angle 90 degrees in the DLS Module The suspended particles are not stationary rather they move about or diffuse in random walk fashion by the process known as Brownian motion caused by collisions of neighboring solvent molecules As a consequence the phases of each of the scattered waves arriving at the PMT detector fluctuate randomly in time due to the random fluctuations in the positions of the particles that scatter the waves Because these waves interfere together at the detector the net intensity fluctuates randomly in time It is important to appreciate that only relatively small movements in particle position are needed to effect significant changes in phase and therefore to create meaningful fluctuations in the final net intensity This is because the laser wavelength is relatively smal
155. ribution of particle radii R or diameters rather than of diffusivities D This is not a problem of course because the Stokes Einstein relation Equation 2 shows that D is simply given by 1 R times a conversion constant Furthermore for relatively small ranges of D a Gaussian distribution in 1 R translates into a Gaussian shape in In R Hence we arrive at approximately a log normal shape for the distribution of particle radii or diameters Using the Stokes Einstein relation Equation 2 and Equations 12b and 13 we can therefore obtain the mean particle diameter d 2R and the standard deviation of the diameter distribution 2 AR This latter parameter is also known as the coefficient of variation and is equal to the square root of the variance it is closely related to the half width of the particle size distribution which is approximately a log normal in shape In the DLS Module we refer to this cumulants method for inverting the autocorrelation function as the Gaussian Analysis It must be stressed that it is 2 parameter fit that is except for the possibility of a change in the baseline there are only two variables which affect the goodness of fit of the quadratic function of C with respect to the reduced data log C t B Equation 11 These are coefficients a and a Coefficient ay has relatively little value in the analysis it is related to the contents of channel 1 of C t which increases with the tota
156. rmation NICOMP INPUT X Menu File Cw388 tbl Minimum Diameter Hu Plot Size 50 Smoothing 2 Plot Range 10 Cancel Menu File Contains the filename of the table that contains the information from the Auto Print Save menu and the Control menu Minimum Diameter This is the smallest size diameter that is displayed This may be any integer from 1 to 1000 nm Plot Size The number of bins or slices into which the diameter axis is divided is provided by this parameter A default value of 45 is initially set The highest resolution of the diameter scale is 60 However a more appropriate choice for most particle size distributions especially those which are more complex than a single narrow peak would be 45 Higher values of Plot Size can produce anomalous results such as false bimodals where a single broad peak is more realistic and multiple split peaks These occurrences are a consequence of the complex mathematical procedure used to generate the Distribution Analysis and are important to the overall resolution of the size distribution i e too large a number of diameter slices Plot Size within a given range of diameters Smoothing Establishes a bias for the Distribution analysis At one extreme Smoothing 1 or 2 distribution results are favored which are either narrow single peaks or bimodals At the other extreme Smoothing 5 or 6 computed fits are favored which produce broad unimodal distrib
157. roduced by Koppel has been used extensively in the past 15 years to obtain estimates of the particle size distribution from DLS In fact until only 5 or 6 years ago it was essentially the only practical method for obtaining such information The conceptual underpinning of the cumulants procedure is simplicity itself as will be seen below Suppose we consider situations for which the plot of log C t B has a relatively small curvature representing a modest deviation from straight line behavior The simplest generalization of the straight line fitting procedure is to find the quadratic function of C which lies closest to the reduced data points i e on a least squares basis The prescription for carrying out a cumulants fit is therefore very simple as summarized below 1 2 loge C t B o ao a t a t 11 A quadratic function of C which we ve indicated by a0 a t a2 t 2 now replaces the trivial straight line function bo b t All that remains is to relate the coefficients of the quadratic function in particular a and a2 to parameters that describe the corresponding particle size distribution In the simple monodisperse case discussed earlier we recall Equations lOa and b that coefficient by equals DK Hence the value of the slope negative divided by the constant K yields the diffusion coefficient D of the uniform particles It turns out in the more general case of a quadratic fit that th
158. s as important a role as the particle size in determining the diffusivity and hence the time scale of the resulting intensity fluctuations In any real situation of interest of course there are many more than two particles in suspension which contribute to the scattered intensity signal However the principle of interference remains the same The resulting signal will be observed to fluctuate average level which is proportional to the number of particles illuminated detected volume and their individual scattering power Equations 1a and 1b The time scale of the fluctuations depends on the particle diffusivity and hence on the particle size This is illustrated in Figures 4a b and c for small medium and large size particles using the same time scale on all three horizontal axes Again it must be stressed that the fluctuations in the net scattered intensity are not caused by the addition or subtraction of particles in the illuminated detected volume Rather they are the result of the variations in position of an essentially fixed number of particles within the scattering volume Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 5 A DLS THEORY HH Figure 4 a b c Representative intensity vs time for small a medium b and large c size particles Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page2 6 rity Til iT jT DLS THEORY AAL Obtaining particle size from the diffusion coefficient The goal of the DLS
159. s true a value for the viscosity is known for a particular temperature which becomes the basis for choosing the latter The index of refraction on the other hand has only a slight temperature dependence which can be ignored A useful expression which shows the dependence of the computed particle diameter d on the solvent viscosity n and its index of refraction n d constant X n n C 1 where T is the characteristic decay time of the autocorrelation function assuming a single particle size and a single decaying exponential function Equation C 1 may prove to be very useful when a size measurement using a solvent for which values for both n and n are unavailable at the time of the run One may then assume the values for water run the sample and correct the resulting diameter distribution plots later when the correct values have been located All size values e g diameter bin values peak diameter locations mean diameters etc on the plots can be scaled up or down to the correct values using the following equation obtained for Equation C 1 d nw 8ng Ns nw dw C 2 Here nw and ng are respectively the viscosities of water and actual solvent used at the temperature at which the measurement is performed ny and ns are respectively the indices of refraction of water and the solvent The original diameter values on the printouts are denoted by dw the final desired values are given by ds above Nicomp 380 Manual
160. selected displays to the right of the file Selecting colors for 1 5 distributions is especially useful when overlaying more than one sample distribution a Click on the Bar Color Custom colors Define Custom Colors gt gt Soc EM box to access the Color window b Select the desired color for the bars of the distribution by clicking on the actual color box c Click OK d Click OK again once the Change Graph Color window re displays e Select a distribution to view the new color selected for the bar color Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 42 NICOMP SOFTWARE f Click on the Frame Coler box to access the Color window Basic colors a Ge eee mI ee LE m Tan mm MAA Custom colors mummmmmu BEEBE EEE Define Custom Colors gt gt Lox e g Select the desired color for the frame of the bars on the distributions by clicking on the actual color box h Click OK i Click OK again once the Change Graph Color window re displays j Select a distribution to view the new color selected for the bar frame Initialize ND Filter This option initializes the neutral density filter First it locates the zero point of the neutral density filter and then it resets the neutral density filter to the position defined in the Autodilution ND position defined in the Control menu Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page
161. ser may choose this operating mode to conduct the measurement but not save any data The measurement will be made according to the choices made in the Control F3 and Auto Print Save F2 menus of the CW388 software Stop Autocorrelator e Used to stop the collection of data in the autocorrelator Clear Correlator Data Li Clears data and closes any files that are open and displayed in the software Prior to clearing the data a confirmation window will display prompting for action Start Autodilution Starts collecting data using the proprietary Autodilution technique U S Patent 4 794 806 foreign patents pending using this option Click on this tool to start Autodilution The automatic dilution of the sample then begins The PUMP switches to ON LOW SPEED pulling diluent into the unit When the appropriate particle concentration that results in a scattering intensity of 300 kHz Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 13 mir roman AM N NICOMP SOFTWARE Start Measurement The instrument starts taking measurements once this option is initiated Change Weighting A Three weightings may be viewed for a particle size distribution using this option Volume weighted Displays the relative particle volume vs diameter The value of the volume weighted particle size distribution is calculated assuming that the particle are spheres of uniform density which scatter light according to cla
162. serves memory It is recommended NOT overwrite old data files rather copy them to disk and then manually delete them from the hard drive The default choice is set to N No If this choice is not changed then any data saving operation which attempts to create a data file having the same NAME EXT as an existing one will cause the new NAME to be changed by adding the character exclamation point automatically to NAME As a result a new data file will be saved having the new file name NAME EXT instead of NAME EXT If a second attempt is taken to create a new file having the same file name as the original NAME EXT another file will be created this time using the added characters resulting in the new file name NAME EXT This procedure will be invoked yet a third time resulting in the new file name NAMEI EXT After three such substitutions a new data file will no longer be saved Also no error message displays when this occurs Click on the square located next to the selection and a black check mark will display An existing file in the data directory that has the same NAME and EXT values will be overwritten Return to Main Menu After all of the choices have been made and reviewed in this Menu exit to the Main Menu by clicking on OK Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 35 NICOMP SOFTWARE Nicomp Input Menu Manual setting of the NICOMP parameters may be set by specifying the following info
163. sing the CW380 CFG file This is not a mistake Rather after running and exiting the CW380 program a CW380 CFG file will automatically be generated and stored in the disk drive and sub directory which are specified in the Auto Print Save Menu Particle Sizing Window CW388 TBL includes all of the information contained in the Auto Print Save Menu and the Control Menu also accessed from the Particle Sizing Window The former menu includes such parameters as Data Storage Directory Data File Name Printout ID Run Time etc The F3 Menu includes the following parameters taken collectively to define the conditions of data collection printout and storage B Autoset Channel Width B Liquid Viscosity B Sample Temperature B Liquid Index of Refraction In some cases a new CW388 software diskette that is sent will be missing the CW380 TBL file This is not a mistake In the absence of this file a default set of parameters will be used to define The parameters in the two menus that are accessed by pressing the F2 and F3 Menu keys After the CW388 program is run and exited a new CW388 TBL file will automatically be generated and stored in the disk drive and sub directory where the main CW388 EXE program is located Generally it is recommended that the CW388 software files be copied to and run from the hard disk drive The software on the diskette can then be saved as a backup copy to be reloaded in the event of accidental erasure of a file or cr
164. ssian and Nicomp calculations simultaneously m SUMMARY RESULT Run time Data Ch 1 Avg Intensity Sensitivity OHr 6Min 8Sec 612 6K 0 0kHz 150 Gaussian Analysis Solid Particle NICOMP Analysis Mean Dia Std Dev Chi Sq Base Adj Peak1 Peak2 Peak 3 VOL 295 5 nm 53 3nm 3 86 0 00 230 5 345 9 Fit Error 18 03 38 52 61 48 7 349 INT 288 9 nm 52 1 nm 3 86 0 00 230 8 345 3 Residual 18 03 42 91 57 09 0 000 NUM 263 8 nm 47 6nm 3 86 0 00 230 1 345 3 18 03 47 12 52 28 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 56 Gauss Nicomp NICOMP SOFTWARE Displays the Gaussian and Nicomp overlays of the Intensity Volume and Number Weighted data collected for the sample 100 80 60 40 20 0 20 Diam nm gt Intensity Wt Mean Diameter 264 6 nm Stnd Deviation 27 0 nm 10 2 96 Volume Wi Mean Diameter 265 0 nm Stnd Deviation 27 0 nm 10 2 Number Wt Mean Diameter 255 0 nm Cind Massi EI am fin Printout ID ons 261 standard SM GAUSSIAN DISTRIBUTION Intensity Wt GE Volume Wt 80 60 40 20 0 1 Diam nm gt Number Wt Intensity Wt Mean Diam nm Percent 94 Volume Wt Mean Diam nm Percent 96 Number Wt Mean Diam nm Percent 94 10 20 277 9 100 0 256 4 100 0 224 2 100 0 50 100 200 500 1K Solid Particle Show Distributions Returns the user to the standard distribution
165. ssical Mie Theory Number weighted Displays the relative number of particles in a sample run vs diameter The value of the number weighted particle size distribution is also calculated assuming that the particles are spheres of uniform density which scatter light according to classical Mie Theory Intensity weighted The result first displays from either of the autocorrelation functions Displays the relative Intensity of scattered light vs diameter for a sample run Toggle Gaussian Nicomp Distribution RES ot Toggles between the Gaussian and Nicomp Distribution for a sample file Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 14 rite T T NICOMP SOFTWARE AIA Toggle Solid Vesicle Particle The instrument defaults to solid particle weighting when power is first applied The weighting may be changed using this option The weighting will change when the results of a new analysis is displayed at which time the title will also change to Gaussian Analysis vesicles The original weighting can be re displayed by using this option again Increase Intensity Can be used to increase the sensitivity to achieve the optimum Photopulse Rate Decrease Intensity Can be used to lower the sensitivity to achieve the optimum Photopulse Rate Print the Active Document This option is used to print the appropriately weighted distribution displayed Prior to selecting the print option pressing W will toggle among the d
166. t NumberWt Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 10 APPENDIX AUTOCORRELATION FUNCTION Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm 7 3 DataFile CProgramFilesiParticle Siang SystemsiNicomp388Wwersion 1 68 Bimodal 300 Autocorrelation Function GAUSSIAN SUMMARY Mean Diameter 2889nm Variance P 1 0 033 Sind Deviation 52 1 nm 18 096 Chi Squared 3 861 Norm Sind Dev 0 180 BaselineAcj 0 000 Coeff of Var n ZAvg Diff Coeff 1 61E 008cm2s RunTime 0 Hr 6 Min 8 Sec Wavelength 632 8 nm CountRate 0 KHz Temperature 23 degC Channel 1 6126 K Viscosity 0 933 cp ChannelWidth 35 0 uSec Index ofRef 1 333 counts X1000 AUTOCORRELATION FUNCTION 610 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page A 11 APPENDIX A AUTOCORRELATION DATA Particle Sizing Systems Inc Santa Barbara Calif USA 350nm 220nm7 3 DataFile CProgramFilesiParticle SiangSystemsiNicomp388Wwersion 1 681Bimodal 300 Autocorrelation Function GAUSSIAN SUMMARY MeanDiameter 2889nm Variance P 1 0 033 Sind Deviation 52 1 nm 18 096 ChiSquared 3 861 Norm Sind Dev 0 180 BaselineAdj 0 000 Coeff of Varn ZAvg Diff Coeff 1 615 0080m2 RunTime 0 Hr 6 Min 8 Sec Wavelength 632 8 nm CountRate 0 KHz Temperature 23 degC Channel 1 6126 K Viscosity 0 933 cp ChannelWidth 35 0 uSec Index ofRef 1 333 counts X1000 AUTOCORRELATION FUNC
167. the ability to zoom into the distribution to examine the fine details of the distribution Close will close this option and return to the CW388 Software Window CW388 Version 1 68 NICOMP Particle Sizing Systems EZ 73 Parbcie Sing Systems ine Sania Barhara Cal UBA Data Fie CxProgram Flas Parice Siting Systemeltieomo 388 Emsdal 300 HLLENMIY eishled QAU NMAN ATAMU TON Argira oid Pariet GAL GIAN MAMMARY Moan Diametor 2889 nm Variance P 1 0 033 Sing Deviation 32 1 em t6 ON Cn Squared 3 001 Norm Stnd Dev 0 160 Baseline Adj 0 000 Coert of vera Z Arg DIT Coen 1 616 000 mara MITENS WT CAU AE DAT RH TON Bimoga 300 Cometaere Menuet 23 of distribution lt 50 of dutribution lt 73 N of gisirioution 20 N of gratrioution 99 o distribution S0 N of alricuton Run Time Count Rate 20 o Channel t 264 35 Channel Width 2316 0m 284 nm 320 9 am 398 0 am 4322 0m 330 7 am Hr 6 Min 8 Sec Waveiengm Kur Temperature 26 K Viscosity 0 uSer Index of Rat Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 9 NICOMP SOFTWARE Print Setup This option allows for the setup of the type of printer to be used the orientation of the printout and the size paper to be used Print Setup Printer Name KONICA MINOLTA C350 PCL5c v Properties Status Ready Type KONICA MINOLTA C350 PCL5c Where IP_192 168 0 199 Comment
168. the next given such a small average value If the instantaneous photon rate were to follow Poisson statistics we would expect the rms standard deviation of the number of pulses per time interval to equal N where N is the average number For our example above this gives a standard deviation of 1 7 Hence from purely a statistical point of view we expect the intensity per 10 microsecond time interval to vary from 0 to 5 photopulses with occasionally a 6 7 or larger independent of the effects of diffusion This is simply a consequence of our having chosen a very short time interval relative to the average photopulse rate When diffusion is added to the process the resulting fluctuations in l t become even more pronounced The resulting integer numbers of photopulses per small time interval are of course the values of I t and I t t in Equation 3 which become multiplied together digitally to compute the values of C t A representative sequence of photopulses is shown in Figure 7 We have subdivided the time base t into intervals of equal width At equal to the channel width of the Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 13 DLS THEORY autocorrelator Here the instantaneous intensity I t is defined as the number of pulses in the interval At which lies closest to time t Over each interval we have recorded the instantaneous intensity for that interval simply the number of photopulses produced by the
169. the normal scattering angle for the basic Nicomp system The external angle can be changed allowing the system to compute the resulting actual scattering angle which depends on the geometry of the scattering cell The external angle is made accessible by making the appropriate change in the System Setup Menu The basic operation of the NICOMP is defined using the following set of parameters Autodilution Drop in Drop in Cell Used to make a measurement on a sample that is already at a good concentration using a drop in cell If an attempt to use Autodilution while the Flow Pump Option has NOT been selected in the System Setup window the option will remain selected and the following warning will display Warning Flow pump is not installed Use Drop In Cell only Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 27 T ri Ts roman AM N NICOMP SOFTWARE Flow Cell The process by which a concentrated sample is diluted to achieve the proper concentration required to make a measurement Interactions between particles in concentrated samples can lead to significant error Manual dilution can help eliminate this problem but it is often time consuming and can have a definite effect on sample reproducibility The Autodilution system can automatically dilute the sample in the flow cell to optimum concentration thus eliminating time consuming trial by error manual dilution Important If a change is made from Autodil
170. the result just discussed This is shown in Figure 18d As taught earlier this is the fundamental result which emerges from the Distribution Analysis all of the more useful weightings are derived from Figure 18d using the rules of light scattering Equation 16 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 60 ME T T T DLS THEORY AAL NICOMP Distribution Analysis Solid Particles 2 Fit Error 1 19 Res idua 0 0 Gaussian Chi Squared 24 7 Data 1735 8 K a e a a 1 e 4 4 1 t 1 4 5 3 t 4 t Diameter log o Diam nm INTENS Figure 18d The intensity weighted Distribution Analysis result for the test bimodal corresponding to Figure 18c We now can appreciate why the test sample was deliberately skewed at the start toward the smaller diameter particles Even though there is three times as much latex mass or volume contained in the 91 nm component compared to the 261 nm size fully three fourths of the total scattered light computed by inverting Equation 14 is produced by the 261 nm fraction Had we attempted to analyze a simple 1 1 mixture of the two sizes the algorithm would still have found a bimodal distribution but would have had a harder time correctly establishing the mean diameters of the two peaks especially the weaker one 91 nm Although we appear to have obtained the excellent results of Figures 18a d with relative ease th
171. ticles We find here that the 91 nm particles represent over 98 of the total number with the 261 nm species accounting for only 1 5 The resulting plots on a linear scale lose accuracy because of this very strong skewing toward the smallest particle peak in the distribution Figures 19a b and c show the detailed printout summaries for the three weightings corresponding respectively to the intensity volume and number weighted distributions It should be evident from our earlier discussion of the motivation behind the ILT algorithm that the closer the spacing of the two peaks in a bimodal the more unlikely it is that the Distribution Analysis will produce the right answer As seen in Figure 16 even when there is a 3 fold Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 62 rity Til iT jT DLS THEORY AAL separation of the two sizes the autocorrelation function bears a close qualitative resemblance to that obtained from a broad unimodal distribution In practice the NICOMP ILT algorithm is able to resolve a bimodal having a size separation of 2 1 or even somewhat closer provided the sample is clean i e largely free of aggregates and other large particle contaminants Of course this capability presupposes that there is a reasonably well balanced intensity contribution from each species In general the narrower the separation the more data are needed to permit unambiguous resolution of the bimodal including a large
172. ting value baseline B In effect the value of t describes the characteristic lifetime or duration of a major bump or fluctuation in the scattered intensity I Hence the larger the particles the slower the diffusivity and resulting fluctuations in and the longer the decay time constant t As you might have predicted by now we are able to obtain the diffusion coefficient D of the particles from the decay constant z the precise relation is Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 11 TT T t Tlr lr AM N DLS THEORY 1 1 2DK 8a or D 1 2K2 1 t 8b Here the quantity K is called the scattering wave vector It is a constant that depends on the laser wavelength A in the solvent and the angle 0 at which scattered light is intercepted by the PMT detector 0 90 for the DLS MODULE In effect K acts as an absolute calibration constant which relates the time scale of the diffusion process to the distance scale set by the laser wavelength making interference possible Constant K is given by K 4nn A sin 0 2 9 where n is the index of refraction of the solvent e g 1 33 for water In the case of the DLS Module with 0 90 and A 632 8 nm K equals 1 868 X 10 cm The rationale for particle sizing using the method of DLS should now be clear We detect scattered light at a fixed angle produced by an ensemble of particles suspended in a solvent The intensity fluctuates in time due to
173. tion definition and motivVatiON ooonnnnninnnnnncnnnnnnnnnncononcncnnnnnnnnnnnnann nono 8 Ideal case uniform particle SIZ8 ooooococcccccccnncconcconononnnnnnnnnnnnonnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnns 11 Photon counting and digital autocorrelation fUNCtiONS ooooonnnononccccnnnnnccnnanananncnnnnnnnnnnns 12 THE SIMPLEST APPROACH TO SIZE DISTRIBUTIONS GAUSSIAN ANALYSIS seeee 16 Uniform particle size trivial analysis 040 nnnnnnnnannnnnnennnnnnnnnnnnnennnn nenn 16 Broad unimodal distribution Gaussian Analysis sen 21 Effects of weighting in the Gaussian Analysis eeeeeeeee 30 Importance of acquiring data of sufficient accuracy cccccconnnoconcccnnccccnnnnananonononnnnnnnnnnn nano 36 NIGOMP DISTRIBUTION ANALYSIS He ei fiere een 44 INITIAL HARDWARE SETUP co condicio ia SECTION 3 SOFTWARE INSTALLATION bocce eee a RED EE dsc SECTION 4 NIGOMP SOFWARE diosss iris SECTION 5 FEB A AE 1 DO O 3 Read Ne A A A A AA ee 4 SAV C ETE 5 SAVE ASC ll S dn detur eto E o Roa eoo A adt ics doo totiens 5 VNo O M BIE S r ringen cidad EA ERN 6 Sublract Data POW st AA 6 O cn TTE 7 Print Preview E 9 O reise roseo sive etna iae ie eie rade te anre rese dees tes ea eae E T T 10 Nicomp 380 User Manual PSS 380Nicomp 030806 11 06 Pagei TON 21 1 AR ERRFERELECFERFETFETECWERRTREREETERTINENFETENPETTLPESPERRLTEFETESTERRRRET FIREOHTELTERERFEHERSELESILTETT
174. to be clearly seen Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 18 mene DAL d ERA y PES ITem ly oe So Wasser PADE T0 PREGOHOHEE PORE EOD EUN UNO ot led eee ied Lm llo al iima FEERHER AI OIR PANI EITTERA LODA PANTA IR MARFIL HUTTE RR ei LLL CC POSE TEER CANA ENG SHA ETE ERESELL EGA CARGO LENG LIGA ALIENI BELL ELLE EURE DAUER Rite C MEME A AA E SERAA DONA LADA ROBBE AUCAT TEIN ERE E DRS ERAN GARFE MTM HELL RIA COL LARTER MEER SEDAN 108 90 99 500 00000 TODAS 00 M DADOS DA OGRE SODA AO me enden JLS ce aro rannte rump UNUESEPE ECUICPIHERUIEEPEEEEL UEBER LLL LEDPZLHUBDDNGREIIDNLLCDLNBELNURBBESSBILBZCSELL angus rH mpi n En tnb EemmmENCSEHEE TET ET TRTTETRTET ETC CEST A A AERE MM UM CUELEEOESD UDOSUEE2ON NEGECEESNS DOSEN DOSOD QUUURUAGI FUERAT UUGRELALELULUR IA VU LU LR VACA CDI UEEELVUUELULLLLALLELULCOUETS PROA DIOS PEPA Ona RRA MARI LU LU EAR OGG LM AREA eT ERA CARLA LEA ARO MA SS CEBACOANGU SAGAS E0002 CASES ROAON MANNI MEM UFHLHEH HIM HLUIUE DH LIUM EU EI e FID IIS rH teeta oa At Gat ae et ee RR RR E aati NORRADOIA UURA LALIT UROL ULM REM ADA MUA DELP MAR AMEND ME mirre FUEGO 1OUDUNEGES CXBAOCDEES SBCOLLEUGN HALE BERE ARAS UAC teen Aaa Lal ES RR EI RA ar UN EDOM LETRA RALAG DENT ONIH KHH mi um lu li deeem HIHHIHIBH HELEDUUHLIEHNHE TNT ESPERE ET EE BEHIE DEGRA PUEUL DEES MEE ELLE LLL NIHIN DI BRASIL PNEU LLLA REM LLLA ENTUM DIOS EE ELI HL 014900018901 MAR LEE HELL UU LUUD ot et mni Figure 8c Log C t
175. to the below C380 Control Menu Menu File CAPSS Software cw388 version 1 68Cw388 tbl Drop In Cell 23 E Ug res Flow Cell Liquid Viscosity 0 933 CP Iv Autoset Channel Width Liquid Refractive Index 1 333 V Autoset Sensitivity Intensity Setpoint 300 KHz IV Auto NICOMP Parameter First Channel Used 2 M Auto Baseline Adi Laser Wavelength nm Cum Set Pt leo Ext Fiber Angl Bet eae 30 9 9 autodilution ND Position 100 Scattering Angle deg Cancel NOTE The laser wavelength may differ depending on the instruments hardware setup 6 Click on OK to return to the CW388 Software Menu 7 Prepare the sample by gently inverting the bottle of latex material ten times 8 Add five drops of the concentrated submicron latex standard to 25 mL of distilled filtered water The resulting mixture should be manually agitated to provide a uniform suspension 9 Manually agitate the sample to provide a uniform suspension This is confirmed by visual inspection of the sterile beaker The suspension should be turbid slightly white in color 10 Draw approximately 3 mL of this turbid suspension into a sterile 3 or 10cc syringe 11 Connect syringe to the luer fitting on the top of the injection valve 12 Gently apply manual pressure to provide a finger tight connection 13 Point the valve handle down with the flow pattern on the handle indicating that the fluid flow path into the Nicomp is from the source
176. trol Menu determines the conditions under which raw data will be collected Access to the Control Menu is gained using this option This is one of two menus used to define the basic operation of the Nicomp The other is the AutoPrint SaveMenu 1 Position the highlight bar over the Control Menu option and click once The Control Menu will display C380 Control Menu Menu File CAPSS Software cw388 version 1 681Cw388 tbl Drop In Cell t 23 C Dons C Flow Cell Liquid Viscosity 0333 CP Iv Autoset Channel Width Liquid Refractive Index E 333 V Autoset Sensitivity Intensity Setpoint 300 KHz M Auto NICOMP Parameter First Channel Used 2 IV Auto Baseline Adj Laser Wavelength nm Cum Set Pt Bo External Fiber amp ngl C PM ANS 90 des iodiuionND Position 100 Scattering Angle deg x 7 2 Press TAB to advance to each of the parameters on the screen MenuFile Specifies the file name under which this table of parameters will be saved Channel Width The channel width of the digital autocorrelator in the Nicomp is reported in terms of microseconds or usec In most cases the system should be allowed to set the channel width automatically Ideally the channel width will be adjusted so that the number of decays in the autocorrelation function lies between 1 7 and 2 7 Bypass of the automatic adjustment of the channel width can be made allowing a value to be entered manually This option is preferred when similar samples are goi
177. ts for each fit A good illustration of the typical settling time of the Gaussian Analysis is provided by a tabulation of the results obtained for the IV fat emulsion discussed above A new computed distribution was obtained approximately every 45 seconds whenever a fresh set of analysis results was displayed on the video terminal a summary of these results is shown below in Table 1 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 36 TT rir Tlitt DLS THEORY m Data Run Int Wt Std Dev Vol Wt Chi Sq Base Ch 1 MeanDiam Mean MeanDiam Adj 30K 229 6 nm 35 206 5 nm 0 7 0 0 X 92 228 4 35 203 9 0 2 158 227 1 36 200 5 0 1 i 209 227 1 35 201 7 0 3 261 227 2 34 205 0 0 4 311 227 4 32 207 3 0 8 361 227 3 32 207 7 1 2 411 227 7 32 208 6 2 3 462 228 0 31 210 3 22 513 228 0 30 211 0 2 6 564 227 8 30 211 3 2 8 614 227 3 30 210 1 665 227 1 30 210 6 2 2 716 226 7 30 210 3 2 0 167 226 4 30 209 8 1 8 817 226 3 30 209 5 1 4 868 226 4 30 209 0 920 226 4 30 208 9 1 5 972 226 3 30 208 9 1 2 1023 226 2 30 209 0 1 2 1047 226 1 30 209 1 1 1 Table 1 Time dependence of the Gaussian analysis results for a typical unimodal sample IV fat emulsion Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 37 T mir AM DLS THEORY The results shown in Table 1 for a well behaved fat emulsion nicely illustrate some of the characteristics of the Gaussian Analysis discussed earlier First we notice that the parameter
178. ual has returned to its ideal value of 0 0 Chi Squared has grown to 24 7 which unambiguously rules out acceptance of the Gaussian Analysis result The mean diameters of the peaks are now at 94 2 and 283 4 nm While these values are too high by a few percent a closer examination of the printouts shown below reveals that the error represents diameter displacements of less than one diameter slice for each peak This is an exceptionally good performance given the difficulty of this test sample Indeed the alternative ILT algorithms being used today generally perform much less well when faced with a control sample like this one We have deliberately showed typical results obtained by the DLS Module for this 3 1 91 261 nm test bimodal after 20 30 minutes running time on occasion even better accuracy has been attained Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 59 T mir iman AM N DLS THEORY NICOMP Distribution Analysis Solid Particles 1 Fit Error 1 19 Residual 0 0 Gaussian Chi Squared 24 7 Data 1736 8 K a 1 r b 5 t t t a 1 1 4 a r 4 t H y 1 t gt Diameter log 000 Figure 18c The volume weighted Distribution Analysis result for the test bimodal after Data 1736K 23 mm Now it is instructive to show a plot of the intensity weighted distribution e the set of weighting coefficients f in Equation 14 which corresponds to
179. uire no attention The photopulse preamplifier discriminator has been preset to provide optimal output pulse characteristics and requires no periodic adjustment The thermoelectric temperature regulator has been calibrated at the factory and requires no routine attention SYSTEM MAINTENANCE Preventive maintenance for the Nicomp tubing and fluidics can and should be performed on a daily basis MAINTENANCE Preventive maintenance for the Nicomp Submicron Particle Sizer can and should be performed on a periodic basis 1 Flush the system adequately to achieve the same counts less than 10 mKz to display on the front panel of the instrument 2 Visually inspect the external tubing on a weekly basis for deterioration and signs of wear If any parts of the tubing are cracked or show signs of excessive wear they should be replaced 3 Visually inspect the tube section passing through the peristaltic pump for discoloration or any signs of wear Service related to the Laser associated optics or the sample cell holder must be performed by the factory or a representative of Particle Sizing Systems Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 7 1 VOLUME WEIGHTED GAUSSIAN 350nm 220nm 7 3 GAU SSIAN SUMMARY Mean Diameter Stnd Deviation Norm Stnd Dev Coeff of Var n Diam nm gt Bimodal 300 Cumulative Result 25 of distribution 50 of distribution 75 of distribution 90 of distribution 99 of distri
180. uld be saved under or position the highlight bar over the existing table file name to be updated and click once The parameters set in the Control Menu and Auto Print Save menu will be saved for future use Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 40 NICOMP SOFTWARE Control Buttons Control Buttons Inc Scattering Intensity ALT 2 Dec Scattering Intensity ALT 1 Inc Scatt Angle by 0 9 Deg CNTL F4 Dec Scatt Angle by 0 3 Deg CNTL F5 Laser Power On CNTL F6 Laser Power Off CNTL F8 Close Inc Scattering Intensity Increases the scattering intensity of the detector Dec Scattering Intensity Decreases the scattering intensity of the detector Inc Scatt Angle by 0 9 Deg Increases the scattering angle of the detector by increments of 0 9 degrees Dec Scatt Angle by 0 9 Deg Increases the scattering angle of the detector by increments of 0 9 degrees Laser Power On Applies power to the laser Laser Power OFF Powers down the laser Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 41 NICOMP SOFTWARE Change Graph Color Change Graph Color File 1 1 Data MENE Frame Color File tt 2 MS Bar Color Frame Color File 3 MB ar Color Frame Color File 4 Mmm Bar Color Frame Coler File 5 BE Bar Color Frame Color A window will display all of the colors available that can be used for the bar color of the distributions of the sample runs The new color once
181. ult for the 91 1091 sample Figure 20 nm bimodal For this particular sample preparation we see that the first two results agree very closely with the known situation indeed more closely than could normally be expected for these short data acquisition times After an elapsed time of 42 minutes Figure 22c the results have actually degraded somewhat although the peak mean diameter values are still within a few percent of the correct answers At this point the Fit Error has fallen to 1 0 with an ideal Residual of 0 0 which easily fits our criteria of a finished run When we return 8 hours later Figure 22d we find a nearly perfect result as regards both particle diameter and volume for the two peaks Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 69 DLS THEORY NICOMP Distribution Analysis Solid Particles SIZE nanometers log scale REL VOLUME RESULTS Distribution Analysis 3 7 o Peak 1 4 3 Diameter 30 9 Volume 70 9 Peak 2 Diameter 266 2 Volume 29 1 27 o ed SS 11 Fit Error Residual 0 0 Mean Diameter 143 0 nm Standard Deviation 8 nm Gaussian Analysis Volume Weighting Mean Diameter 189 7 nm Standard Deviation Chi Squared 4 9 Baseline Adjust 0 00 X Data 405 5 K Scale Parameters MIN DIAM 10 SMOOTHING 3 RUN TIME AVG COUNT RATE CHANNEL WIDTH TEMPERATURE VISCOSITY INDEX OF REFRAC PRINT AT DATA OF PRINTOUTS PLOT SI
182. ut of volume weighted Distribution Analysis result for the 3 1 71 91 261 bimodal sample after 10 min essem 71 Figure 22c Printout of volume weighted Distribution Analysis result for the 70 3 1 91 261 bimodal sample after 42 min 72 Figure 22d Printout of volume weighted Distribution Analysis result for the 3 1 73 91 261 bimodal sample after 8 hrs 10 min cocoocnnnncnnnnnoncnonencnnonnnnnnnonononenennnnnonononancnnnnnnnnnns 73 Nicomp 380 User Manual PSS 380Nicomp 030806 11 06 Page vii ME T T T GENERAL INFORMATION JAJA REGISTRATION Please register your software by taking a moment to fill out the registration page provided In keeping with our promise we can easily provide two years of free software upgrades Just call us if you need information about our other products or information about upgrading your existing system TECHNICAL SUPPORT If technical support is needed please contact one of the following offices Particle Sizing Systems 8203 Kristel Circle Port Richey FL 34668 Tel 727 846 0866 Fax 727 846 0865 Or Particle Sizing Systems 201 Woolston Drive Ste 1 C Morrisville PA 19067 Tel 215 428 3424 Fax 215 428 3429 Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 1 1 GENERAL INFORMATION SAFETY CONSIDERATIONS The NICOMP and Autodiluter Submicron Particle Sizer is certified to conform to the app
183. ution mode to Drop in cell and the flow cell remains selected the pump will initiate and spill water into the system Likewise if the drop in cell remains selected and the pump is required to start running Ctrl F will not cause the pump to start running Autoset Channel Width This parameter is used to establish whether the channel width of the autocorrelator will be set automatically during a measurement which is initiated by clicking on the Autodilution icon or the Green G The channel width can be set manually The ideal channel width will produce a 1 7 2 7 decay in the correlation function Autoset Sensitivity This parameter is used to establish whether the sensitivity of the PMT detector will be adjusted automatically during a measurement If this option is selected the sensitivity is set automatically to reach 300 kHz or whatever parameter was set in the Intensity Setpoint field of the Control Menu Auto NICOMP Parameters This parameter enables the system to automatically set the range of parameters for the NICOMP Distribution Analysis or to permit these parameters to be preset for all subsequent measurements If the Automatic NICOMP Parameters option is selected the internal system computer will attempt to optimize the parameter choices However if this option is not selected the parameters set in the Nicomp Input Menu will be used for analysis Please refer to the Nicomp Input Menu section of this manual for details about
184. utions The default value at the start is 3 which represents a reasonable compromise until the STD DEV and CHI SQUARED have revealed more about the probable nature of the particle size distribution This may then suggest a better choice for the Smoothing parameter For Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 36 ME T T T NICOMP SOFTWARE JAIL example in the case of a single narrow distribution of polystyrene latex particles a smoothing value of 1 2 or 3 may be used Also for a mixture of two different latex particles optimal separation of the size peaks especially for a very close spacing a factor of two or closer is best achieved using Smoothing 1 2 or 3 For a broad distribution such as an oil in water emulsion made by homogenization or milled powder a Smoothing value of 4 to 6 should be used Otherwise lower values of Smoothing will tend to produce peak splitting leading to anomalous results like false bimodals If no prior information concerning a sample is known i e whether it consists of a broad distribution of particle sizes or is a bimodal the results of the Gaussian Analysis can be used as a guide in selecting the most appropriate value for the Smoothing The fit autocorrelation function by definition is a unimodal distribution i e approximate log normal The values of the STD DEV and CHI SQUARED serve as guides to help decipher whether to use a low Smoothing value or a high Smoothi
185. value of the Gaussian Chi Squared Even for a widely spaced bimodal however one must always keep in mind the practical requirement that the weakest peak contribute adequate intensity The rules of light scattering as summarized by Equation 16 will alert one to the possibility that a bimodal cannot be measured NOT because of insufficient volume fraction for a given size component typically the smaller size but because that component contributes only 1 or less to the total scattered intensity As a general rule of thumb we can say that a second or third peak will be reliably measured only if its relative intensity is at least 1 2 Hence whenever one is using the Distribution Analysis it is usually a good idea to obtain a printout of the intensity weighted result as well as the desired volume weighted plot Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 2 63 DLS THEORY NICOMP Distribution Analysis Solid Particles 3 Bimodal latex sample 3 1 91 amp 261 nm nanoweters log scale REL INTENSITY l i t t t l l l l l i l t t I t Scale Parameters MIN DIAM 10 PLOT SIZE 45 SMOOTHING 3 RANGE 100 RUN TI ME Hours 22 Mins 50 Secs AVG COUNT RATE z 0 306 4 kH CHANNEL WIDTH 22 0 EMPERATURE T 23 VISCOSITY 9 933 INDEX OF REFRACTION 1 333 COUNT RATE SETPOINT 300 Molec Wt 1 0 Molec Wt 0 50 USEC DEGREES CENT CENTIPDISE kHz E 4 ALPHA BETA
186. wanted characters or press Ctrl and Y to delete the entire 80 character string The caption is composed of an 80 position alphanumeric string The caption will display exactly the way that it was typed 4 Press ENTER when the editing is completed and the caption reads as desired The new caption will be entered into memory Auto Operation Options No Print Save Cycles Use this parameter to establish the number of Operation Cycles which will occur after the start of the measurement process In the example of the Auto Print Save Menu 2 Cycles were chosen The cycles can be defined in two different ways Nicomp 380 Manual PSS 380Nicomp 030806 06 06 Page 5 31 TT JT T MA AM N NICOMP SOFTWARE Using Run Time The first and most convenient way in which an Operation Cycle can be defined is through the Run Time 1 Click on the circle position to the left of the Using Run Time selection A black circle will display in the circle Enter the parameter in terms of minutes This will define a single automatic Operation Cycle A Run Time set to 5 minutes means that after approximately 5 minutes of data acquisition the firstcycle will be completed and all of the Operating Functions which have been specified in the Auto Print Save Menu will be implemented If the No Auto Operation Cycles listed in the AutoPrint Save Menu is larger than one another cycle based on the Run Time will begin This process will continue for the
187. y necessary to sample the value of l t very frequently i e to choose small values of t between sampled pairs in order to obtain an accurate autocorrelation function which is sensitive to rapid changes in ls caused by rapid diffusion of the particles For this reason it is therefore necessary to define ls in terms of the photopulse rate using a very small unit of time In this way the measurement of I can be made as frequently as necessary and approaches being an instantaneous value For example when 100 nm 0 1 micron particles are measured by the DLS Module the sampling of I t is performed approximately every 10 microseconds In this case therefore l t is arbitrarily defined to be the number of photopulses which occur during a given 10 microsecond interval This short a time interval or smallest increment in t is needed to compute the relatively rapid decay of C t versus t which occurs for these rapidly diffusing small particles Of course for smaller particles an even smaller unit time interval would be needed to define I t Two observations should immediately be evident First given such small time intervals used to define l t the resulting number of pulses must be very small Consider our example of a typical average photopulse rate of 300 000 per second this corresponds to an average instantaneous intensity of 3 pulses per 10 microseconds Second we should expect this number to change greatly from one time interval to

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