The OD600DiluPhotometer™ User Manual
Contents
1. The OD6OO DiluPhotometer User Manual Version 1 3 phone 49 0 89 726 3718 O Fax 49 0 89 726 3718 54 Email info implen de www implen de INEL CH OD600 DiluPhotometer User Manual Implen GmbH Schatzbogen 52 D 81829 Munich Declaration of Conformity This is to certify that the Implen ODGOO conforms to the requirements of the following directives 13 23 EEC amp 89 336 EEC Standards to which conformity is declared EN 61 010 1 2001 Safety requirements for electrical equipment for measurement control and laboratory use EN 61326 1998 Electrical equipment for measurement control and laboratory use EMC requirements Signed Dated September 1 2008 wail A Ly Dr Thomas Sahiri Managing Director Implen GmbH Version 1 3 Page 2 14 INEL CN OD600 DiluPhotometer User Manual TABLE OF CONTENTS 1 ESSENTIAL SAFETY NET E eegEesgieeteeengeteee ege eeh 4 1 1 Unpacking Positioning and Installation ccceeeseeeeesseeeeeeseeeeenseeseeesseeseeaseseeseeeeeenseesoenseesonenenes 4 2 INTRODUC TION D 5 21 YOU Speco pHOtOMETE aag AEEA AAE Ea EAA 5 22 ampie Handling TDS EAE E a EAA EA 5 Bed OVD AGAIN RE e EE 6 DESEN 7 31 DIC SCH UO E 7 32 el EN engl E e sssini aa aaa a a aa E i 7 Sid OPECINCANONS ranar aA S a 8 A OD600 DILUCELL APPLICATION s sssssssssssssesessesessssrscsessesesssacsessescsesaeseseacsesaescsesansesesaesnsoraeseeaeatsess 9 4 1 Bacterial Cell Culture2. Print Print results stored in memory Display There is a memory number indicator and a battery indicator Version 1 3 Page 6 14 INEL CN OD600 DiluPhotometer User Manual 3 DILUCELL 3 1 Description DiluCell is especially designed for the usage of the OD6OO DiluPhotometer for the analysis of samples at a wavelength range of 340 nm to 950 nm Due to the reduced pathlength DiluCell provides a virtual dilution without the need of a physical dilution of higher concentrated samples The two different available versions DiluCell 10 and DiluCell 20 allow an automatic 1 10 and 1 20 dilution of the sample The required sample volume is 200 ul or 100 ul respectively The following scheme explains the functionality of the DiluCell technology Lambert Beer Law I Abs log exexd Abs measured absorbance value on your instrument screen incoming light outgoing light extinction coefficient concentration pathlength standard 1 cm OO OD IMPLEN DiluCell 3 2 Operation Instructions Step 1 Place the DiluCell into the cuvette sample compartment Ensure the correct positioning of the DiluCell The light beam of the ODGOO is directed horizontal from the front of the instrument to the back Important Note DiluCell will not work in any other orientation do not mix other cuvette types within a DiluCell measurement series for blank etc Step 2 Pipett
3. This enables readings to be taken at for example an incubator and downloaded to a PC in a different laboratory The results remain in the memory even when the instrument is switched off Step 1 Step 2 Step 3 Step 4 Step 5 Step 6 Step 7 Step 8 Step 9 Switch the instrument on by pressing the ON OFF button Press mem button to display MEM if not already displayed Place a reference into the cuvette compartment Press the blank button The display will show 0 00 but the memory number will not change Insert the sample and press the sample button The result will be displayed and automatically stored and the Memory Number will increase by one To retrieve the results press recall print This will print out all of the results held in the memory if the instrument is connected to a PC or printer and cause the memory number to flash Repeated pressing of the button will display the results in the memory on the screen in reverse order scrolling back to the beginning Press reset or mem to go back to the latest result Pressing reset when the latest result in the memory is showing will cause the screen to flash rt and d If no further action is taken the screen will revert to its normal state after 7 seconds If reset is pressed again whilst the screen is flashing all of the memory positions will be cleared Important Note The instrument can store up to 99 readings Version 1 3 The hundredth reading will automatically overwrite
4. continuous development we reserve the right to alter specifications without notice The product does not fulfil the specific requirements of the IVD Warranty e IMPLEN guarantees that the product supplied has been thoroughly tested to ensure that it meets its published specification The warranty included in the conditions of supply is valid for 12 months only if the product has been used according to the instructions supplied IMPLEN or your supplier can accept no liability for loss or damage however caused arising from the faulty or incorrect use of this product Version 1 3 Page 14 14
5. number using an alternative technique for example microscope slide method and converting to OD 600 using the rule of thumb that 1 OD 600 8 x 108 cells ml for E coli Clumping together of cells will also affect readings so the medium they are suspended in will also make a difference 4 1 2 Operation Instructions Making a measurement Step 1 Switch the instrument on by pressing the ON OFF button Step 2 Place a reference into the cuvette compartment Step 3 Press the blank button The display will show 0 00 Step 4 Remove the reference and replace with the sample solution in a cuvette Step 5 Press the sample button The display will show the OD of the sample in absorbance units Important Information If using DiluCell remember to multiply the reading from the OD6OO after the measurement with the proper virtual dilution factor Virtual dilution factor are 10 for DiluCell 10 20 for DiluCell 20 Multiple samples can be compared with the same reference by placing different samples in the cuvette chamber and making measurements for each one It is recommended to re reference with the reference solution every 10 to 15 minutes to avoid any slow instrument drift Please re reference always if in doubt Version 1 3 Page 9 14 cd OD600 DiluPhotometer User Manual Using the memory function The instrument can store up to 99 readings in the memory The results can then be viewed printed or downloaded at a later time
6. transit If any damage is discovered please inform your supplier immediately Ensure your proposed installation site conforms to the environmental conditions for safe operation Indoor use only Temperature 5 C to 35 C Maximum relative humidity of 80 up to 31 C decreasing linearly to 50 at 40 C The instrument is powered by the internal rechargeable battery or by mains electricity using the supplied power adapter Using the instrument with the mains adapter will automatically recharge the battery The battery will last approx 1 month when fully charged with normal use A full battery recharge will take approx 12 hours overnight Please read through this user manual prior to use Please contact your original supplier in the first instance if you experience technical or sample handling difficulties If this equipment is used in a manner not specified or in environmental conditions not appropriate for safe operation the protection provided by the equipment may be impaired and instrument warranty withdrawn Version 1 3 Page 4 14 INEL CN OD600 DiluPhotometer User Manual 2 INTRODUCTION 2 1 Your Spectrophotometer Your OD6GOO is a small easy to use instrument that is dedicated to measuring samples at a wavelength of GOO nm It is suitable for measuring growth rates of all types of cell e g E coli and yeast Combining ODGOO with DiluCell 10 or DiluCell 20 facilitates measurements with low sample volumes of 20
7. 0 ul and 100 ul respectively and a virtual sample dilution of either factor 10 or 20 With a bandwidth of 40 nm the OD600 enables also measurements nearby 600 nm like the protein quantification assay according to Bradford 595 nm A 600 nm LED source in combination with a fibre optic is used to obtain the measurement The instrument can be used in incubation cabinets and under anaerobic conditions 2 2 Sample Handling Tips e Note that the light beam is directed horizontal from the front of the instrument to the back therefore please ensure the cell is inserted in the correct alignment e The optical height is 8 5 mm e The cell holder supplied with the instrument accepts SS tier virtual Dilution Adapter pue 40 200 4l DiluCell 20 100 pi Macro cuvette 10 mm 1ml he max fill volume 4 5 ml Semi micro cuvette 10 mm 0 5 ml max fill volume 1 4 ml 10 mm diameter tubes 12 mm diameter tubes 16 mm diameter tubes Depends on tube being used IMPORTANT WARNING Always wear protective clothing when handling bacteria or other cells Version 1 3 Page 5 14 INEL CN OD600 DiluPhotometer User Manual 2 3 Keypad and Display OD600 LCD Display Recall Print _ cai print Reset mem Memory AN _ Blank Reference Sample On off key we ooo p owopse key owopse Turns the instrument on off Set reference to 0 000 OD at 600 nm on a reference Recall
8. Measurement ccccecceessseeesceensseeeeceeasseeceeasseeeeeonaaseeeseonanseesseoeaseeseoesonas 9 Aa Rtl e NEE 9 412 Operation INStrUCHONS vessccasaehcocesae lt deavasdcndenansacadecoscudessasd cadence dadestaciconevaessdeaverssedpssesaceeacosdeds socneiua 9 5 OD600 STANDARD CUVETTE APPLICATION s ssssssssessssssesessssesesessssescsesssatscsesseseansesesseansesssanaeseeess 11 5 1 Protein Quantification with Bradford ASS Y cccccsssssseeeeesseesseseeeeeeeeeeeesenseeeeeeeeoeesnneeeeeeeoeeenes 11 S11 KT eet EEN 11 5 1 2 Operation Instructions eeh eege uerge esg gege eeeE heeten EEN 11 6 TROUBLE SHOOTING Je UE 12 7 FAN Go ei el 13 8 MAINTENANCE csc re ace eee ce tegen eco ec OOE 13 8 1 Cleaning and General Care of the INStrumMment ccceeeeeeeeesenseeeeeeeeceeenneeeeeeeeeeoeeneeeseseeoeeens 13 8 2 De contamination Procedure cccccssssseeesssessseeesceesseeseeensseeeeeeeesseeeeeoeasseeeseoesseeesooesseessonnssessenons 13 9 SPECIFICATION AND WARRANT aaaaaaaaaaaaaanannnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn nnmnnn 14 Version 1 3 Page 3 14 INEL CN OD600 DiluPhotometer User Manual 1 1 1 ESSENTIAL SAFETY NOTES Unpacking Positioning and Installation Check the contents of the pack against the packing list If any shortages are discovered please inform your supplier immediately Inspect the instrument for any signs of damage caused in
9. Page 8 14 INEL CN OD600 DiluPhotometer User Manual 4 OD600 DILUCELL APPLICATION 4 1 Bacterial Cell Culture Measurement 4 1 1 Description e The stage of growth of a bacterial culture needs to be monitored to ensure that the cells are harvested at the optimum point for the greatest density of live cells An exemplary growth curve is given below Cells should be harvested towards the end of the log phase The optical density of the sample indicates when this point has been reached This value varies dependent on the cells being grown Routinely the cells are grown until the absorbance at 600 nm known as OD 600 reaches approximately 0 4 prior to induction or harvesting A linear relationship exists between cell number density and OD 600 up to approx 0 6 stationary Decline Log Phase Lag e It is important to note that for turbid samples such as cell cultures the absorbance measured is due to light scattering and not the result of molecular absorption The amount of scatter is affected by the optics of the system distance between the cell holder and instrument exit slit geometry of this slit and the monochromator optics Different spectrophotometer types therefore give different responses for the same turbid sample to compare results they must be normalized using calibration curves H A calibration curve can be determined by comparing measured OD 600 to expected OD 600 Expected OD 600 is determined by counting cell
10. charged If only one or no bars are present the battery needs to be recharged Connect the instrument to the electric power supply using the adaptor recharge unit The battery will be fully recharged in 12 hours The OD600O value is different to When you measure turbid solutions you do not measure the that obtained on another instrument absorbance transmittance of light at the detector you measure the amount of in the lab scattered light that reaches the detector Thus optical geometry is very important the further the distance from the sample to the detector the greater the effect of the scattered light Thus instead of harvesting at 0 4 OD for example you have do it at 0 8 OD A simple conversion factor can be calculated from the OD 600 of your existing instrument compared to that of the OD600 instrument Version 1 3 Page 12 14 INEL CN OD600 DiluPhotometer User Manual 7 ACCESSORIES DiluCell 10 DC 10 DiluCell 20 DC 20 Adapter set for 10 and 12 mm tubes B 80 3000 57 8 MAINTENANCE 8 1 Cleaning and General Care of the Instrument The instrument has no serviceable parts The instrument requires little maintenance but the following are considered good practice 1 Keep the instrument clean and dry Wipe off any spilt liquids immediately Clean with a slightly damp cloth a non abrasive water based soap or detergent may be used The instrument may be wiped 2 Remove the cuvettes from the instrument when not i
11. e your reference or sample into the DiluCell DiluCell is very easy to fill the geometrical setup Supports air bubble free sample application It is recommended however that you check for air bubbles before the measurement In case of an air bubble in the cell please remove it by gently tapping the cell Volume requirements are 200 ul for DiluCell 10 100 ul for DiluCell 20 Important Information Remember to multiply the reading from the OD6GOO after the measurement with the proper virtual dilution factor Virtual dilution factor are 10 for DiluCell 10 20 for DiluCell 20 Important Note It is not recommended to reuse DiluCell due to e high probability of cross contamination e high probability of air bubbles when refilling the cell e decontamination not easily possible for cell culture measurements e g Version 1 3 Page 7 14 INEL CN OD600 DiluPhotometer User Manual 3 3 Specifications DiluCell 10 DiluCell 20 Material PMMA PMMA Wavelength Range 340 nm 950 nm 340 nm 950 nm Volume Requirement 200 ul 100 ul Pathlength at 8 5 mm centre height 1 0 mm 0 5 mm Tolerance 5 10 Ordering Information pack with 96 cells DC 10 DC 20 Limitations Do not autoclave DiluCell The DiluCell is approved only for the usage with the OD600 DiluPhotometer For solvent compatibility please contact your local Implen partner or the Implen Team at www implen de Version 1 3
12. n use 3 Store in a cool place away from corrosive chemicals or fumes 8 2 De contamination Procedure To decontaminate we recommend that the instrument is wiped with ethanol or other antibacterial detergent as required A soaked cloth may be inserted into the cuvette chamber or ethanol sprayed directly into the compartment The instrument can be sterilised using formaldehyde or ethylene oxide but not with UV light due to plastic degradation For severe contamination it is possible to remove the 4 screws in the base and separate the top and bottom covers taking care to not drop the battery inside the instrument The contaminated areas in the instrument may then be wiped with a suitable anti bacterial detergent Version 1 3 Page 13 14 INEL CN OD600 DiluPhotometer User Manual 9 SPECIFICATION AND WARRANTY Technical Specification Range Optical Density 0 3 Ato 1 99 A Controls 6 button power blank sample memory reset recall print Optical height 8 5mm Cuvette holder Fixed with drain hole Accepts DiluCell 10 DiluCell M20 10 mm pathlength semi micro and macro cuvettes or 14 16 mm round tubes 99 readings Power requirements External power adaptor 110 to 220 V 50 60 Hz 20 VA or internal rechargeable NiMH battery 180 x 150 x60 mm 0 6 kg Specifications are measured after the instrument has warmed up at a constant ambient temperature and are typical of a production unit As part of our policy of
13. ons Step 5 Press the sample button The display will show the OD of the standard solutions in absorbance units Step 6 Create a standard curve Step 7 Measure the absorbance of sample solutions Step 8 Determine the protein concentration of unknown samples according to the standard curve Version 1 3 Page 11 14 INEL CN OD600 DiluPhotometer User Manual 6 TROUBLE SHOOTING NOTES ERROR INDICATION SOLUTION A flashing Absorbance reading of This indicates an Absorbance of more than 1 99 and is therefore out of range 2 00 Abs is obtained The sample needs to be diluted or use DiluCell 10 or DiluCell 20 for virtual dilution A negative reading is obtained In normal measurements the test sample has a positive Absorbance compared to that of the Reference Negative readings will be obtained if the Reference and Test cuvettes are mixed up A flashing Absorbance reading of This indicates an Absorbance of less than 0 30 Abs and is therefore out of Unexpected results are obtained Any bubbles in solution will produce considerable error TTT TE REF is displayed when sample is The baseline has not been set Replace the sample with a blank or reference No reading is obtained when using Check that there is sufficient battery power available the instrument in battery mode The battery power available is indicted by the battery symbol at the bottom right hand corner of the display Three bars in the battery indicate that it is fully
14. the first reading Page 10 14 INEL CN OD600 DiluPhotometer User Manual 5 OD600 STANDARD CUVETTE APPLICATION 5 1 Protein Quantification with Bradford Assay 5 1 1 Description Bradford Assay is a rapid and accurate method used to determine the total protein concentration of a sample by quantifying the binding of a dye Coomassie Brilliant Blue G 250 to an unknown protein The assay is based on the observation that the absorbance maximum for an acidic solution of Coomassie Brilliant Blue G 250 shifts from 465 nm to 595 nm when binding to protein occurs By comparing this binding to that of different Known concentrations of a standard protein the concentration of the unknown protein can be calculated Standard curves with bovine serum albumin BSA are given below as an example Protein Quantification Bradford 1 25 ug ml Protein Quantification Bradford 100 2000 ug ml o 5 10 15 20 25 0 500 1000 1500 2000 concentration of standard ug ml concentration of standard ug ml 5 1 2 Operation Instructions Important Note Please follow the detailed protocols supplied with your Bradford assay kit to ensure accurate results Making a measurement Step 1 Switch the instrument on by pressing the ON OFF button Step 2 Place a reference into the cuvette compartment Step 3 Press the blank button The display will show 0 00 Step 4 Remove the reference and replace with standard soluti
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