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WellPro 96 and 384 - Phenix Research Products

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1. 7 4 ESTABLISHING A FILE USING THE CONTROLLER 8 4 1 INTRODUCTION RR DN C OT 8 4 2 THE KEYPAD CONTROLLER eee sese sn sn sse sess sse sese sense essen esses ense sns sn sensa 8 4 2 1 The Cor Pr 8 4 2 2 The Keypad eicit e eI 9 4 2 5 ClO AM mcr pr LR 9 4 2 4 jun E EEUU 9 4 2 5 SCOP eMe cM CEN MD MN dM MN M DIURNO 10 4 2 6 C180 o er I EE 10 4 2 7 I II 10 4 2 8 pm 10 4 2 9 MA opp M RN 11 422 10 AENT REIR ee ER e 11 4 3 ENTERING DATA Rey dune 11 4 4 Ql tee D 11 AEN QU Wii BED e E 12 5 1 QVERVIEW AS anun ss ced 12 5 2 EDITING A PLATE FILLING FILE J J u uu sse seen u 12 5 5 STORING OVERWRITING THE FILE 16 5 4 RUNNING A PLATE FILLING FILE J u u u u 16 5 5 SAMPLE PLATE FILLING EXPERIMENT ccccscccscesssescesscesccescescccsccecccsseessesseessesseescesscentecceceteesseessecseestessceatceceeeseesee 17 WellPro User Manual 01 11 04 V Table of Contents 6 SERIAL D
2. 2221 esses ns 5 1 5 USER MANUAL L C S N AS aa SSS Q usa 4 2 5 2 1 P 5 2 2 LOCATION OF THE UNIT IN THE LABORATORY 5 2 5 POWER REQUIREMENTS u u u s sns ense ensem s a u 5 2 4 UNPACKING AND INSPECTION SE S SS SSS S s TER 6 2 5 INITIAL INSTALLATION OF THE WELLPRO 384 LIQUID HANDLER J 6 me cuales iiligmee H TP 7 3 1 INTRODUCTION 2 5 eee e cere to e Su oue te v ORE Soi Mu KSO Sus SD 7 3 2 WELLPRO WP384 CoNTROLLER J J u esse sense sess uu u 7 3 3 THE WELLPRO WP384 LIQUID HANDLING UNIT u u 7 3 4 THE WELLPRO REAGENT CONTAINER J u u uu u J
3. File 22 is a plate to plate transfer in which a small sample from the diluted plate from File 21 is pipetted to a new assay plate The last line in the file indicates that the operation should be transferred to File 34 at the termination of File 22 LINK TO STORED FILE 0 45 0 STOP 34 After the WellPro liquid handler has finished File 22 the operation of the system is transferred to File 34 which is used to add reagent via the manifold system This is the final step in the overall operation as indicated by LINK TO STORED FILE 0 45 0 STOP 0 57 Note The number of the files that are linked can be any order the file number is merely used for identification purposes 44 Section 11 Adjusting the Tip Height 11 ADJUSTING THE TIP HEIGHT 11 1 Overview The depth of the tips in the wells must be properly set in order for the system to accurately aspirate fluid When standard microplates are used this depth is set by choosing one of the preset values FLAT U or V that is presented in the PLATE TYPE parameter in the various files However when other plates are used in a few special situations it will be necessary to manually set the tip height using File 70 The figures below illustrate the height positions that this file provides gt 2 200 2 200 gt gt 2 080 2 080 gt D gt 1 360 1 360 gt D 800 Fig 11 1 Important Height Values 11 1 1 Editing Running
4. Take a tube in one hand and with the other hand lower a tip into the tube just far enough so that the fins don t touch the top of the tube and the tip doesn t seal with the sides of the tube Mark the tube to show the position o the bottom of the tip This will represent the artificial bottom Manually pipette water into the tube until the meniscus reaches the mark you have made This volume of water represents the void volume Record the value for void volume on the worksheet Place the tube into a rack and insert the rack in the middle cavity Use File 70 to determine the height of the water in the tube and record the TIP NOW AT number on a worksheet as the value for WELL BOTTOM HEIGHT Pipet 50 ul of water into the tube Determine the height of liquid in the tube and record the TIP NOW AT number on a worksheet as the 50 UL HEIGHT 47 Section 11 Adjusting the Tip Height 557 Note When using tall or narrow tubes respond to the prompts the display by entering the values you have just determined for WELL BOTTOM HEIGHT and 50 UL HEIGHT Before carrying out any liquid handling operations that involve tips you must add an extra volume of liquid to each tube that equals the void volume determined above The VOLUME IN WELL AT START does not include this void volume The void volume is excluded because it is below the artificial well bottom level so the value you give for VOLUME IN WELL AT START reflects only the amount
5. 12 W Mean Weight Volume Dispensed Standard Deviation Vi VI 23 Coefficient of Variation SD x 100 Calibration Setting 56 Section 12 Calibration Procedures 12 6 Quality Control Procedures IES Note All laboratories should have a functioning quality control program to provide verification of the accuracy and precision of its test results The use of assayed control samples is helpful in monitoring the overall performance of the WellPro liquid handler and the reagents used in an assay For example when serum samples are studied it is a good practice to assay serum controls in the normal and abnormal ranges with each batch of samples When using commercial control sera the value of each control should fall within the manufacturer s stated acceptable range for the analyte When a commercial control is not available a control can be generated by adding known amounts of the analyte to a normal serum pool For research specimens add known amounts of analyte to the appropriate matrix Aliquot into containers and store at a temperature that is optimum for the sample s stability For maximal control choose at least three levels of concentration low mid range and high You may wish to choose additional concentrations at decision points or at questionable points on the standard curve where necessary A Levey Jennings control chart as shown in Figure 12 1 provides the simplest way to present daily con
6. 70 TIP HEIGHT 10 Section 4 Establishing a File using the Controller 4 2 9 Yes No The Yes and No keys are used to respond to questions asked by the WellPro controller and tell it whether you want it to do certain tasks Each option will be underlined in sequence until you press step 4 2 10 Enter 1 If the Enter key is pressed after an alphanumeric key has been selected the indicated value is included in the file 2 If the Enter key is pressed at the END OF FILE RUN the run will start 43 Entering Data into a File Each of the template files contains a series of steps that are used to enter the appropriate data for the task to be performed and you can scroll through the steps via the Step key When you come to a step that must be edited the appropriate selection can be made in one of two ways e The user must enter a numeric value within the range As an example the VOLUME IN WELL AT START step Figure 4 3 is used to tell the liquid handler how much liquid is in each well before any transfer has been made The WellPro uses this number to determine how far down into the wells the tips should go and also to prevent the instrument from attempting to add more liquid than each well can hold VOLUME IN WELL AT START 0 350 Fig 4 5 The Volume Well at Start Step You can set the value of this parameter from 0 to 50 uL in 1 uL increments via the numeric keypad After the desired value has been entered p
7. RUN STORE END OF FILE is used to indicate what the system should do after you have completed the editing of the file e f you select RUN the file will start f you select STORE the file will be copied into memory see Section 5 5 for additional information about storing files 887 Note You can also review the file you have just made by continuing to press the STEP key 17 ENTER FIRST TIP ROW ENTER FIRST TIP ROW A P 170 15 Section 5 Plate Filling 53 Storing Overwriting the File To store a new file 1 At the END OF FILE statement item p Section 5 2 select STORE The display will appear as shown below FILE STORE ENTER USER 2 Enter your user number or project number this number is provided for reference only and does not affect the use of the system then press ENTER The display will indicate FILE STORE ENTER FILE XX where xx is the file number assigned by the system it will be the lowest unoccupied file If you want to assign a different number enter it 5 Press ENTER The display will present YES TO STORE USER 2222 FILE XX 4 Press YES to accept the assignment 5 4 Running a Plate Filling File When you are ready to run a Plate Filling file 4 Select the desired file 2 Press START If the MICROTROF does not contain the required amount of liquid to perform the desired operation the message PLEASE FILL MICROTROF wil
8. Specifications Chapter 14 In addition a series of appendices include the warranty and ordering information Section 2 Installation Procedures 2 INSTALLATION PROCEDURES 2 1 Overview The WellPro 384 Liquid Handler is designed to be installed by the operator This chapter describes how the unit is to be unpacked and assembled 2 2 Location of the Unit in the Laboratory The unit should be installed in a facility with the following conditions e The temperature should be maintained between 20 26 68 79F To prevent temperature fluctuations do not place the WellPro 384 liquid handler in direct sunlight near equipment that gives off heat or under heating or air conditioning outlets e flammable or toxic solvents are to be used a suitable ventilation system should be provided Theuse of open flames the laboratory should be prohibited e Corrosive vapors or dust should not be present as these materials can adversely affect the long term performance of the system The Well Pro fluidics module requires approximately xx mm yy of linear bench space In addition the controller requires xx cm yy 557 Note Place the WellPro 384 a stable level bench or hood Excessive jarring or vibration during operation may affect its performance 23 Power Requirements The WellPro 384 Liquid Handler to designed to operate at 100 120 220 or 240 V 50 60 Hz with a maximum power consumption of 560 watts The operating
9. WARNING WARNING For protection from injury by the WellPro liquid handler moving parts Do Not Remove the blue plastic cover while the WellPro liquid handler is operating or the power is on 13 2 Tips The WellPro liquid handler has been calibrated using WellPro 384 disposable tips The design of the tips and the manufacture of their internal volumes orifice diameters lengths etc to tight tolerance limits assure the accuracy and precision of the liquid handler system The length of the tips is of particular importance since the proper draw off of liquid from the tip depends heavily on its depth in the liquid that it is in contact with Tips not manufactured by ProGroup may decrease the accuracy and precision of the system 13 3 Power Interruptions If the power supplied to the WellPro has been interrupted the following actions will happen when the power is restored 1 If you were editing a file you will lose the file and will have to re edit the file 2 If you had just stored a file the WellPro will store the file 3 If you in the middle of a run you will lose the run Homing the instrument will sequence it back to a point where you can start over 59 Section 13 Operating Hints and Maintenance 13 4 Maintenance 13 4 1 Preventive Maintenance To ensure trouble free operation keep the WellPro liquid handler clean and free of chemicals dirt and dust To ensure trouble free operation you should keep your WellPro
10. rows B P will be active EMPTY LAST ROW TO EMPTY LAST ROW TO OFF 1ST ROW MAG MICROTROF3 EMPTY LAST ROW TO is used to tell the system where to place the extra volume of liquid transferred into the last row of wells after a serial dilution 1 If you select OFF the extra liquid will be left in the last row of wells 2 If you select 1ST ROW the extra liquid will be removed from the last row of wells and return it to the first row of wells you chose in ROWS TO DILUTE 5 If you select MAG the system will remove the extra liquid and discard it into the tip magazine 4 If you select MICROTROFS the system will remove the extra liquid and discard it into the rear reservoir of a MICROTROF container located in the P2 cavity of the template 557 Note Note will appear the display only when the 24 wells across PLATE 11 ORIENTATION is selected LIQUID HANDLING SPEED LIQ HANDLING SPEEDS RAPID MEDIUM SLOW LIQUID HANDLING SPEEDS is used to select the rate sample delivery from the tips rapid medium or slow 21 12 13 14 Section 6 Serial Dilution LINK TO STORED FILE LINK TO STORED FILE 0 45 0 STOP 0 LINK TO STORED FILE is used to indicate what the system should do after it completes the file it is currently running If you want the system to stop at the end of the file enter O If you want the system to perform another file after the p
11. 3 1 Introduction The WellPro 384 Liquid Handler system includes the following components e WellPro Controller Section 5 2 e WellPro WP384 Liquid Handling Unit Section 3 3 e WellPro Reagent Containers 3 2 WellPro WP384 Controller The WellPro WP384 Controller includes a keyboard and display which provides the operator with complete control of all liquid handling operations The basis of operation is via a user generated method which includes a series of primary operations The method can be stored and retrieved as desired Each primary operations provides for user control of a given aspect of the operation Typical operations include the volumes to transfer mix or dispense the transfer speed and whether to change tips between transfers 3 3 WellPro WP384 Liquid Handling Unit The WellPro WP384 Liquid Handling Unit is an electro mechanical transfer unit that includes a table and a vertical head assembly The table accommodates interchangeable templates which position reagent containers a magazine with rows of disposable tips etc During operation the table slides back and forth horizontally to bring the appropriate workstation under the head The head assembly consists of three basic parts Aplunger housing which contains nozzles that press into the tips to form airtight seals Aplunger block which moves up and down to aspirate and deliver samples e tip ejector plate which removes used tips The head assembly
12. across the front of the instrument as shown in Figure 1 2 Fig 1 2 Standard Template e Serial Dilution Template 16 Across The Serial Dilution template 16 Across is used when serial dilution is to be performed lengthwise on a plate In this case the plate is oriented such that the 16 wells of a column face the front of the instrument Figure 1 3 Fig 1 3 Serial Dilution Template Section 1 Introduction 1 3 About the User Manual This User Manual has been written for the end user of the system e g a laboratory technician and provides information about the operation and maintenance of the WellPro WP384 Liquid Handler Read the manual in its entirety prior to operating the instrument This User Manual has been designed to give you the information you need to Review safety precautions Install the system Use the system in routine jobs and research Perform basic maintenance procedures This User Manual also describes features and specifications of the WellPro WP384 Liquid Handler This manual includes the following sections Installation Chapter 2 System Components Chapter 3 Establishing a File Chapter 4 Plate Filling Chapter 5 Serial Dilution Chapter 6 Plate to Plate Transfer Chapter 7 Emptying a Plate Chapter 8 Time Delay Files Chapter 9 Linking Files Chapter 10 Adjusting the Tip Height Chapter 11 Calibration Procedures Chapter 12 Operating Hints and Maintenance Chapter 15
13. if you intend to work outside the normal operating range If desired you can make a volume correction by modifying the volume settings entered into the WellPro liquid handler when creating a file For example if the operating temperature is 55 and 51 pL are dispensed when 50 pL is anticipated you may enter 49 pL into the file to compensate for the change If you will be continually working outside the specified temperature range a permanent change can be made to make the volume correction automatically For details call ProGroup Technical Service at 1 800 471 1916 12 2 1 Materials Required To perform a gravimetric verification you will need the following 1 Analytical Balance 2 Calibrated Thermometer 3 Calibration Weighing Wells 4 Calibration Plate 5 Forceps 6 Water high purity 7 New WellPro Tip Magazine and Tips 8 Photocopy of the Gravimetric Worksheet From This Manual Table 12 2 50 Section 12 Calibration Procedures 12 3 Calibration Procedure 1 Assemble all items and allow them to equilibrate at room temperature for at least one hour During this time use File 70 to determine the well bottom and 200 uL heights for the calibration plate detailed discussion of this procedure is presented in Chapter 10 Weigh each well of the calibration plate Record this and all other data on the worksheet Place a 24 across template on the table Place a magazine of tips in the back cavity of the temp
14. is used to indicate what the system should do after you have completed the editing of the file If you select RUN the file will start If you select STORE the file will be copied into memory see Section 6 3 for additional information about storing files ENTER FIRST TIP ROW ENTER FIRST TIP ROW A P E ENTER FIRST TIP ROW is used to specify which row of tips to use Rows of the tip magazine are designated by letters A through P back to front 50 Section 7 Plate to Plate Transfer 7 4 Storing Overwriting the File To store a new file 1 At the END OF FILE statement item p Section 5 2 select STORE The display will appear as shown below FILE STORE ENTER USER 2 Enter your user number or project number this number is provided for reference only and does not affect the use of the system then press ENTER The display will indicate FILE STORE ENTER FILE XX Where xx is the file number assigned by the system which will be the lowest unoccupied file If you want to assign a different number enter it 8 Press ENTER The display will present YES TO STORE USER zzzz FILE XX 4 Press YES to accept the assignment 7 5 Running a Plate to Plate Transfer When you are ready to run a plate to plate transfer 1 Select the desired file 2 Press START If the MICROTROF does not contain the required amount of liquid to perform the desired operation the
15. liquid handler clean and free of chemicals dirt and dust liquids spilled on the WellPro liquid handler should be wiped off immediately Keep your WellPro liquid handler free of any corrosive reagents Wipe spills immediately Clean non contaminated work surfaces regularly with a damp cloth 13 4 2 Cleaning up Spills If solution is accidentally spilled on the instrument clean them up immediately Although all electrical components within the instrument have been either shielded or sealed against fluid contact there may be long term deleterious effects from corrosion or salt accumulation To ensure maximal safety and trouble free operation clean up spills immediately Turn off and unplug the WellPro liquid handler before wiping up spills 13 4 3 Piston Plungers On a periodic basis gently apply a light silicone grease to the piston plungers of the 24 channel liquid head using a clean cloth or lint free towel This will keep them sealing well and inhibit corrosion 13 4 4 Table Rails The rails on which the WellPro table runs must be kept clean and free of foreign matter to ensure smooth operation Periodically clean these by wiping with a clean cloth ALWAYS BE CERTAIN POWER IS TURNED OFF BEFORE CLEANING THE TABLE RAILS 13 4 5 Cleaning the Plastic Cover The plastic covers should be cleaned using a dilute solution of a mild nonabrasive cleaner and then rinsed with a soft clean damp cloth Note Do not use organic solve
16. message PLEASE FILL MICROTROF will appear on the screen This prompt allows you to simply enter the volume you are adding to the MICROTROF reservoir without requiring you to step back through each of the file s parameters At the conclusion of the operation the system will stop or proceed to the file that is linked to it 51 Section 7 Plate to Plate Transfer 7 6 Sample Serial Dilution Experiments File 63 Plate to Plate Transfer The following example will show you how to perform a simple transfer of 30 uL of a simulated cell suspension from plate 2 to plate 1 which contains no liquid in its wells prior to transfer There will be four 30 uL medium speed mixes in each well before solution is transferred as might be the case when you want to gently agitate cells into suspension before transferring them You will be using flat bottomed plates 1 Placea 24 across template on the table 2 Place a magazine of tips in the back cavity of the template 3 For demonstration purposes manually pipette 50 of water into each of the 384 wells of plate 2 4 Place the plate in the middle cavity of the template 5 Place empty plate in the front cavity of the template 6 Press file 63 and then enter to access File 63 7 Edit the file using the following keystrokes 52 Section 7 Plate to Plate Transfer Display Before Editing Keystrokes Display After Editing PLATE TO PLATE STEP STEP TO EDIT FILE 65 TRANSFER D
17. operation Once the WellPro liquid handler has paused you may either abort the file by pressing the Stop key again or go through the file one motion at a time by pressing the Step If you are using the Tip Height File File 70 the Stop key is also used to exit from the file If you are using the Home Pump Prime File the Stop key will stop priming the pump 4 2 7 Help 1 Help is normally used to determine the numeric range for a key sequence 2 During file editing the Help key allows you to override the range limits on many parameters Normally the WellPro WP384 Liquid Handler will verify the validity of a parameter e g the transfer volume when it is entered by checking against the allowable range of values for the parameter which is shown on the display By pressing the help key the controller in instructed to accept ANY entered value subject only to physical limitations of the instrument 4 2 8 File A File is a stored set of instruction and conditions which the WellPro WP384 Liquid Handler uses to perform the desired activity The user selects the desired file via the alphanumeric keypad and presses the File followed by the Enter key to load it 70 files are available Files 51 through 70 are pre programmed in WellPro WP384 and you can modify them to create new files Files 1 through 45 are user storage files The initial standard files are as follows 0 HOME PUMP PRIME 61 PLATE FILLING 62 SERIAL DILUTION 65 PLATE TO PLATE
18. should go and also to prevent exceeding the volume capacity of the wells This parameter can be set from 0 to 50 uL in 1 uL increments 5 TRANSFER MODE TRANSFER MODE 1 TO 1 1 TO SEVERAL TRANSFER MODE is used to tell the system whether to transfer one row of plate 1 to the corresponding row of plate 2 or to transfer one row of plate 1 to several different or multiple rows of plate 2 27 6 10 Section 7 Plate to Plate Transfer TRANSFER VOLUME TRANSFER VOLUME 0 50 25 ul TRANSFER VOLUME is used to tell the system how much sample to withdraw from a source well for transfer to a new well during a plate transfer This parameter can be set from O to 50 uL in 1 uL increments The system calculates the maximum TRANSFER VOLUME for the row by dividing the VOLUME IN WELL AT START by the number of transfers specified for that row MIXES BEFORE TRANSFER MIXES BEFORE TRANSFER 0 9 0 MIXES BEFORE TRANSFER is used to indicate the number of aspiration delivery cycles that should be performed before transferring a sample For example when you are diluting cells which may settle you should agitate them back into suspension before transfer by mixing This parameter can be set from 0 to 9 0 indicates no mixing MIXES AFTER TRANSFER MIXES AFTER TRANSFER 0 9 0 MIXES AFTER TRANSFER is used to indicate the number of aspiration delivery cycles that should be performed after transf
19. slides up and down vertically to bring the tips to the appropriate height to perform the designated operations A 24 channel tip air displacement pipetting head is included on the WellPro liquid handler as most protocols are designed to be performed in the 24 channel mode but serial dilutions can be performed in the 16 channel mode to allow for maximum dilution capability with the 384 well plate The table assembly drives the plate which holds the well plates and the Microtrof reagent containers 3 4 WellPro Reagent Container The WellPro liquid handler is used with a Microtrof which is a reagent container The Microtrof container consists of up to three reservoirs each of which is used to contain a reagent or solution In some cases fluid from the wells is deposited in the Microtrof The WellPro WP384 liquid handler uses templates to position microplates reagent containers and tip magazines on the movable table Two templates are provided as described in Section 1 5 4 Section 4 Establishing a File using the Controller 4 ESTABLISHING A FILE USING THE CONTROLLER 4 1 Introduction The basis of operation of the WellPro 584 Liquid Handler is the which is a set of instructions and conditions that are used to perform the desired operation The operator can establish store and execute files via the controller module The system includes a number of template files that can be used to establish files for actual operation To
20. wells the tips should go If you are not using a standard microplate select HEIGHT and determine the amount to raise or lower the tip height by using File 70 the Tip Height File Section 10 The height parameter can be set from 100 to 3500 in increments of 1 MIXES BEFORE EMPTY MIXES BEFORE EMPTY 0 9 0 MIXES BEFORE EMPTY is used to tell the system how many mixes aspiration delivery cycles should be performed during the mixing process before waste removal You can set the value of this parameter from 0 to 9 mixes in increments of 1 The numeral 0 indicates no mixing 57 8 10 11 Section 8 Empting a Plate ROWS TO EMPTY ROWS TO EMPTY A B C D E F G H I J K L M N O P ROWS TO EMPTY is used to indicate the rows of the plate to empty and which to leave illed Rows are designated by letters A through P To select row A press YES The underline will remain and B will be active if you do not want to use a row press NO to advance to the next row If you want all rows to be emptied press the STEP key After the last NO selection has been made press STEP i e if you want all rows except for A press NO for A The active row will become B If you press STEP rows B P will be active LIQUID HANDLING SPEEDS LIQ HANDLING SPEEDS RAPID MEDIUM SLOW LIQUID HANDLING SPEEDS is used to select the rate sample withdrawal by the tips rapid medium or slow LINK TO STORED FILE LIN
21. 50 ul by the number of rows a transfer will be made to two rows C and D The maximum acceptable volume for the transfer is therefore 25 ul 54 Section 7 Plate to Plate Transfer Display Before Editing Keystrokes Display After Editing TRANSFER VOLUME 25 ENTER TRANSFER VOLUME 0 50 50 ul 0 50 25 ul MIXES BEFORE TRANSFER STEP MIXES BEFORE TRANSFER 0 9 0 0 9 0 MIXES AFTER TRANSFER STEP MIXES AFTER TRANSFER 0 9 0 0 9 0 5 5 CHANGE TIPS YES NO YES NO PLATE 1 TYPE STEP PLATE 1 TYPE FLAT U V HEIGHT TUBE FLAT U V HEIGHT TUBE PLATE 2 TYPE STEP PLATE 2 TYPE FLAT U V HEIGHT TUBE FLAT U V HEIGHT TUBE LIQ HANDLING SPEEDS STEP LIQ HANDLING SPEEDS RAPID MEDIUM SLOW RAPID MEDIUM SLOW LINK TO STORED FILE STEP LINK TO STORED FILE 0 45 0 STOP O 0 45 0 STOP 0 END OF FILE ENTER RUN STORE PLATE TO PLATE START START TO RUN FILE 63 557 Note WellPro liquid handler allows you to create and store multiple versions of File 63 Plate to Plate Transfer so as to transfer from row A of plate 1 to rows A and B of plate 2 from row B to rows C and D from row C to rows E and F from row D to rows G and H and then link these files together The result is that you can conveniently transfer from the rows of one plate to duplicate rows of another plate for assay using a single start keystroke Details on file linking are provided in Section 10 55 Section 8 Empting a Plate 8 EMPTYING A PLA
22. 75 0 200 75 MIXES AFTER TRANSFER 4 MIXES AFTER TRANSFER MIX VOLUME 25 MIX VOLUME 0 50 50 25 PLATE TYPE U PLATE TYPE FLAT U V HEIGHT TUBE FLAT U V HEIGHT TUBE FILL PLATE ROWS C FILL PLATE ROWS AB C D E F C H K L M N O P A B CD E F O H E HCL M N O P MICROTROF VOL AT START 70 MICROTROF VOL AT START 20 80 77 mL 20 80 70 CHANGE TIPS YES CHANGE TIPS YES NO YES YES NO YES HANDLING SPEEDS HANDLING SPEEDS RAPID MEDIUM SLOW RAPID MEDIUM SLOW LINK TO STORED FILE 0 LINK TO STORED FILE 0 45 O STOP 0 0 45 0 5 0 END OF FILE RUN STORE Section 6 Serial Dilution 6 SERIAL DILUTION 6 1 Overview Serial Dilution is the progressive dilution and redilution of a liquid In this process a sample is added to a diluent and the two are mixed An aliquot is then removed for addition to the next diluent containing well This sequence is repeated until the desired number of dilutions have been obtained serial dilutions As an example if equal volumes of sample and diluent were mixed during each step of the series the sample concentration would be 1 2 1 4 1 8 1 16 1 32 and so on in sequential rows The template for Serial Dilution is File 62 This file can be used as a template to establish a user file which can be stored and used as desired and it can be as a template for additional serial dilution files 6 2 Editing a Serial Dilution File When you open File 62 or any other Serial Dilution file the opening step
23. E FLAT U V HEIGHT TUBE TRANSFER ROWS A B C D E F G H I J K L M N O P LIQ HANDLING SPEEDS RAPID MEDIUM SLOW LINK TO STORED FILE 0 45 0 5 0 Section 12 Calibration Procedures 9 Quickly weigh each well of the row dispensed 10 Repeat steps eight through ten for each row 11 Subtract the initial weighing from the final weighing for each well 12 4 Calculations To calculate the accuracy and precision of the WellPro liquid handler 1 Determine the arithmetic mean average weight of each column by adding all sample net weights together and dividing by the number of samples 2 Calculate the mean volume at the determined water temperature using the mean weight and Volume Conversion Factor Table 12 1 using equation 12 1 Volume Weight x Volume Conversion Factor 12 1 Table 12 1 Volume Conversion Factor Water Temperature Conversion Factor 15 0 1 002 17 5 1 002 20 0 1 005 22 5 1 005 25 0 1 004 27 5 1 005 50 0 1 005 5 Determine the standard deviation SD of the individual volumes within each column using the equation 11 2 SD XIWiW Px 41 l ViVi Volume Conversion Factor 12 2 1 density N 1 Where Wi is the weight of an individual well WI is the average of the weight of the individual wells Vi is the volume of an individual well Vl is the average of the volume of the individual wells N is the number of wells 24 4 Determine the coefficient of variation CV using the equation 12 5 C
24. E STORE ENTER FILE XX Where xx is the file number assigned by the system which will be the lowest unoccupied file If you want to assign a different number enter it 8 Press ENTER The display will present YES TO STORE USER zzzz FILE XX 4 Press YES to accept the assignment 9 4 Using Time Delay File A time delay file is used with linked files A detailed discussion of linked files is presented in Section 10 While a Time Delay file is active the display will show the time remaining 42 Section 10 Linking Files 10 LINKING FILES 10 1 Overview The FILE LINK feature is provided to allow you to join two or more stored files as a sequence enabling the user to automate very complex liquid handling operations A file parameter LINK TO STORED FILE appears just before the END OF FILE in each file type 0 see Chapter 7 This prompt permits you to direct the WellPro liquid handler to transfer control to another file immediately after completing the file that it is currently running instead of terminating the operation of the file 10 2 Using Linked Files When the system is operating using a linked file sequence the sequence will use the tips indicated in the ENTER FIRST TIP ROW prompt for the first file in the sequence This prompt appears regardless of whether the tips are to be changed in the first file The user should respond by specifying the row of tips A P the standard magazine plus t
25. E TYPE PLATE TYPE FLAT U V HEIGHT TUBE PLATE TYPE is used to indicate the design of the bottom of the plate This information is used to determine the height of the liquid in each well and thus calculate how far down into the wells the tips should go If you are not using a standard microplate select HEIGHT and determine the amount to raise or lower the tip height by using File 70 the Tip Height File Section x xx The height parameter can be set from 100 to 3500 in increments of 1 PLATE ORIENTATION PLATE ORIENTATION 24 16 WELLS ACROSS PLATE ORIENTATION is used to indicate how the plate is positioned The value that is selected indicates the number of wells that will be under the pipette tips at one time 20 10 Section 6 Serial Dilution ROWS TO DILUTE ROWS TO DILUTE A B C D E F G H I J K L M N O P ROWS TO DILUTE is used to indicate the rows of the plate are to be included in the serial dilution Rows are designated by letters A through P back to front in the 24 across configuration and 1 through 16 in the 16 across configuration To select row A press YES The underline will remain and B will be active if you do not want to use a row press NO to advance to the next row If you want all rows to be filled press the STEP key After the last NO selection has been made press STEP i e if you want all rows except for A press NO for A The active row will become B If you press STEP
26. ICROTROF is especially important with cells or other inhomogeneous samples This parameter can be set between 0 and 9 0 indicates no mixing MICROTROF MIX VOLUME MICROTROF MIX VOLUME 0 200 50 MICROTROF MIX VOLUME is used to indicate how much liquid to aspirate into the tip and dispense back into the MICROTROF in order to mix the MICROTROF contents This parameter can be set from O 50 in 1 ul increments 0 indicates no mixing Mix efficiency may vary with MICROTROF volume DELIVERY VOLUME DELIVERY VOLUME 0 200 50ul DELIVERY VOLUME indicates how much solution should be dispensed into each well from the MICROTROF reservoir This parameter can be set from 0 50 in 1 uL increments MIXES AFTER TRANSFER MIXES AFTER TRANSFER 0 9 2 MIXES AFTER TRANSFER is used to specify the number of mixes after the aliquot has been delivered to the plate This parameter may be set from 0 9 in increments of 1 0 indicates no mix after transfer MIX VOLUME MIX VOLUME 0 50 50ul MIX VOLUME is used to specify the volume of the aspirate delivery cycles used to mix the contents of the well after transfer The upper limit is calculated as 75 of the transfer fill volume This parameter can be set from 0 to the upper limit in increments of 1 uL PLATE TYPE PLATE TYPE FLAT U V HEIGHT TUBE PLATE TYPE is used to indicate the design of the bottom of the plate This informati
27. ILUTION iioi eiae oe eic Peste ote ote cecidere obe oido ans 19 6 1 OVERVIEW ate inum sau ou 19 6 2 EDITING A SERIAL aeree SS cete ba pea soe ud vae ve 19 6 5 STORING OVERWRITING THE FILE 25 6 4 RUNNING SERIAL DILUTION FILE cccccccscccsccsssescesssesccescescceccececsseessesseessesseescessceatecsseeseesseessecceecesseeateeceeeseesee 23 6 5 SAMPLE SERIAL DILUTION EXPERIMENT u u u u u 24 7 PLATE TO PLATE TRANSFER uu u Hn u Qu aqu snn nas ss panapas h gea 26 7 1 OVERVIEW aa as 26 7 2 TYPES OF PLATE TO PLATE TRANSFFER J J u u u u u 26 7 5 EDITING PLATE TO PLATE TRANSFER FILE J a sete senes u 27 7 4 STORING OVERWRITING THE 51 7 5 RUNNING A PLATE TO PLATE TRANSFFER l u uuu sess u u 31 7 6 SAMPLE SERIAL DILUTION EXPERIMENTS u u u u u 32 8 EMPTYING A PLATE uu e
28. ING HINTS AND 2222 59 13 1 OVERVIEW a 59 13 2 59 15 5 u eee ieee ee repe ee EEG Eee x RE eve ac 59 13 4 LP geo 60 13 41 Preventive Maintenance u 60 15 4 2 Cleaning UP SPIS nihin nnn 60 1 5 4 5 Pist n PI ngers eee eere e rv nee e ee e O ua ERR awa RD SR FREE o 60 15 4 4 x 60 15 4 5 Cleaning the Plastic COVER tette te tecto ee ce EQ EE e ee e Re gutter 60 15 46 F se Re R NN YT 61 13 5 TROUBLESHOOTING esee eer e ege Ee ec pei eg ede Eea ee use eee de epe Pe 61 14 WELLPRO WP384 SPECIFICATIONS 62 WellPro User Manual 01 11 04 vii Section 1 Introduction 1 INTRODUCTION 1 1 Overview The ProGroup WP384 Liquid Handler Figure 1 1 is designed to perform the following liquid handling activities on a user programmable basis Precisely transfer liquids in volumes fro
29. IRECTION STEP TRANSFER DIRECTION P2 TO P1 P1 TO P2 VOL IN WELL PLATE 1 0 50 0 ul VOL IN WELL PLATE 2 0 50 TRANSFER MODE 1 TO 1 TO SEVERAL TRANSFER VOLUME 0 50 MIXES BEFORE TRANSFER 0 9 MIXES AFTER TRANSFER 0 9 MIX VOLUME 0 22 CHANGE TIPS YES NO PLATE 1 TYPE FLAT U V HEIGHT TUBE PLATE 2 TYPE FLAT U V HEIGHT TUBE TRANSFER ROWS A B C D E F GH I J K L M N O P LIQ HANDLING SPEEDS RAPID MEDIUM SLOW LINK TO STORED FILE 0 45 O STOP 0 END OF FILE RUN STORE PLATE TO PLATE START TO RUN FILE 65 30ul IO IO STEP STEP STEP 70 ENTER 4 ENTER STEP 22 ENTER STEP STEP STEP STEP NO STEP STEP ENTER START 55 P2 TO P1 P1 TO P2 VOL IN WELL PLATE 1 0 50 VOL IN WELL PLATE 2 0 50 TRANSFER MODE 1 TO 1 TO SEVERAL TRANSFER VOLUME 0 50 MIXES BEFORE TRANSFER 0 9 MIXES AFTER TRANSFER YESNO 0 9 MIX VOLUME 0 22 ul 22 ul CHANGE TIPS PLATE 1 TYPE FLAT U V HEIGHT TUBE PLATE 2 TYPE FLAT U V HEIGHT TUBE TRANSFER ROWS A B C D E F GH I J K L M N O P LIQ HANDLING SPEEDS RAPID MEDIUM SLOW LINK TO STORED FILE 0 45 0 STOP 0 30ul Ie IO File 63 Plate to Plate Transfer Mode and Direction Section 7 Plate to Plate Transfer The following example will show you how to use the TRANSFER MODE capability in combination with File 63 Plate to Plate Transfer to transfer 35 uL of w
30. K TO STORED FILE 0 45 0 STOP 0 LINK TO STORED FILE is used to indicate what the system should do after it completes the file it is currently running If you want the system to stop at the end of the file enter O If you want the system to perform another file after the present file is complete enter the desired file number The LINK TO STORED FILE command permits more efficient operation because you don t need to start each individual file manually with the file A detailed discussion of Linked files is presented in Section 10 END OF FILE END OF FILE RUN STORE END OF FILE is used to indicate what the system should do after you have completed the editing of the file If you select RUN the file will start If you select STORE the file will be copied into memory see Section 5 5 for additional information about storing files 557 Note You can also review the file you have just made by continuing to press the STEP key 58 Section 8 Empting a Plate 12 ENTER FIRST TIP ROW ENTER FIRST TIP ROW A P P ENTER FIRST TIP ROW is used to specify which row of tips to start with Rows of the tip magazine are designated by letters A through P from back to front 8 3 Storing Overwriting the File To store a new file 1 At the END OF FILE statement item 11 Section 8 2 select STORE The display will appear as shown below FILE STORE ENTER USER 2 Enter yo
31. START is used to how much liquid is in each well before any solution has been removed The system uses this number to determine how far down into the wells the tips should go and also to prevent the instrument from attempting to withdraw present This parameter can be set from to 50 uL in 1 increments 56 5 VOLUME TO EMPTY Section 8 Empting a Plate VOLUME TO EMPTY 0 50 50 ul VOLUME TO EMPTY is used to indicate how much liquid to remove from each well and transfer to the waste container a reservoir or the MICROTROF container This parameter can be set from 0 to 50 uL in 1 pL increments 4 WASTE TO MICROTROF WASTE TO MICROTROF T1 T2 T3 WASTE TO MICROTROF is used to indicate which reservoir of the MICROTROF container to deliver its waste solution to This parameter can be set to T1 T2 or T3 which indicate respectively the front middle and rear reservoirs 5 CHANGE TIPS CHANGE TIPS YES NO CHANGE TIPS is used to indicate whether you want it to get new tips from the tip magazine during the waste removal thus decreasing carryover If you don t want to change tips the system will use the same tips for the whole plate PLATE TYPE PLATE TYPE FLAT U V HEIGHT TUBE PLATE TYPE is used to indicate the design of the bottom of the plate This information is used to determine the height of the liquid in each well and thus calculate how far down into the
32. TE 8 1 Overview The WellPro liquid handler can remove all or part of the solution in a plate and replace it with another solution which is delivered from a MICROTROF reservoir The steps that the WellPro liquid handler uses to empty a plate are 1 Aspirate sample from plate 2 Deliver to T3 waste reservoir The template for Emptying a plate is File 64 which is used as a template to establish a user file that can be stored and used as desired and it can be used as a template for additional Emptying a plate files 557 Note This file is designed to make rapid exchanges of culture media reagents diluents etc within a single plate For refilling with a new reagent link to an edited FILE 61 plate fill file 8 2 Editing an Emptying a Plate File When you open file 64 or any other Serial Dilution file the opening step is presented and the step command should be selected to view the file on a screen by screen basis You can change the value of each parameter as desired to provide the desired program to fill the plates After a value is entered press ENTER To advance to the next step press STEP A detailed discussion of role of each key is presented in Chapter 4 1 OPENING STEP PLATE EMPTY STEP TO EDIT FILE 64 When File 64 is opened the OPENING STEP display is presented If you want to view edit the file press STEP 2 VOLIN WELL AT START VOL IN WELL AT START 0 50 50 ul VOLUME IN WELL AT
33. The WellPro Automated Liquid Handling System m ProGroup Instrument Corp 1 800 471 1916 www wellpro us WellPro 96 and 384 Liquid Handler EES Eee User Manual User Manual Rev 2 0 November 2004 NOTE This manual is written to be used for either the WellPro 96 or WellPro 384 instruments The instruments vary in that the liquid heads are specifically designed for either a 96 or 384 well microplate The WellPro 96 uses 12 channel liquid head with 200ul capacity tips and the WellPro 584 uses a 24 channel liquid head with 50ul capacity tips The examples given in the text have been written for the WellPro 384 ie Rows A to P Columns 1 to 24 transfer volumes 50ul max etc When using the WellPro 96 the appropriate volumes and positions will be displayed on the hand controller The tutorial examples given can be followed for either instrument WellPro 96 specs are included on this CD ROM under the WellPro 96 Brochure If you prefer a hard copy of this manual exclusively for the WellPro 96 please contact Customer Service at 1 800 471 1916 1 618 258 7395 or www wellpro us Trademarks WellPro Automated Liquid Handling System is a registered trademark of ProGroup Instrument Corp All other trademarks and registered trademarks are property of their respective holders Disclaimer ProGroup reserves the right to change its products and services at any time to incorporate technological developments This manual is subject to change without
34. V SD X 100 12 5 Mean Volume A Gravimetric Worksheet presented as Table 12 2 can be used to assist in calculating the CV Four columns are shown for the sake of clarity This worksheet should contain 12 columns 55 Section 12 Calibration Procedures TABLE 12 2 Gravimetric Worksheet Date Tech Temperature GRAVIMETRIC WORKSHEET 1 2 3 4 ROW IW FW NW W FW NW IW FW NW W FW NW A B C D E F G H W MT P Vol NE X E SD TE _ E CV IW Initial Weight FW Final Weight NW Net Weight 54 Section 12 Calibration Procedures 12 5 Calibration of Non Aqueous or Viscous Solutions 12 5 1 Introduction The WellPro is normally calibrated with water as it is the most common liquid used in clinical and biological applications Water has unique viscosity surface tension and binding characteristics If you are using a different solvent it is probable that these characteristics will differ radically from that of water and the calibration using water might not be valid Alternatively if you are pipetting viscous fluids the procedure described below should be used to calibrate the system 887 Note solutions such as 20 glycerol exhibit radical viscosity changes when cooled 12 5 2 Procedure 1 Determine the weight of a specific volume of the test solution at the temperature you plan to use routinely by weighing a known volume in a volumetric flask 2 Calculate the density
35. advance to the next step press STEP A detailed discussion of role of each key is presented in Chapter 4 1 OPENING STEP PLATE FILLING STEP TO EDIT FILE 61 When File 61 is opened the OPENING STEP display is presented If you want to view edit the file press STEP 2 PLATE POSITION STEP PLATE POSITION 1 2 PLATE POSITION is used to whether the plate to be filled is in the FRONT cavity of the template Position 1 the second cavity of the template Position 2 or both Underline the desired position s 5 VOLUME IN WELL AT START VOL IN WELL AT START 0 350 Qul VOLUME IN WELL AT START is used to indicate tell how much liquid is in each well before any transfer has been made This number is used to determine how far down into the wells the tips should go and also to prevent the instrument from attempting to add more liquid than each well can hold This parameter can be set from 0 to 50 in 1 uL increments 4 DELIVER FROM DELIVER FROM T1 T2 T3 DELIVER FROM is used to indicate the MICROTROF reservoir that contains the liquid to be dispensed 1 T2 refers to the front middle and rear reservoirs of MICROTROF container Underline the desired choice 12 10 Section 5 Plate Filling MIXES IN MICROTROF MIXES IN MICROTROF 9 3 MIXES IN MICROTROF is used to specify mixing of the MICROTROF contents before an aliquot is taken Mixing of a M
36. ange of possible sequences to customize your own complex serial dilution files 25 Section 7 Plate to Plate Transfer 7 PLATE TO PLATE TRANSFER 7 1 Overview Transferring an aliquot of sample from one plate to another is called plate to plate transfer or replica plating This technique allows for transfer of samples in a SOURCE plate to a RECEIVER plate for assay culture etc The steps required to perform the plate transfer are 1 Aspirate sample from source plate 2 Deliver sample to receiver plate 5 Replace used tip with fresh one and proceed to next sample row etc The template for Plate to Plate Transfer is File 65 which is used as a template to establish a user file which can be stored and used as desired and it can be as a template for additional serial dilution files 7 2 Types of Plate to Plate Transfer There are two general types of Plate to Plate Transfer 1 transfer any single row one plate to any other single row in another plate 2 transfer any single row in one plate to two or more rows in another plate The program allows you to specify the direction of the transfer The default direction is from plate 2 to plate 1 but the system also allows you to transfer from plate 1 to plate 2 so that either plate can be used as the DESTINATION plate As examples you can transfer from row A of plate 1 to row B of plate 2 or you can transfer from row A of plate 2 to rows A B and C of plate 1 This enables y
37. ater simulating sample The WellPro liquid handler allows you to specify the direction of the transfer You can designate either plate as the SOURCE or the DESTINATION plate In this case the transfer will be from row B of plate 1 to rows C and D of plate 2 1 Placea 24 across template on the table 2 Place a magazine of tips in the back cavity of the template 5 Place the source plate into the front cavity of the template This will be the source plate 4 Place an empty plate in the middle cavity of the template This will be the destination plate plate 2 5 Press file 63 and then enter to access File 63 6 Edit the file using the following keystrokes Display Before Editing Keystrokes Display After Editing PLATE TO PLATE STEP STEP TO EDIT FILE 63 TRANSFER DIRECTION NO STEP TRANSFER DIRECTION P2 TO P1 P1 TO P2 VOL IN WELL PLATE 1 0 50 VOL IN WELL PLATE 2 0 50 TRANSFER MODE 1 TO 1 1 TO SEVERAL SOURCE ROW A B C D E F GH DESTINATION ROWS A B C D E F GH STEP 50 ul STEP dul STEP NO STEP NO NO YES YES NO NO NO NO STEP P2 TO P1 P1 TO P2 VOL IN WELL PLATE 1 0 50 50 ul VOL IN WELL PLATE 2 0 50 TRANSFER MODE 1 TO 1 1 TO SEVERAL SOURCE ROW A B C D E F GH DESTINATION ROWS CD 957 Note in the next step the WellPro controller automatically calculates the acceptable range of the TRANSFER VOLUME by dividing the VOL IN WELL PLATE 1
38. b u S uQ as q dens 36 8 1 OVERVIEW a UAR ERE EA S mu Sa a Sasa 36 8 2 EDITING AN EMPTYING A PLATE FILE J a u u u u 36 8 3 STORING OVERWRITING THE FILE u u uu uuu u 39 8 4 RUNNING AN EMPTY A PLATE FILE a J a u u u uu u u u 39 d zz5idlb c M 40 9 1 OVERVIEW NN LT E 40 9 2 EDITING A TIME DELAY FILE J u u nete sete u u 40 9 3 STORING OVERWRITING THE FILE J u u u u uu u 42 9 4 USING A TIME DELAY FILE 1 u u u u u 42 10 LINKING FIEES Tu u P 43 10 1 bes Su Re eon Une 45 10 2 IBI eH dz alldcc sss 43 10 3 TYPICAL SERIES OF OPERATION USING LINKED FILES l l l u u 43 11 ADJUSTING THE TIP HEIGHT snn a a Nee eie eiae eere ettet Peto be Weaie eee tos ido
39. cal representative If internal parts are missing or damaged report this to the shipping company and your local representative as soon as the problem is observed es Note If there us any apparent damage to the system the user should investigate the nature of the damage before plugging the unit into the mains to ensure that powering up of the system will not create a hazardous condition or damage internal components If the damage appears significant call your local representative before connecting the unit to the mains 2 5 Initial Installation of the WellPro 384 Liquid Handler Assembly installation and final inspection of the WellPro 384 Liquid Handler will be done by the user To install the unit 1 Connect the Controller Cord to the Controller and the rear on the liquid handling unit 2 Place the power cord to the line into the socket into the rear panel of the liquid handling unit and plug the unit into the line Power up the unit The liquid handler will initialize and the controller will present Figure 2 1 The pipette tip will move to determine the zero If the head does not move contact your local service representative for assistance HOME START TO RUN FILE 0 Figure 2 1 The Keypad Controller Press the START button The pipette tip will move in the X Y and Z direction If the head does not move contact your local service representative for assistance Section 3 System Components 3 SYSTEM COMPONENTS
40. crements from this lower limit to 20 mL 60 mL or 80 mL for the front middle and rear reservoirs respectively for the MICROTROF reservoir After the file has been run the display will update the value to reflect the decreased amount of liquid in the reservoir CHANGE TIPS CHANGE TIPS YES NO YES CHANGE TIPS is used to indicate if the tips should be removed after the first liquid is dispensed and a new set of tips from the tip magazine should be placed on the pipette head thus decreasing carryover If you don t want to change tips then the same tips will be used throughout the entire operation LIQUID HANDLING SPEED LIQ HANDLING SPEEDS RAPID MEDIUM SLOW LIQUID HANDLING SPEEDS is used to select the rate sample delivery from the tips rapid medium or slow 14 Section 5 Plate Filling 15 LINK TO STORED FILE LINK TO STORED FILE 0 45 0 STOP 0 LINK TO STORED FILE is used to indicate what the system should do after it completes the file it is currently running If you want the system to stop at the end of the file enter O If you want the system to perform another file after the present file is complete enter the desired file number The LINK TO STORED FILE command permits more efficient operation because you don t need to start each individual file manually with the file A detailed discussion of Linked files is presented in Section 10 16 END OF FILE END OF FILE
41. d and tested to insure the proper volume delivery It is recommended that only qualified personnel be permitted to calibrate the instrument If recalibration becomes necessary you should call ProGroup Technical Service at 1 800 471 1916 12 2 The Gravimetric Method The gravimetric method of verification involves weighing each individual well containing water dispensed from the WellPro Liquid Handler The analytical balance used in the procedure should be well maintained calibrated with an appropriate set of National Institute of Science and Technology certified weights NIST Class S Weights NBS Handbook 105 1 Circular 547 or equivalent and should be capable of reading 0 001 g 0 0005 0 The gravimetric method also requires a thermometer which should be calibrated and readable to 0 5 C 49 Section 12 Calibration Procedures CAUTION CAUTION Since It is possible that water which has been added to the wells may evaporate during the calibration we strongly recommend that you perform this procedure by dispensing water into one row at a time then weighing the amount of water added to each well A row of 24 wells typically takes 10 minutes to weigh Repeat this procedure for each subsequent row until all sixteen rows have been weighed C Note The WellPro liquid handler has been calibrated in the operational range of 20 26 68 79 F Operation outside this range may affect the volumetric calibration and the system should be calibrated
42. de is selected in this file In that instance the SOURCE row specified acts as the single source row for transfer to several receiving or DESTINATION rows DESTINATION ROWS DESTINATION ROWS A B C D E F G H I J K L M N O P DESTINATION ROW is used to indicate which row of the DESTINATION plate is used to place liquid in during transfer This parameter only appears if the 1 to SEVERAL mode is selected in this file In that instance the SOURCE row specified acts as the single source row for transfer to several receiving or DESTINATION rows 29 16 17 18 19 Section 7 Plate to Plate Transfer LIQ HANDLING SPEEDS LIQ HANDLING SPEEDS RAPID MEDIUM SLOW LIO HANDLING SPEEDS is used to select the rate of sample delivery from the tips rapid medium or slow LINK TO STORED FILE LINK TO STORED FILE 0 45 0 STOP 0 LINK TO STORED FILE is used to indicate what the system should do after it completes the file it is currently running If you want the system to stop at the end of the file enter O If you want the system to perform another file after the present file is complete enter the desired file number The LINK TO STORED FILE command permits more efficient operation because you don t need to start each individual file manually with the file A detailed discussion of Linked files is presented in Section 10 END OF FILE END OF FILE RUN STORE END OF FILE
43. e display will present YES TO STORE USER zzzz FILE XX 4 Press YES to accept the assignment 6 4 Running a Serial Dilution File When you are ready to run a Serial Dilution 1 Select the desired file 2 Press START If the MICROTROF does not contain the required amount of liquid to perform the desired operation the message PLEASE FILL MICROTROF will appear on the screen This prompt allows you to simply enter the volume you are adding to the MICROTROF reservoir without requiring you to step back through each of the file s parameters At the conclusion of the operation the system will stop or proceed to the file that is linked to it 25 6 5 Section 6 Serial Dilution Sample Serial Dilution Experiment File 62 Serial Dilution 1 2 Ratio The following example will show you how to perform a simple 1 2 1 4 1 8 1 16 etc serial dilution on 5 uL samples in an 16 across microplate Each well of the plate will be pre filled with 25 uL of solution There will be four mixes in each well after every transfer 1 2 Fill an empty plate with 25 uL of diluent Place a 16 across template on the table Place the magazine of tips in the back cavity of the template Manually pipette 50 uL samples into row 1 of the plate Use water in this example Place the plate in the front cavity of the template with samples Press file 62 and then enter to access File 62 Edit the file using the following k
44. en file 65 or any other Time Delay file the opening step is presented and the step command should be selected to view the file on a screen by screen basis You can change the value of each parameter as desired to provide the desired program to fill the plates After a value is entered press ENTER To advance to the next step press STEP A detailed discussion of role of each key is presented in Chapter 4 1 OPENING STEP TIME DELAY STEP TO EDIT FILE 65 When File 65 is opened the OPENING STEP display is presented If you want to view edit the file press STEP 2 DELAY DELAY 0 MINS 0 SEC DELAY is used to indicate to the system to wait a specified interval after completing a file in a linked sequence before beginning the next file in that sequence You can set the length of the delay from 0 to 999 59 minutes seconds When the DELAY prompt first appears the underline will allow you to enter the minutes field When you press the Enter key the seconds field will be activated in the orig that have the delay parameter only is 0 999 and 0 99 DELAY 0 99 0 MIN 0 SEC 40 Section 9 Time Delay File 5 PAUSE WHEN LINKING PAUSE WHEN LINKING NO YES PAUSE WHEN LINKING is used to tell the system whether to pause and await re start after operator attention or whether it should continue on to the next file without halting This feature is useful for changing plates or tips addin
45. erring a sample For example when you are diluting cells which may settle you should agitate them back into suspension after transfer by mixing This parameter can be set from 0 to 9 0 indicates no mixing MIX VOLUME MIX VOLUME 0 25 25 ul MIX VOLUME is used to tell the system how much solution should be used in the Mix step Typically this volume is set to 50 of the total volume in the well to be mixed but will be more or less depending on the application For example when mixing cells it may be desirable to use a lower percentage of the total volume The value of this parameter may not exceed 75 of the total volume in the well you are mixing The system will automatically calculate this amount and indicate it on the display as the upper limit in the range This parameter can be set in 1 uL increments from 0 uL to the upper limit CHANGE TIPS CHANGE TIPS YES NO CHANGE TIPS is used to indicate if the tips should be removed after the first liquid is dispensed and a new set of tips from the tip magazine should be placed on the pipette head thus decreasing carryover If you don t want to change tips then the same tips will be used throughout the entire operation 28 11 12 15 14 15 Section 7 Plate to Plate Transfer PLATE 1 TYPE PLATE 1 TYPE FLAT U V HEIGHT TUBE PLATE 1 TYPE is used to indicate the design of the bottom of plate 1 This information is used to determi
46. establish a file for a specific operation e Open the template file that corresponds to the action that you want to perform e Edit the method to meet the specific requirements of your operation e Save the file As an example if you want establish a file to be used to add 20 uL to each well in a plate open File 67 which is the template file for adding fluid to a plate edit the file as appropriate and save it using a different file number e g File 27 When you are ready to run the operation simply recall the file and press Start When a file is executed the steps are performed in the indicated order In this chapter we describe the keypad controller and explain the general approach to file editing Discussion of the individual file templates is presented in Chapters 5 9 42 The Keypad Controller 4 2 1 The Display When the system is powered up the display will be as shown in Figure 4 1 The nature of the information on the display will depend on the status of the system as described below HOME START to run file 0 Figure 4 1 The Keypad Controller Section 4 Establishing a File using the Controller Figure 4 2 The Controller 4 2 2 The Keypad The controller includes ten alphanumeric keys and nine control keys The general role of the control keys is described below For many keys the action is dependent on the status of the program 4 2 3 Clear 1 Removes information that you have entered on the disp
47. eystrokes 24 Section 6 Serial Dilution Display Before Editing Keystrokes Display After Editing SERIAL DILUTION STEP STEP TO EDIT FILE 62 VOL IN WELL AT START 25 ENTER VOL IN WELL AT START 0 50 25 ul 0 50 25 ul TRANSFER VOLUME TRANSFER VOLUME 0 50 5 STEP 0 50 5ul MIXES BEFORE TRANSFER STEP MIXES BEFORE TRANSFER 0 9 0 9 5 MIX AFTER TRANSFER STEP MIX AFTER TRANSFER 0 9 0 9 5 MIX VOLUME MIX VOLUME 0 5 0 5 Sul CHANGE TIPS STEP CHANGE TIPS YES NO YES NO PLATE TYPE NO PLATE TYPE FLAT U V HEIGHT TUBE STEP FLAT U V HEIGHT TUBE PLATE ORIENTATION NO PLATE ORIENTATION 24 16 WELLS ACROSS STEP 24 16 WELLS ACROSS DILUTE ROWS 1 2 5 STEP DILUTE ROWS 1 2 5 4 5 6 7 8 9 10 11 12 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 13 14 15 16 17 18 19 20 21 22 23 24 20 21 22 23 24 EMPTY LAST ROW TO STEP EMPTY LAST ROW TO OFF 1ST ROW MAG TROF3 OFF 1ST ROW MAG TROF3 LIQ HANDLING SPEEDS STEP LIQ HANDLING SPEEDS RAPID MEDIUM SLOW RAPID MEDIUM SLOW LINK TO STORED FILE STEP LINK TO STORED FILE 0 45 0 STOP 0 0 45 0 STOP 0 END OF FILE ENTER RUN STORE SERIAL DILUTION START START TO RUN FILE 62 Serial dilutions can be very cumbersome and time consuming when performed manually The WellPro liquid handler has the capability to perform very complex serial dilution sequences As you gain familiarity with the WellPro liquid handler and especially the powerful FILE LINKING features experiment with the r
48. g instructions for the WellPro Model 384 Liquid Handler Read the manual in its entirety prior to operating the instrument This User Manual has been designed to give you the information you need to Review safety precautions e Install the WellPro Model 384 Liquid Handler e Use the WellPro Model 384 Liquid Handler in routine analyses and research e Perform basic maintenance procedures This User Manual also describes features and specifications of the WellPro Model 584 Liquid Handler hardware and on board software WellPro User Manual 01 11 04 iv Table of Contents TABLE OF CONTENTS SAFETY SYMBOLS AND MARKINGS Il 6 tee IV TABLE OR CONTENTS eti cores a i o S DE EE Run u Qu u V 1 INTRODUCTION u do derent ettet itte tn tet aaa OR HE EUN IR ETE 1 1 1 OVER E 5 eoe coe tele ue ete ve EE 1 1 2 PRINCIPLE OF OPERATION u u u sns ense u sess ss a 2 1 2 1 c EE 2 1 2 2 Liquid Handling BUT e RR 2 1 2 5 WellPro Reagent 2 4 2 1 2 4 WellPro Templates
49. g reagents etc 4 LINK TO STORED FILE LINK TO STORED FILE 0 45 0 STOP 0 LINK TO STORED FILE is used to indicate what the system should do after it completes the file it is currently running If you want the system to stop at the end of the file enter O If you want the system to perform another file after the present file is complete enter the desired file number The LINK TO STORED FILE command permits more efficient operation because you don t need to start each individual file manually with the file A detailed discussion of Linked files is presented in Section 10 5 END OF FILE END OF FILE RUN STORE END OF FILE is used to indicate what the system should do after you have completed the editing of the file e f you select RUN the file will start e If you select STORE the file will be copied into memory see Section 5 3 for additional information about storing files 887 Note You also review the file you have just made by continuing to press the STEP key 41 Section 9 Time Delay File 9 3 Storing Overwriting the File To store a new file 1 At the END OF FILE statement item 5 Section 8 2 select STORE The display will appear as shown below FILE STORE ENTER USER 2 Enter your user number or project number this number is provided for reference only and does not affect the use of the system then press ENTER The display will indicate FIL
50. he tips for the MICROTROF container that you want the WellPro liquid handler to select first The WellPro uses the same counter to keep track of which tips have been used whether you have specified tip changes or not Accordingly each time the WellPro liquid handler links to the next file in a chain it sequences through the tip magazine to select the next row of fresh uncontaminated tips Likewise if you start a new file or restart the same file the counter automatically increments causing the WellPro liquid handler to select the next row of unused tips When the last row of tips row A has been used the WellPro liquid handler stops automatically and displays the ENTER FIRST TIP ROW prompt again Change the magazine and specify the row of tips that you want to start with usually row H for the magazine or row J if you have also changed the tips for the MICROTROF container 10 3 A Typical Series of Operation Using Linked Files This section describes the use a linked file in a typical scenario In this example 1 Asample will be diluted File 21 2 Asmall fraction of the diluted sample will be transferred to a new plate File 22 5 Areagent is added to each well File 54 File 21 is a user customized Serial Dilution file The last line in the file indicates that the operation should be transferred to file 22 at the termination of File 21 LINK TO STORED FILE 0 45 0 STOP 22 45 Section 10 Linking Files
51. ion This section describes how to verify the accuracy and precision of the performance of the WellPro 584 Liquid Handler using a gravimetric procedure which utilizes the weight of water to determine the volume dispensed While this method is the most accurate and controllable method other methods could be used Alternative methods which could be used involve the transfer of e asolution of chromophore such as potassium dichromate and the determination of the absorbance e asolution of fluorescein or rhodamine and the determination of the fluorescence e asolution of a radioisotope and the determination of the radioactivity These methods are more difficult to perform and are subject to variations in detection therefore they are to be considered secondary and are not recommended The frequency of calibration is dependent on the needs of your facility and is based on parameters such as the types of samples used how frequently the WellPro liquid handler is used and the maintenance program you have instituted When you are using a new WellPro liquid handler you should test eight samples from each tip to determine precision and accuracy Subsequent verification can be carried out using the procedures in the Quality Control section Section 12 6 We recommend that you maintain a log of calibration reports as this collected data will help you and WellPro Technical Service to identify any problems The WellPro Liquid Handler is factory calibrate
52. is presented and the step command should be selected to view the file on a screen by screen basis You can change the value of each parameter as desired to provide the desired program to fill the plates After a value is entered press ENTER To advance to the next step press STEP A detailed discussion of role of each key is presented in Chapter 4 1 OPENING STEP SERIAL DILUTION STEP TO EDIT FILE 62 When File 62 is opened the OPENING STEP display is presented If you want to view edit the file press STEP 2 VOLIN WELL AT START STEP VOL IN WELL AT START 0 350 20 ul VOLUME IN WELL AT START is used to tell the system how much diluent is in each well before any transfer has been made This parameter does not pertain to the first row which contains the sample The system uses this number to determine how far down into the wells the tips should go and also to prevent the instrument from attempting to withdraw more sample than is actually present This parameter be set from 0 to 50 uL in 1 uL increments screen says 0 350 but text says 50 8 TRANSFER VOLUME TRANSFER VOLUME 0 100 5 ul TRANSFER VOLUME indicates how much sample should be withdrawn from the original well for transfer to the new well during a serial dilution This parameter can be set from O to 50 uL in 1 uL increments A transfer volume of 0 may be used if you wish to simply mix the contents of the well 19 Sec
53. l appear on the screen This prompt allows you to simply enter the volume you are adding to the MICROTROF reservoir without requiring you to step back through each of the file s parameters At the conclusion of the operation the system will stop or proceed to the file that is linked to it 16 5 5 Section 5 Plate Filling Sample Plate Filling Experiment File 61 Plate Filling The following example will show you how to perform a plate filling operation using a 24 across template The plate will be in position 1 Each well of the plate will be pre filled with 25 uL of solution There will be four mixes in each well after every transfer 1 2 Fill an empty plate with 25 uL of diluent Place a 24 across template on the table Place the magazine of tips in the back cavity of the template Place the plate in the front cavity of the template with samples Press file 61 and then enter to access File 612 Edit the file using the following keystrokes 17 Section 5 Plate Filling Display Before Editing Keystrokes Display After Editing PLATE FILLING STEP STEP TO EDIT FILE 61 PLATE POSITION STEP 1 1 VOL IN WELL AT START 25 ENTER VOL IN WELL AT START 0 50 25 ul 0 50 25 ul DELIVER FROM 1 ENTER DELIVER FROM T1 T2 T3 T T2 T3 MIXES IN MICROTROF 5 ENTER MIXES IN MICROTROF 9 3 5 MICROTROF MIX VOLUME STEP MICROTROF MIX VOLUME 0 200 50ul 0 3 3 ul DELIVERY VOLUME DELIVERY VOLUME 0 200 50
54. l they are level with the top of the tubes Record the TIP NOW AT number on a worksheet as the TUBE HEIGHT 557 Note Note the maximum number in the acceptable range at the lower left of the display Subtract the TIP NOW AT number from the maximum that is indicated The resulting value is very important This difference allows you to determine which operations you can safely carry out with the tubes in question Figure 10 2 defines the operations you can and cannot carry out for differences in particular ranges D is the difference between two values The height of tube as it stands in the rack and the maximum number shown in the range for the TIP NOW AT prompt of File 70 Tip Height File 46 Section 11 Adjusting the Tip Height Fig 11 2 Tube Heights Using Tall or Narrow Tubes If your tubes are very tall and narrow it is possible that the tips may not be able to reach the bottom of the tubes without forming a seal with the tube or the fins on their upper edges may strike the top of the tubes In the same vein if the bottom of the tube is a sharply angled V the tips may get stuck in the tubes ARTIFICAL 20041 HEIGHT 77271 tl E ARTIFICAL _ __ WELL VOLUME VOID VOLUME QA terete ie aah Fig 11 3 Tall and Narrow Tubes If any of these problems are observed it will be necessary to establish an artificial WELL BOTTOM HEIGHT above this point as described below 1
55. late Manually pipette 50 ul of water into each of the 384 wells of plate 26 Place plate 2 in the middle cavity of the template Place the calibration plate containing the pre weighed wells in the front cavity of the template Access File 65 by pressing File 65 and Enter Edit File 65 as described below 51 Section 12 Calibration Procedures Display Before Editing Keystrokes Display During Review PLATE TO PLATE STEP STEP TO EDIT FILE 63 TRANSFER DIRECTION P2 TO P1 P4 TO P2 VOL IN WELL PLATE 1 0 50 VOL IN WELL PLATE 2 0 50 50 UL TRANSFER MODE 1 TO 1 1 TO SEVERAL TRANSFER VOLUME 0 50 50 UL MIXES BEFORE TRANSFER 0 9 o MIXES AFTER TRANSFER 0 9 9 CHANGE TIPS YES NO PLATE 1 TYPE FLAT U V HEIGHT TUBE PLATE 2 TYPE FLAT U V HEIGHT TUBE TRANSFER ROWS A B C D E F G H IJ K L M N O P LIQ HANDLING SPEEDS RAPID MEDIUM SLOW LINK TO STORED FILE 0 45 0 STOP 0 END OF FILE RUN STORE PLATE TO PLATE START TO RUN FILE 63 STEP STEP STEP STEP START STEP STEP STEP 0 50 STEP 40 ENTER STEP STEP STEP STEP ENTER 52 TRANSFER DIRECTION P2 TO P1 P1 TO P2 VOL IN WELL PLATE 1 0 50 OUL VOL IN WELL PLATE 2 50 UL TRANSFER MODE 1 TO 1 1 TO SEVERAL TRANSFER VOLUME 0 100 40 UL MIXES BEFORE TRANSFER 0 9 9 MIXES AFTER TRANSFER 0 9 9 5 YES NO PLATE 1 TYPE FLAT U V HEIGHT TUBE PLATE 2 TYP
56. lay before it is entered into the WellPro controller This key is most commonly to erase information and start over when you are editing a file 2 Erases an entire user created file by pressing the File key the appropriate number keys the Enter key the File key and the Clear key in that order 4 2 4 Start 1 Instructs the system to start to carry out the instructions and conditions in the active file 2 Restarts a file that has been paused by the user before completion The file will start where it left off when it was paused Section 4 Establishing a File using the Controller 4 2 5 Step The Step key has several uses At the start of a file Step indicates that you want to edit or review the file While editing a file Step is used to indicate that the present setting for the parameter is acceptable and you want to see the next parameter When the End of File message is displayed start of the file will be presented for editing or review After you have pressed Stop during the execution of a file the activity indicated in the next line in the file will be performed 4 2 6 Stop The Stop Function has several uses If you are editing a file the END OF FILE message will be presented At this point you have two choices If you press Enter and the press Start the file will be run If you press Stop again you will be able to enter a new file If you are running a file the WellPro liquid handler will pause stop the present
57. m 1 uL to 50 pL using disposable tips Perform serial dilutions in almost any ratio Perform serial dilutions of 16 wells or 24 wells simultaneously Precisely mix the contents of a microplate Transfer samples to or from microplates Perform mother daughter repli plating Add a reagent to the wells on a plate at a user specified time Fig 1 1 WellPro WP384 Liquid Handler Section 1 Introduction 1 2 Principle of Operation The WellPro WP384 Liquid Handler consists of a hand held controller and a liquid handling unit A variety of reagent containers can be used to hold liquids that will be added to the cells in a plate or have been removed from a plate 1 2 1 Controller The WellPro WP384 Liquid Handler includes a keyboard and display that are used to program all operations of the unit The controller allows the user to control parameters such as the volumes to transfer mix or dispense the transfer speed and whether to change tips between transfers 1 2 2 Liquid Handling Unit The WellPro WP384 Liquid Handler is an electro mechanical transfer unit that is comprised of two basic parts a table and a vertical head assembly The table accommodates a series of interchangeable templates which position the reagent containers and a magazine with the disposable tips etc During operation the table slides back and forth to bring the appropriate component under the head assembly The head assembly consists of three basic parts a
58. ne the height of the liquid in each well and thus calculate how far down into the wells the tips should go If you are not using a standard microplate select HEIGHT and determine the amount to raise or lower the tip height by using File 70 the Tip Height File Section x xx The height parameter can be set from 100 to 3500 in increments of 1 PLATE 2 TYPE PLATE 2 TYPE FLAT U V HEIGHT TUBE PLATE 2 TYPE is used to indicate the design of the bottom of plate 2 This information is used to determine the height of the liquid in each well and thus calculate how far down into the wells the tips should go If you are not using a standard microplate select HEIGHT and determine the amount to raise or lower the tip height by using File 70 the Tip Height File Section x xx The height parameter can be set from 100 to 3500 in increments of 1 SOURCE ROW SOURCE ROW A B C D E F G H I J K L M N O P SOURCE ROW is used to indicate which row of the SOURCE plate is used to withdraw samples from This parameter only appears if the 1 to SEVERAL mode is selected in this file In that instance the SOURCE row specified acts as the single source row for transfer to several receiving or DESTINATION rows TRANSFER ROW TRANSFER ROW A B C D E F G H I J K L M N O P TRANSFER ROW is used to indicate which row of the TRANSFER plate is used to place liquid in during transfer This parameter only appears if the 1 to SEVERAL mo
59. nei 45 11 1 OVERVIEW ias NTIS RTS 45 11 1 1 Editing Running the Tip Height Adjustment 2 22 2 2 2222 45 11 1 2 Setting the Tip Height 1 1 46 12 CALIBRATION PROCEDURES 42221 49 12 1 INTRODUCTION a am eo decore etr Aer petto ee reve eue eve pe ou vues teen Bee No e Ute uo 49 12 2 THE GRAVIMETRIC METHOD J estes ense u uu u 49 12 2 4 u LLL u cases u u e e tect eer Sau uay euer us 50 12 3 CALIBRATION PROCEDURE J u uu uuu u u u 51 12 4 CALCULATIONS 5 Cana A u AEO ua aa EVEN 53 12 5 CALIBRATION OF NON AQUEOUS OR VISCOUS SOLUTIONS 55 42 54 INtFOd ctIi n cuu u T 55 42 5 2 Procedure 55 12 6 QUALITY CONTROL PROCEDURES a u u asas asan Neve ETETE E EAI TANE ESENCE aE 57 12 6 1 Verification RR 58 WellPro User Manual 01 11 04 vi Table of Contents 13 OPERAT
60. nts that may damage the cover Decontamination may be performed by chemical means Chlorine bleach is an acceptable decontamination solution Ethanol and gluteraldehyde should not be used on the plastic surfaces DO NOT AUTOCLAVE 60 Section 13 Operating Hints and Maintenance 13 4 6 Fuse Replacement Turn the power switch off and disconnect the power cord from the power outlet before replacing the fuse This procedure will insure against electrical shocks For continued protection against electrical overload and fire hazard replace the fuse only with one of the same type and rating 5 amps 250V Normal blow 13 5 Troubleshooting If you encounter difficulties in the use of the instrument please contact your local ProGroup Technical Service Department A technical service representative may be able to resolve the problem over the telephone using a basic check out procedures If the problem cannot be resolved in this manner the technical service representative will arrange for a field service representative to visit your lab to repair the unit If the unit cannot be repaired in your lab arrangements will be made to return the unit to ProGroup for factory repair Only qualified personnel should attempt service or repair work on the WellPro operating circuits or mechanism 61 Section 13 WellPro WP384 Specifications 14 WELLPRO WP384 SPECIFICATIONS Dimensions Controller Liquid Unit Height 2 1 2 16 Width 6 14 12 Dep
61. of liquid above the artificial well bottom level So for example if the void volume of a set of tubes was 100 ul and the tube contained 1700 Hl all the VOLUME IN WELL AT START would be1600 File 70 Tip Height The following example shows how to determine experimentally the two numbers required to edit a file so that a microtitration plate other than a standard microplate can be used 1 Placea 24 across template on the table 2 Place the magazine in the back cavity of the template 3 Accurately fill row P of a microtitration plate with 50 ul of water 4 Place the plate in the middle cavity of the template 5 Press file 70 and then enter to access File 70 6 Press start 7 When NOW AT is displayed press 2800 and then enter This will bring the tips down to the plate 8 Press the f yes or 4 no keys to raise or lower the tips until they just touch the meniscus of the water Record the TIP NOW at number on a photocopy of the Tip Height Worksheet as the 50 ul Height 9 Press the J no key or enter numbers until the tips are just above the bottom of the wells Record the TIP NOW at number on the worksheet as the WELL BOTTOM HEIGHT 10 Press the stop key and the display will read HEIGHT COMPLETE FILE 70 11 Press file the new file number that you want to use and then enter 48 Section 12 Calibration Procedures 12 CALIBRATION PROCEDURES 12 1 Introduct
62. of the solution using the following formula d weight volume 5 Perform a shortened version of the Calibration Procedure at the temperature and with the file sequence you plan to use routinely Weigh the wells of Row B at this same temperature 4 Calculate the arithmetic mean average weight W by adding all sample weights together and dividing by the number of samples 24 5 Calculate the volume dispense by dividing the mean weight dispensed by the density 6 Calculate the standard deviation SD of the individual volumes using equation 12 4 SD xXIWI WP x 1 YlVi Vb 12 4 25 density 25 7 Calculate the coefficient of variation CV using equation 12 5 CV SD x 100 12 5 mean volume 8 If desired a volume correction can be made by modifying the volume setting entered into WellPro liquid handler when creating File for example if 46 uL is dispensed for a viscous solution when 50 uL are anticipated enter 54 uL 50 x 50 46 into the WellPro controller to compensate for the change The Non Aqueous or Viscous Solution Worksheet presented as Table 12 5 can be used to assist in calculating the coefficient of variation This worksheet can be reproduced as desired for calibration activities 55 Section 12 Calibration Procedures Table 12 5 Non Aqueous or Viscous Solutions Date Solution Temperature Tech Density Volume Weighed D Density Weight Volume Weight WellPro Weighing Row B 10 11
63. on is used to determine the height of the liquid in each well and thus calculate how far down into the wells the tips should go If you are not using a standard microplate select HEIGHT and determine the amount to raise or lower the tip height by using File 70 the Tip Height File Section 11 The height parameter can be set from 100 to 3500 in increments of 1 15 11 12 15 14 Section 5 Plate Filling FILL PLATE ROWS FILL PLATE ROWS A B C D E F G H I J K L M N O P FILL PLATE ROWS is used to indicate the rows of the plate to fill and which to leave unfilled Rows are designated by letters A through P To select row A press YES The underline will remain and B will be active if you do not want to use row press NO to advance to the next row If you want all rows to be filled press the STEP key After the last NO selection has been made press STEP i e if you want all rows except for A press NO for A The active row will become B If you press STEP rows B P will be active MICROTROF VOLUME AT START MICROTROF VOL AT START 20 80 77ml MICROTROF VOLUME AT START is used to indicate how much liquid is in the reservoir you have chosen before any liquid has been removed The system uses the delivery volume and the number of rows to be filled to calculate the minimum volume needed to run the file and indicates it on the display as the lower limit You can set the value of this parameter in 1 mL in
64. ou to set up an assay plate to do duplicates triplicates etc The TRANSFER MODE DIRECTION is a feature that is especially useful when it is combined with FILE LINK because it allows you to transfer duplicate samples from the rows of one plate to another plate Thus using these functions in combination makes it possible to transfer from row A of plate 1 to rows A and B of plate 2 then automatically continue with another file which transfers from row B of plate 1 to rows C and D of plate 2 and so on TRANSFER MODE DIRECTION also allows you to use a 384 well plate as a reagent MICROTROF to deliver multiple reagents to a single assay plate You can place the same or different reagents in each of the 24 columns and or each of the 16 rows of the source plate These reagents can be delivered to specified rows on the destination plate This arrangement minimizes reagent waste When you use TRANSFER MODE DIRECTION feature with the TUBE HANDLING feature option you can carry out transfers from larger containers such as micro test tubes or segmented troughs and use them as reagent containers Using the help key enables you to extend the acceptable range for TRANSFER VOLUME and VOLUME IN WELL AT START so you can make full use of the larger volume capacity of these containers 26 Section 7 Plate to Plate Transfer 7 3 Editing a Plate to Plate Transfer File When you open file 63 or any other Serial Dilution file the opening step is presented and
65. plunger housing which contains nozzles that press into the tips to form airtight seals e aplunger block which moves up and down to aspirate and deliver samples atip ejector plate which removes used tips The head assembly slides up and down to bring the tips to the appropriate height to perform the designated operations In most instances a 24 channel tip air displacement pipetting head is used on the WellPro WP384 Liquid Handler and most protocols are designed to be performed in the 24 channel mode but serial dilutions can be performed in the 16 channel mode to allow for maximum dilution capability with the 584 well plate 1 2 3 WellPro Reagent Containers The WellPro liquid handler can be used with a variety of MICROTROF reagent containers each of which consists of three reservoirs Each reservoir holds a different amount of liquid e g 20 mL 58 mL and 77 mL You can use the reservoirs to hold reagents that will be added to a plate or waste that has been removed from a plate In addition some MICROTROFs include a station for storage of microtips Section 1 Introduction 1 2 4 WellPro Templates The WellPro WP384 liquid handler uses a template to position microplates reagent containers and tip magazines on the movable table Two templates are provided Standard Template 24 Across A general purpose template Template 1 24 Across is used for liquid handling operations when the 24 well rows on the plate are oriented
66. potential is set at the factory and is indicated on the serial number tag on the rear panel of the system WARNING WARNING Verify that this voltage is correct for your location before installation CAUTION CAUTION WellPro 384 Liquid Handlers shipped to locations in the United States and Canada are supplied with a standard three conductor power cable and a three terminal plug that provides an earth ground The unit MUST be connected to a three terminal socket that is properly wired according to the U S National Electrical Code and is grounded to a true earth ground The use of an adapter to a two terminal outlet is PROHIBITED Section 2 Installation Procedures 24 Unpacking and Inspection The WellPro 384 Liquid Handler is shipped in one carton that contains the items listed in Table 2 1 When the system is received carefully unpack the unit and verify receipt of all components We recommend that you save the shipping container and packing material in case you need to move or ship the instrument Table 2 1 PACKING LIST Quantity Description Part Number 1 WellPro WP384 Automated Liquid Handler Controller Controller Cord Plastic Cover Operators Manual Microtrof Pipette Tip Rack Template Horizontal Format 24 Across Sey TT iMa Template Vertical Format 16 Across If there is external damage to the shipping carton report the damage to the shipping company and your lo
67. prior notice as part of a continuous product development Although this manual has been prepared with every precaution to ensure accuracy ProGroup assumes no liability for any errors or omissions nor for any damages resulting from the application or use of this information This manual supersedes all previous editions No liability for consequential damages ProGroup shall not be liable for any damages whatsoever arising out of the use or inability to use this product Printed in the United States Safety Sybols and Markings SAFETY SYMBOLS AND MARKINGS These symbols are intended to draw your attention to essential information and alert you to the presence of hazards as indicated Some of these symbols may not appear in the manual or on the product SAFETY SYMBOLS used in the WellPro Model 384 Liquid Handler Power ON Power OFF WARNING MARKINGS used in the documentation Caution risk of electric shock Caution biohazard risk Caution risk of personal injury to the operator or a safety hazard to the surrounding area Caution risk of damage to the instrument other equipment or loss of performance or function in a specific application P P P P WellPro User Manual 01 11 04 About the User Manual ABOUT THE USER MANUAL This User Manual has been written for the user e g laboratory technician and provides information on the WellPro Model 384 Liquid Handler This manual contains the installation and operatin
68. resent file is complete enter the desired file number The LINK TO STORED FILE command permits more efficient operation because you don t need to start each individual file manually with the file A detailed discussion of Linked files is presented in Section 10 END OF FILE END OF FILE RUN STORE END OF FILE is used to indicate what the system should do after you have completed the editing of the file If you select RUN the file will start and step 14 will be presented If you select STORE the file will be copied into memory see Section 6 3 for additional information about storing files ENTER FIRST TIP ROW ENTER FIRST TIP ROW A P P ENTER FIRST TIP ROW is used to specify which row of tips to use Rows of the tip magazine are designated by letters A through P back to front 22 Section 6 Serial Dilution 6 3 Storing Overwriting the File To store a new file 1 At the END OF FILE statement item p Section 5 2 select STORE The display will appear as shown below FILE STORE ENTER USER 2 Enter your user number or project number this number is provided for reference only and does not affect the use of the system then press ENTER The display will indicate FILE STORE ENTER FILE XX Where xx is the file number assigned by the system which will be the lowest unoccupied file If you want to assign a different number enter it 8 Press ENTER Th
69. ress Enter the range says 350 e The user must choose one of a set of discrete values or selections As an example the PLATE TYPE step Figure 4 4 is used to tell the WellPro liquid handler whether your plate has a FLAT U shaped or V shaped bottom The WellPro liquid handler uses this information to determine the height of the liquid in each well and thus how far down into the wells the tips should go One and only one value is used for a given plate PLATE TYPE FLAT U V HEIGHT TUBE Fig 4 4 The Plate Height Step To select the desired option press the YES or NO key until it the desired type is highlighted then press Enter 44 File 0 File 0 which is presented when the unit is powered up is used to initialize the system 11 Section 5 Plate Filling 5 PLATE FILLING 5 1 Overview Plate filling refers to the addition of a user specified quantity of liquid to the wells in a plate The template for plate filling is File 61 which is used to establish a user file which can be stored and used as desired and can be as a template for additional plate filling files 5 2 Editing a Plate Filling File When you open file 61 or any other plate filling file the opening step is presented and the step command should be employed to view the file on a screen by screen basis You can change the value of each parameter to provide the desired program to fill the plates After a value is entered press ENTER To
70. th 654 28 Weight 70 Ibs 30 2kg Power Requirements 120 220 240 VAC 47 63 Hz 2 amps at 120 VAC 1 amp at 240 VAC Fuse 3 amps 250V Normal blow Room Temperature 23 3 C 68 79 Volume Range 0 to 50 ul programmable in 1 ul increments Accuracy Typically 50 ul 5 pl 10 pl 1 ul with phosphate buffered saline at 25 Precision Single and multi channel head typical coefficient of variation less than 1 at 10 ul with deionized water at 23 C Time Delay Programmable from 1 second to 999 minutes in 1 second steps Accurate to 1 part in 1000 or better over ambient temperature range 384 Well Microtitration Plates 384 well 24 X 16 5 X 3 microtitration plates typically used 62
71. the Tip Height Adjustment File When you open file 70 the opening step is presented and the Start command should be selected to view the file on a screen by screen basis You can change the value of each parameter as desired to provide the desired program to fill the plates After a value is entered press ENTER To advance to the next step press STEP A detailed discussion of role of each key is presented in Chapter 4 45 Section 11 Adjusting the Tip Height 1 HEIGHT HEIGHT START TO RUN FILE 70 Press Start to initiate the file 2 SELECT FUNCTION SELECT FUNCTION TIPS WASH HEAD Select Function is used to select measuring plate dimensions for use with the disposable tips 8 TIP NOW AT xxxx NOW 20 3500 GO TO TIP NOW AT is used to raise lower the tips so that the tip is in a position that approximates the position you are trying to determine You can set this parameter is desired from 20 to 3500 in increments of 1 Each step is equivalent to lowering the tip by approximately 0 00124 inch As an alternative you can use the zi o OR T keys to either raise or lower the tips ten steps at a time 0 0124 inch in order to fine adjust the position of the tips When the tip is properly adjusted press Enter to present the Tip Height screen 4 TIP HEIGHT TIP HEIGHT COMPLETE FILE 70 11 1 2 Setting the Tip Height Lower the tips unti
72. the step command should be selected to view the file on a screen by screen basis You can change the value of each parameter as desired to provide the desired program to fill the plates After a value is entered press ENTER To advance to the next step press STEP A detailed discussion of role of each key is presented in Chapter 4 1 OPENING STEP PLATE TO PLATE STEP TO EDIT FILE 63 When File 63 is opened the OPENING STEP display is presented If you want to view edit the file press STEP 2 TRANSFER DIRECTION TRANSFER DIRECTION P2 to P1 P1 to P2 TRANSFER DIRECTION is used to indicate whether the system should transfer from plate 2 to plate 1 or from plate 1 to plate 2 i e either plate 1 the plate in the front cavity of the template or plate 2 the plate in the middle cavity can be used as the source plate 5 VOLIN WELL PLATE 1 VOL IN WELL PLATE 1 0 50 Qul VOLUME IN WELL PLATE 1 is used to indicate how much liquid is in each well of plate 1 The system uses this value to determine how far down into the wells the tips should go and also to prevent exceeding the volume capacity of the wells This parameter can be set from 0 to 50 uL in 1 uL increments 4 VOLIN WELL PLATE 2 VOL IN WELL PLATE 2 0 50 50 ul VOLUME IN WELL PLATE 2 is used to indicate how much liquid is in each well of plate 2 The system uses this value to determine how far down into the wells the tips
73. tion 6 Serial Dilution MIXES BEFORE TRANSFER MIXES BEFORE TRANSFER 0 9 3 MIXES BEFORE TRANSFER is used to indicate the number of aspiration delivery cycles that should be performed before transferring a sample As an example if you are diluting cells which may settle the MIX BEFORE TRANSFER step might be used to agitate them back into suspension before transfer by mixing This parameter can be set from O to 9 0 indicates no mixing MIX VOLUME MIX VOLUME 0 75 Sul MIX VOLUME is used to tell the system how much solution should be used in the Mix step Typically this volume is set to 50 of the total volume in the well to be mixed but will be more or less depending on the application when mixing cells it may be desirable to use a lower percentage of the total volume The value of this parameter may not exceed 75 of the total volume in the well you are mixing The system will automatically calculate this amount and indicate it on the display as the upper limit in the range This parameter can be set in 1 uL increments from 0 to the upper limit CHANGE TIPS CHANGE TIPS YES NO YES CHANGE TIPS is used to indicate if the tips should be removed after the first liquid is dispensed and a new set of tips from the tip magazine should be placed on the pipette head thus decreasing carryover If you don t want to change tips then the same tips will be used throughout the entire operation PLAT
74. trol data The chart is a sheet of rectangular coordinate graph paper marked with days on the horizonal axis and assay units on the vertical axis The mean value as well as the limits of acceptable error are indicated on the chart and are based on past laboratory experience typically mean x 2 standard deviations Plot the daily value on the control chart so you can easily see any trends or any point falling beyond the limits of acceptable error Figure 12 1 Levey Jennings Control Chart 57 Section 12 Calibration Procedures 12 6 1 Verification Verification of the WellPro liquid handler calibration without the contribution of the reagents can also be part of the quality control system in your laboratory Weigh only Row A wells or a T of wells consisting of Row A and Column 6 as illustrated in Figure 12 2 and construct a Levey Jennings control chart using the mean average values for the Row and for the Column 58 Section 13 Operating Hints and Maintenance 13 OPERATING HINTS AND MAINTENANCE 13 1 Overview The WellPro 384 Liquid Handler is designed to be used with a minimum of maintenance In this chapter we describe routine operations that can be performed by the operator CAUTION CAUTION Do not open the Backpack Electronics or disassemble the controller unit There are no operator serviceable components in the instrument other than those describe the SERVICE AND MAINTENANCE and OPERA TING INSTRUCTIONS sections
75. ur user number or project number this number is provided for reference only and does not affect the use of the system then press ENTER The display will indicate FILE STORE ENTER FILE XX Where xx is the file number assigned by the system which will be the lowest unoccupied file If you want to assign a different number enter it 8 Press ENTER The display will present YES TO STORE USER zzzz FILE XX 4 Press YES to accept the assignment 84 Running an Empty a Plate File When you are ready to run an Empty a Plate file 1 Select the desired file 2 Press START At the conclusion of the operation the system will stop or proceed to the file that is linked to it 59 Section 9 Time Delay File 9 TIME DELAY FILE 9 1 Overview A Time Delay file allows you to insert a delay between two files that have been linked into a sequence This time delay can be specified from 1 second in length up to several hours Extremely long time delays are possible by linking two or more delay files in sequence This lets you more fully automated the liquid handling portions of assays As an example of the utility of this feature you can direct the system to add a reagent wait a specified period of time and then add another reagent You can also use a plate to plate transfer to start a reaction pause to incubate and then use a plate fill operation to stop the reaction 9 2 Editing a Time Delay File When you op

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