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Biotek EL800, Operator'S Manual

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Biotek EL800 Operator'S Manual Download Page 106

80  

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Chapter 3: Operation

 

 

BioTek Instruments, Inc. 

 

C  U  R  V  E  -  F  I T   T Y P E :   C - S P L  I  N  E 

 

  C  -  S  P  L  I N E  

 

  P T - P T   * M  O  R  E 

• 

Linear curve fit:

 A simple best-fit straight line is plotted using the 

values of the standards. 

• 

Quadratic or “Quad” curve fit:

 A curve fit that uses the quadratic 

equation “ax

2

 +bx +c = y” to plot the standard’s values. Utilizing this 

curve, any data point for a standard that deviates from the ideal value 
will not affect the entire curve. 

• 

Cubic curve fit:

 A curve fit that uses the equation  

“ax

3

 + bx

2

 + cx + d = y” to plot the standard’s values. This type of curve 

fit is affected even less than the quadratic fit when any particular 
standard has a poor value. 

 

• 

2-P (LOGIT/LOG):

 A curve fitted to the standard values, which is 

characterized by a skewed sigmoidal (S-shaped) plot that eventually 
becomes asymptotic to the upper and lower standard values. The 
logistic equation is algebraically transformed to a simpler form in which 
experimentally determined values are used for the responses at 
concentrations of zero and infinity. 

Note:

 Uses linear axis type only. 

• 

Cubic Spline (C-Spline) curve fit:

 A piecewise polynomial 

approximation consisting of joining a set of data points by a series of 
straight lines, which is then smoothed by using a cubic fit. 

• 

4-Parameter Logistic or “4-P”: 

 A curve fitted to the standard values, 

which is characterized by a skewed sigmoidal (S-shaped) plot that 
becomes asymptotic to the upper and lower standard values. The 4 
parameters are: Left asymptote, Right asymptote, Slope and Value at 
the Inflection point. This fit is most recommended for immunoassay 
data, and is more exact than Logit/Log. 

Note:

 Uses linear axis type 

only. 

• 

Point to Point or “PT to PT”:

 A plot that connects each standard point 

with a line, with no averaging of the values to “smooth” the curve at 
each standard. 

• 

Press

 SOFT KEYS 1, 2, 3, 

or

 4

 to select the curve-fit type that is 

displayed above the soft key. Select 

*MORE

 

to display additional 

options. 

• 

Press 

ENTER

 to save the selection and advance to the next screen. 

Summary of Contents for EL800

Page 1: ...Operator s Manual Absorbance Microplate Reader ELx800 ...

Page 2: ... ...

Page 3: ...ement kits are available from BioTek PN 7330513 is required for standard models and PN 7330516 is required for ELx800UV and ELx800NB models The kit contains complete instructions for replacing and aligning the bulb Please ignore the replacement alignment instructions currently supplied in this manual in Chapter 5 If you have any questions or require further assistance please contact BioTek s Techn...

Page 4: ... ...

Page 5: ...ELx800 Absorbance Microplate Reader Operator s Manual October 2006 2006 Part Number 7331000 Revision P BioTek Instruments Inc ...

Page 6: ...rademark and ELx800 Gen5 KC4 KCjunior and EL800 are trademarks of BioTek Instruments Inc Microsoft Windows and the Windows logo are registered trademarks of Microsoft Corporation in the United States and other countries All other trademarks are the property of their respective holders Restrictions and Liabilities Information in this document is subject to change and does not represent a commitment...

Page 7: ... xv Warnings xvi Hazards and Precautions xvi Hazards xvi Precautions xvii Directive 89 336 EEC Electromagnetic Compatibility xix Directive 73 23 EEC Low Voltage xix Directive 2002 96 EC Waste Electrical and Electronic Equipment xix Directive 98 79 EC In Vitro Diagnostics xix Electromagnetic Interference and Susceptibility xx User Safety xx Safety Symbols xxi Chapter 1 Introduction 1 Introducing th...

Page 8: ...ain Menu 18 Configuring Global Default Options 19 Connecting a Printer to the ELx800 20 Setting Up the Serial Port for Communications With Other Devices 22 Attaching the Cable 22 Installing Software on the Host PC 22 Setting Communication Parameters 22 Installing Additional Filters 24 Checking the Reader s Filter Table Setting 26 Repackaging and Shipping the ELx800 28 Before Repackaging the Instru...

Page 9: ...ns 95 Setting the Date and Time 95 Viewing Editing the Filter Table 96 Specifying Data Output and Reporting Options 97 Selecting Read Options 99 Chapter 4 Instrument Qualification 101 Recommendations for Achieving Optimum Performance 102 Recommended Qualification Schedule 103 Qualification Procedures 104 System Test 105 Checksum Test 107 Absorbance Plate Test 108 Description 108 Requirements 110 D...

Page 10: ...es 134 Procedure 134 Chapter 6 Troubleshooting and Error Codes 137 Overview 138 System Test Description 138 Glossary of Terms 138 Error Codes 139 General Errors 140 Fatal Errors 146 Appendix A Computer Control 147 Overview 148 Controlling the Reader With Gen5 149 Setting Up Gen5 149 Problems 150 Getting Started With Gen5 150 Controlling the Reader With KC4 152 Setting Up KC4 152 Problems 153 Getti...

Page 11: ...ndix E Instructions for Programming a New Assay 179 Sample ANA Screen Enzyme Immunoassay Kit 180 Intended Use 180 Background 180 Principle of the Assay 180 Materials 181 Quality Control and Results 182 Expected Values 182 Programming the ANA Screen Enzyme Immunoassay Kit 183 Sample Human Anticardiolipin IgG Enzyme Immunoassay Kit 185 Intended Use 185 Background 185 Principle of the Assay 185 Mater...

Page 12: ...ttaching the End Caps 33 Figure 13 Placing the Unit in the Box 34 Figure 14 Keyboard 38 Figure 15 Options Available from the Main Menu 42 Figure 16 Sample Output for the System Test Sheet 1 of 2 106 Figure 17 Sample Output for the System Test Autocal Analysis Sheet 2 of 2 107 Figure 18 Sample Standards Certificate for the Absorbance Test Plate 111 Figure 19 Sample Test Plate Analysis Report 111 Fi...

Page 13: ...arameters 169 Figure 38 Samples With Calls on Matrix Report 172 Figure 39 Curve Fit Report 173 Figure 40 Samples With Calls on Column Report 174 Figure 41 Column Report Without Samples 175 Figure 42 Panel Report 176 Figure 43 Assay Detail Report Sheet 1 of 2 177 Figure 44 Assay Detail Report Sheet 2 of 2 178 List of Tables Table 1 Recommended Qualification Schedule 103 Table 2 Typical Enzyme Subst...

Page 14: ... free in the U S 802 655 4740 outside the U S Fax 802 655 7941 E Mail customercare biotek com Service TAC Phone 800 242 4685 toll free in the U S 802 655 4740 outside the U S Fax 802 655 3399 E Mail tac biotek com European Coordination Center BioTek Instruments GmbH Kocherwaldstrasse 34 D 74177 Bad Friedrichshall Germany Internet www biotek de Phone 49 0 7136 9680 Fax 49 0 7136 968 111 E Mail info...

Page 15: ...tes Warning A Warning indicates the potential for bodily harm and tells you how to avoid the problem Caution A Caution indicates potential damage to the instrument and tells you how to avoid the problem DEFINE Text in COURIER font represents menu options as they appear on the instrument s display Note Bold text is primarily used for emphasis This icon calls attention to important information ...

Page 16: ... Updated Elx800 and EL800 model comparison table in Appendix F I 2 99 Incorporated Manual Updates Changed European addresses J 9 99 Changed printer compatible description Changed the number of samples necessary if there aren t controls standards or blanks defined Corrected the number of stored plates to 8 Corrected the positions for re use of standard curves Removed the M command from Appendix B C...

Page 17: ...Optional Accessories list Chapter 1 page 1 8 Changed callout in Figure 2 2 from Place unit in bag to Unit in plastic bag Chapter 2 page 2 2 Added KC4 startup information to Appendix E Removed references to ELx800UV and EL800UV from Appendix F Standardized the presentation of significant digits throughout Changed Abs to OD throughout N 10 04 Restructured manual according to new template Removed ref...

Page 18: ...r 4 Changed title from Performance Verification and IQ OQ PQ Procedures to Instrument Qualification Add Gen5 instructions for System Test and Absorbance Plate Test In Recommended Qualification Schedule changed PQ frequency from monthly and semiannually to monthly and quarterly and clarified criteria for running Liquid Tests 1 2 or 3 In Absorbance Plate Test section added a sample test plate report...

Page 19: ...elopment or other nonclinical purposes If there is no such label then the instrument may only be used for research and development or for other nonclinical purposes Quality Control It is considered good laboratory practice to run laboratory samples according to instructions and specific recommendations included in the assay package insert for the test to be conducted Failure to conduct Quality Con...

Page 20: ...nstrument s power supply or power cord must be connected to a power receptacle that provides voltage and current within the specified rating for the system Use of an incompatible power receptacle may produce electrical shock and fire hazards Warning Electrical Grounding Never use a two prong plug adapter to connect primary power to the external power supply Use of a two prong adapter disconnects t...

Page 21: ...uct quality control checks could result in erroneous test data Warning Reader Data Reduction Protocol For readers with onboard assay software the software will flag properly defined controls when they are out of range The software will present the data with the appropriate error flags for the operator to determine control and assay validity For readers operated via computer control no limits are a...

Page 22: ... supply within the range of line voltages listed on it Caution Shipping Hardware Shipping straps and a shipping block must be removed before operating the reader They must be reinstalled before repackaging the reader for shipment See Chapter 2 Installation Caution Disposal This instrument contains printed circuit boards and wiring with lead solder Dispose of the instrument according to Directive 2...

Page 23: ...Part 1 General requirements Directive 2002 96 EC Waste Electrical and Electronic Equipment Disposal Notice This instrument contains printed circuit boards and wiring with lead solder Dispose of the instrument according to Directive 2002 96 EC on waste electrical and electronic equipment WEEE Directive 98 79 EC In Vitro Diagnostics Product registration with competent authorities Traceability to the...

Page 24: ...t his her expense Canadian Department of Communications Class A This digital apparatus does not exceed Class A limits for radio emissions from digital apparatus set out in the Radio Interference Regulations of the Canadian Department of Communications Le present appareil numerique n met pas du bruits radioelectriques depassant les limites applicables aux appareils numerique de la Class A prescrite...

Page 25: ...nu et courant alternatif Gleich und Wechselstrom Corriente continua y corriente alterna Corrente continua e corrente alternata Earth ground terminal Borne de terre Erde Betriebserde Borne de tierra Terra di funzionamento Protective conductor terminal Borne de terre de protection Schutzleiteranschluss Borne de tierra de protección Terra di protezione On Supply Marche alimentation Ein Verbindung mit...

Page 26: ...ntrappolarsi Warning hot surface Attention surface chaude Warnen heiße Oberfläche Precaución superficie caliente Attenzione superficie calda Consult instructions for use Consulter la notice d emploi Gebrauchsanweisung beachten Consultar las instrucciones de uso Consultare le istruzioni per uso In vitro diagnostic medical device Dispositif médical de diagnostic in vitro Medizinisches In Vitro Diagn...

Page 27: ...ion if technical assistance is needed Introducing the ELx800 Absorbance Microplate Reader 2 Hardware Features 3 Software Features 3 Package Contents 4 Optional Accessories 5 Specifications 6 Standard Model 6 Ultraviolet UV Model 7 Narrow Beam NB Model 8 Product Support Service 10 Contacting the Technical Assistance Center 10 Returning Instruments for Service Repair 10 ...

Page 28: ...ge is from 400 nm to 750 nm UV instruments have an extended range from 340 nm to 750 nm Kinetic analysis can be performed using computer control e g via Gen5 KC4 or KCjunior running on a host PC Assay definitions consisting of protocols templates and formulas and the data they produce are managed by an onboard processor via a 2 line x 24 character LCD screen and membrane switch Data can be stored ...

Page 29: ...nds read capability to include 384 well and 60 72 96 well Terasaki microplates External 24 volt power supply compatible with 100 240 V 10 0 50 to 60 Hz One serial COM port 25 pin male connector One parallel port 25 pin female connector Software Features An easy to use menu driven interface Endpoint calculations Curve Fitting with 4 parameter cubic quadratic linear 2 P cubic spline and point to poi...

Page 30: ...ne blank filter The UV model includes a 340 nm filter Operator s Manual PN 7331000 Printer cable PN 71072 Dust cover PN 7332040 RS 232 serial cable PN 75053 Unpacking instructions PN 7331006 packing instructions PN 7331034 Shipping document PN 94075 that includes Warranty Statement and Certificate of Compliance and Calibration Set of shipping materials PN 7332062 Declaration of Conformity PN 73310...

Page 31: ... PN 7330531 for 60 72 and 96 well Terasaki plates for ELx800NB and ELx800RNB Absorbance Test Plate PN 9000547 6 filter or PN 7260522 7 filter Gen5 s Reader Diagnostics Utility PN 5320201 for the Absorbance Test Plate Absorbance Liquid Test Solutions BioTek Wetting Agent Solution PN 7773002 BioTek QC Check Solution No 1 PN 7120779 25 ml or PN 7120782 125 ml Epson LX 300 printer or equivalent 110V o...

Page 32: ...1 0 from 0 000 to 2 000 OD at 405 nm 3 0 from 2 000 OD to 3 000 OD 450 nm Repeatability STD 0 5 0 005 OD from 0 000 to 2 000 OD 405 nm The following specifications apply to 96 well flat or round bottom plates single wavelength measurements with a 30 second read rapid read mode Absorbance Measurement Range 0 000 to 3 000 OD Accuracy 2 0 0 020 OD from 0 000 to 2 000 OD 405 nm Linearity 2 0 from 0 00...

Page 33: ...o 2 000 OD 405 nm Linearity 1 0 from 0 000 to 2 000 OD 405 nm 3 0 from 2 000 to 3 000 OD 450 nm Repeatability STD 0 5 0 005 OD from 0 000 to 2 000 OD 405 nm Optical specifications for the 340 to 400 nm range 50 second read in normal read mode Absorbance Measurement Range 0 000 to 2 000 OD Accuracy 2 0 0 010 OD from 0 000 to 2 000 OD 340 nm Linearity 2 5 from 0 000 to 2 000 OD 340 nm Repeatability ...

Page 34: ... 405 nm 450 nm 490 nm and 630 nm The following specifications apply to 96 well flat or round bottom plates with a 50 second read normal read mode Absorbance Measurement Range 0 000 to 3 000 OD Accuracy 1 0 0 010 OD from 0 000 to 2 000 OD 405 nm Linearity 1 0 from 0 000 to 2 000 OD 405 nm 3 0 from 2 000 to 3 000 OD 450 nm Repeatability STD 0 5 0 005 OD from 0 000 to 2 000 OD 405 nm The following sp...

Page 35: ...bance Measurement Range 0 000 to 3 000 OD Normal Read Mode Dual wavelength 4 minutes 45 seconds Accuracy 2 0 0 020 OD from 0 000 to 2 000 OD 405 nm Linearity 2 5 from 0 000 to 2 000 OD 405 nm Repeatability STD 1 5 0 010 OD from 0 000 to 2 000 OD 405 nm Rapid Read Mode Dual wavelength 3 minutes 25 seconds Accuracy 2 5 0 020 OD from 0 000 to 2 000 OD 405 nm Linearity 2 5 from 0 000 to 2 000 OD 405 n...

Page 36: ...rovide the following information Your name and company information A daytime phone or fax number and or an e mail address The product name model and serial number The software part number and basecode version available via the keyboard by selecting UTIL TESTS CHKSUM For troubleshooting assistance or instruments needing repair the specific steps that produce your problem and any error codes display...

Page 37: ...Power Up and System Test 18 ELx800 Main Menu 18 Configuring Global Default Options 19 Connecting a Printer to the ELx800 20 Setting Up the Serial Port for Communications With Other Devices 22 Attaching the Cable 22 Installing Software on the Host PC 22 Setting Communication Parameters 22 Installing Additional Filters 24 Checking the Reader s Filter Table Setting 26 Repackaging and Shipping the ELx...

Page 38: ...als This packaging should protect the instrument from damage during shipping Inspect the shipping box packaging instrument and accessories for signs of damage If the reader is damaged notify the carrier and your manufacturer s representative Keep the shipping cartons and packing material for the carrier s inspection The manufacturer will arrange for repair or replacement of your instrument immedia...

Page 39: ...nual Unpacking the Instrument and Its Accessories 1 Carefully open the top of the box and remove the power supply shelf Figure 1 2 Remove the manual Declaration of Conformity and other accessories Figure 1 Removing the power supply and shelf Shelf Power supply ...

Page 40: ...of the box and remove the left and right foam end caps Figure 2 Place the reader on a level surface and remove the instrument from the 2 Mil plastic bag Figure 2 Removing the foam end caps Left end cap Unpacking instructions Unit inside 26 x 32 2 Mil poly bag Right end cap ...

Page 41: ...urface 5 Use the slotted screwdriver to remove the four mounting screws from the top cover Figure 3 and unhook the shipping straps from the post on the bottom of the instrument 6 Set aside the four screws you will use them again in step 9 Figure 3 Removing the top cover mounting screws Top cover Shipping straps Post ...

Page 42: ...s and shipping block with the packaging materials 9 Replace the top cover turn the instrument upside down and use the slotted screwdriver to replace the cover s four mounting screws 10 Remove the shipping straps from the top of the carrier and store them with the packaging materials 11 Turn the instrument right side up Figure 4 Removing the shipping block Shipping block Shipping straps Carrier ...

Page 43: ... power supply to ensure proper operation of the unit The ELx800 has a universal 24 VDC 2 1 A power supply that functions from 100 to 240 V 10 0 50 to 60 Hz without external switching Never use a two prong plug adapter to connect primary power to the ELx800 power supply Use of a two prong adapter disconnects the utility ground creating a severe shock hazard Always connect the power supply cord dire...

Page 44: ...SYSTEM TEST PASS The power up system test does not produce a printed results report To run the test manually and obtain a printout of the system test values start at the Main Menu and press UTIL TESTS SYSTEM See System Test and Checksum Test in Chapter 4 or Connecting a Printer to the ELx800 on page 20 for more information ELx800 Main Menu Following successful power up of the ELx800 the Main Menu ...

Page 45: ...a computer or to both Additional options include report format Column and or Matrix and whether or not to print standard curves READ Enable or disable read time prompting for Plate ID Sample ID and Sample Count Specify whether or not to read in Rapid mode Note Refer to Using the Utility Options in Chapter 3 for more specific information on setting up the ELx800 READY 09 14AM 09 08 04 READ DEFINE R...

Page 46: ...l port requires a 25 pin D sub connector A parallel cable PN 71072 designed to connect the reader to a printer is provided with the ELx800 To attach a printer to the ELx800 1 Turn the reader off 2 Place the printer in a location adjacent to the ELx800 3 Attach one end of the cable to the parallel port on the printer 4 Attach the other end of the cable to the parallel port on the ELx800 5 Tighten t...

Page 47: ...Setting Up the ELx800 21 ELx800 Operator s Manual Figure 6 Connectors for printer parallel computer serial and power supply Serial port Parallel port printer port Power supply jack ...

Page 48: ... software Attaching the Cable 1 Power down the computer and the ELx800 2 Connect the appropriate serial cable to both machines The serial port on the reader is a DTE configuration with a 25 pin pin male D sub connector 3 Power up the reader and the computer 4 Ensure that the ELx800 and the computer are operating with the same communications settings Installing Software on the Host PC Refer to the ...

Page 49: ... I O N D A T E T I M E F I L T E R M O R E E D I T S E T U P I N F O R M A T I O N R S 2 3 2 C A L P L A T E M O R E 4 The SELECT BAUD RATE screen will appear showing the currently defined Baud Rate S E L E C T B A U D R A T E 9 6 0 0 1 2 0 0 2 4 0 0 9 6 0 0 V I E W Select the desired baud rate Select VIEW if you wish to see the reader s other communication settings R S 2 3 2 S E T T I N G N O P A...

Page 50: ...50 490 630 and 340 nm Important Keep track of all filter locations The physical location of the filters must match the filter locations mapped in the reader s software filter table The filter wheel must have no empty locations all locations must be filled with either a filter or a blank plug Install all filters with the arrow denoting the light direction pointing downward Figure 7 Filter wheel ass...

Page 51: ...e cover The filter wheel sits on a pin and can be lifted off 8 Remove the filters by turning the wheel upside down over a cloth The filters which are labeled with the wavelength and light direction arrow should easily slide out 9 When replacing the filters in the wheel handle them from the edges Do not touch the glass portion of the filter Clean any filters that appear dirty with lens paper and is...

Page 52: ... I N E R E P O R T U T I L S E L E C T U T I L I T Y O P T I O N T E S T S S E T U P O U T P U T R E A D 2 From the SELECT UTILITY OPTION menu press SETUP The Edit Setup Information screen appears on the display E D I T S E T U P I N F O R M A T I O N D A T E T I M E F I L T E R M O R E 3 From this menu press FILTER 4 The wavelength for Filter 1 will be displayed To advance to Filter 2 press ENTER...

Page 53: ...L T E R 1 W A V E L E N G T H 4 0 5 When the last filter has been entered the software exits the filter routine and displays the following screen E D I T S E T U P I N F O R M A T I O N D A T E T I M E F I L T E R M O R E 6 Press the Main Menu key to return to the main menu ...

Page 54: ...id the warranty If the original packing materials have been damaged or lost contact BioTek for replacement packing see Product Support Service in Chapter 1 for contact information Warning If the reader has been exposed to potentially hazardous material decontaminate it to minimize the risk to all who come in contact with the reader during shipping handling and servicing Decontamination prior to sh...

Page 55: ...800 and Its Accessories Refer to Figures 8 through 13 when repackaging the ELx800 1 Move the carrier to the home position 2 Turn off the instrument and unplug the power supply 3 Wrap the shipping straps around the carrier Figure 8 Figure 8 Securing the carrier Carrier Shipping straps ...

Page 56: ...the instrument over and hook the shipping straps over the post Figure 9 5 Using a slotted screwdriver remove the four top cover mounting screws 6 Lift the instrument off the cover and turn it over Figure 9 Hooking the shipping straps over the post Post Mounting screws Top cover Shipping straps ...

Page 57: ...Lx800 Operator s Manual 7 Install the shipping block place the groove in the shipping block over the shaft and use a Phillips screwdriver to attach the two mounting screws Figure 10 Figure 10 Installing the shipping block Shipping block Shaft ...

Page 58: ...32 Chapter 2 Installation BioTek Instruments Inc 8 Turn the instrument over and install the top cover that was removed in step 6 Figure 11 Figure 11 Reinstalling the top cover Top cover Mounting screws ...

Page 59: ...Turn the instrument right side up place it in the 2 Mil plastic bag then attach the left and right end caps Figure 12 10 Place the instrument with the end caps into the box Figure 13 Figure 12 Reattaching the end caps Left end cap 26 x 32 2 Mil poly bag Right end cap ...

Page 60: ...oTek Instruments Inc 11 Place the shelf into the notched out area in the end cap and the power supply into an 8 x 11 bubble bag Place the power supply on the shelf Figure 13 Figure 13 Placing the unit in the box Shelf Power supply ...

Page 61: ...her the reader requires calibration cleaning periodic maintenance warranty work and or repair Make a note of any error messages displayed and their frequency 3 Provide BioTek with the name and contact information of a person who may be contacted if questions arise 4 Close the box and tape it shut 5 Write RMA and the RMA number in large clear letters on the outside of the shipping container and shi...

Page 62: ...36 Chapter 2 Installation BioTek Instruments Inc ...

Page 63: ...42 Define 44 Define Method Map Formula and Curve 47 Defining METHOD 47 Defining MAP 51 Defining FORMULA 67 Defining CURVE 79 Reading a Microplate 86 Selecting an Assay to Run 87 Printing Reports 91 Editing Standard Outliers 92 Printing Results 94 Using the Utility Options 95 Setting the Date and Time 95 Viewing Editing the Filter Table 96 Specifying Data Output and Reporting Options 97 Selecting R...

Page 64: ...nc ELx800 Front Panel Main Menu Previous Screen Options Main Menu Previous Screen Options 7 8 9 1 2 3 0 G H A B C 4 5 6 D E F Shift CLEAR ENTER STOP READ R E A D Y 9 4 5 A M 0 9 1 2 0 4 R E A D D E F I N E R E P O R T U T I L Figure 14 Keyboard ...

Page 65: ...ally saves the current screen settings and advances to the next screen in a series Pressing Previous Screen generally saves the current screen settings and returns control to the screen most previously viewed Press CLEAR to reset a numeric value to 0 or to clear all characters when editing an assay name Tip Press Shift Clear at the Map Generation screen to clear a previously defined manual map Mov...

Page 66: ...ttoms can cause loss of accuracy especially with U and V bottom polyvinyl microplates Check for this by reading an empty microplate Dual wavelength readings can eliminate this problem or bring the variation in density readings to within acceptable limits for most measurements Inaccuracy in pipetting has a large effect on measurements especially if smaller volumes of liquid are used For best result...

Page 67: ... 4 for more information Refer to Chapter 6 Troubleshooting and Error Codes to interpret error codes and Chapter 1 Introduction for information on contacting BioTek Instruments Technical Assistance Center TAC P o w e r u p S e q u e n c e V x x x I n i t i a l i z i n g B i o T e k I n s t r u m e n t s S y s t e m S e l f T e s t IMPORTANT Some ELx800 readers have custom programmed software instal...

Page 68: ...llows the creation of a reading and data reduction protocol You will be prompted to select an assay from an assay definition list REPORT option Allows printing of results and protocol descriptions You will be prompted for the name of a previously run assay with valid data UTIL option You will be prompted to select options from the list of onboard utilities READY 09 14AM 09 08 04 READ DEFINE REPORT...

Page 69: ...tion are unavailable for programming within Quick Read You can quickly access the Quick Read assay by selecting READ from the main menu The Quick Read assay default DEFINE settings are shown below and cannot be edited except where noted Method Single Wavelength 405 nm editable Map 96 well plate geometry Blank on Air Automap Map starting location A1 Samples only no blanks standards or controls Samp...

Page 70: ... E 3 Optional Edit the assay name and then press ENTER See Editing the Assay Name on page 46 for detailed instructions The DEFINE menu will appear D E F I N E M E T H O D M A P F O R M U L A C U R V E The following options are available within the DEFINE menu METHOD Define the wavelength type single or dual wavelength s and plate geometry page 47 MAP Specify the plate layout using blanks controls ...

Page 71: ... the assay 1 From the Main Menu press DEFINE MAP and continue to press ENTER until ENTER NUMBER OF SAMPLES is displayed Enter at least one sample The software will lock up if zero 00 samples are selected E N T E R N U M B E R O F S A M P L E S 0 1 2 From the Main Menu press UTIL READ and continue to press ENTER until PROMPT SAMPLE COUNT is displayed Select YES P R O M P T S A M P L E C O U N T Y E...

Page 72: ... below CLEAR Clears the reader s display MAIN MENU Returns the display to the Main Menu screen PREVIOUS SCREEN Returns the display to the previous screen ENTER Saves the current settings and advances to the next screen Editing the Assay Name Use the EDIT NAME screen to edit the name currently assigned to the assay The assay name can contain up to 16 alphanumeric characters N A M E H B S A G 1 S P ...

Page 73: ...e following functions METHOD Specify the wavelength type wavelength filter s and plate geometry MAP Specify mapping information FORMULA Access the formula entry screens CURVE FIT Specify curve fit options Defining METHOD Defining a method includes selecting Single or dual wavelength Filter s Plate geometry The options appear on the display in the order that they were programmed in the assay If the...

Page 74: ...n the two measurements The final reported optical density is the difference between the two readings the delta OD Dual wavelength readings can significantly reduce optical interference caused by scratched or fingerprinted microplates since the scratches or fingerprints reduce the amount of light on both wavelengths The currently selected wavelength appears on the top line of the display and the av...

Page 75: ...ll Costar 96 well Costar 384 well Nunc square 96 T Terasaki 72 T Terasaki 60 T Terasaki Used only on ELx800NB version The PLATE TYPE selection screen allows you to select the geometry of the plate that will be used for the assay Press MORE to cycle through the available options Note The NB version of the reader includes choices for 384 well or 72 and 96 well Terasaki plate formats An adapter PN 73...

Page 76: ... 4 8 9 6 9 6 H M O R E P L A T E T Y P E 9 6 W E L L S 9 6 M M O R E Press MORE to cycle through the available options 6 6 well 2 x 3 96 96 well 8 x 12 12 12 well 3 x 4 96H 96 well Hellma Quartz 8 x 12 24 24 well 4 x 6 96M 96 well Metric 8 x 12 9 mm well 48 48 well 6 x 8 spacing ...

Page 77: ...er of Standard Replicates Averaging of Standards Concentration and Location of Standards Number of Controls Control Type Definition Number of Control Replicates Control Location Number of Samples Number of Sample Replicates Sample Location Important The NB reader using 384 well geometry does not have access to map formula or curve options The 384 well plate type is preset for the map to have 384 s...

Page 78: ...ch blanks controls standards and or samples are assigned to specific locations on the plate M A P G E N E R A T I O N M A N U A L A U T O M A N U A L Select AUTOMATIC PLATE MAP GENERATION to instruct the software to automatically generate a plate map after the blanks controls standards and or samples have been defined Select MANUAL PLATE MAP GENERATION to indicate that the well assignments will be...

Page 79: ...ER to save the selection and continue Replication Direction This option allows you to specify how replicates are mapped on the plate The currently selected Replication Direction appears on the top line of the display and the available options appear on the bottom R E P D I R E C T I O N A C R O S S D O W N A C R O S S Select DOWN to map the replicates down the column following the direction of the...

Page 80: ...Rep Direction ACROSS 1 2 3 4 5 6 7 8 9 10 11 12 A STD1 STD1 STD2 STD2 STD3 STD3 STD4 STD4 STD5 STD5 PC PC B NC NC SMP SMP SMP SMP SMP SMP C D E F G H Map Direction DOWN Rep Direction ACROSS 1 2 3 4 5 6 7 8 9 10 11 12 A STD1 STD1 B STD2 STD2 C STD3 STD3 D STD4 STD4 E STD5 STD5 F PC PC G NC NC H SMP SMP Map Direction ACROSS Rep Direction DOWN 1 2 3 4 5 6 7 8 9 10 11 12 A STD1 STD2 STD3 STD4 STD5 PC ...

Page 81: ...ping S T A R T M A P P I N G A T W E L L L O C A T I O N A 0 1 Use the numeric and alpha keys to enter a letter or number at the cursor location For any well location only the alpha keys are active for the first character and numeric for the second and third characters The valid entry range is from A01 to the last well on the plate depending on the plate type and the number of blanks standards con...

Page 82: ...and P DOWN are displayed on three screens B L A N K M A P F U L L A I R F U L L C O N S T M O R E B L A N K M A P F U L L R O W C O L U M N M O R E B L A N K M A P F U L L P A C R O S S P D O W N M O R E Select the BLANK MAP type see the descriptions on the next page Press MORE to cycle through the available options ROW or COLUMN and P ACROSS or P DOWN Press ENTER to save the well location and con...

Page 83: ...ach well in the row Use manual mapping to position blanks controls standards and samples COLUMN enables a single blank well or an average of blank wells to be placed in each column Since the maximum number of blanks is 12 and if all 12 columns are used each column can have only one blank Manual mapping is recommended in this case Replicates follow in the same column as the first well of each sampl...

Page 84: ...st editable field and advances automatically Press CLEAR to clear the value on the display Press ENTER to continue Number of Blanks The NUMBER OF BLANKS field allows you to enter the number of blank wells in the assay This entry screen is only displayed when Full Column or Row blank maps is selected Any previously defined value is displayed E N T E R N U M B E R O F B L A N K S 0 2 Use the numeric...

Page 85: ...table field Press ENTER to continue Number of Standards This option allows you to enter the number of standard groups that will be used in the assay Any previously defined value will be displayed on the screen Note If the number of standards is altered the number of replicates for the standard automatically reverts to 1 E N T E R N U M B E R O F S T A N D A R D S 0 2 Use the numeric keys to enter ...

Page 86: ... wells on the plate Press CLEAR to clear the value on the display Press ENTER to continue Average Standards The AVERAGE STANDARDS option allows you to select whether or not to average the replicates of each standard group This average is used to calculate the standard curve instead of using the individual replicates of each standard If the number of standard replicates is 1 this option is not avai...

Page 87: ...lable for viewing only Pressing ENTER advances to the concentration value entry for the next standard If manual mapping is selected the location must be defined Pressing ENTER from the standard concentration entry moves the cursor to the LOCATION field Pressing ENTER from the LOCATION field advances to the concentration value entry for the next standard Valid Well Locations When defining the repli...

Page 88: ...y 53 cannot be applied to samples on a plate to be run in Assay 51 To reuse a standard curve you must first program an assay in positions 31 through 55 and then run the assay During the defining process you will be prompted to enter the number of standards the number of standard replicates and the standard concentrations The following screen will appear after these prompts R E U S E S T A N D A R ...

Page 89: ...x was chosen as the starting well in the map usually well A01 If manual mapping was used to map the plate the map is not regenerated the reader will not produce results for the well positions that originally were standards Auto mapping is recommended if the standard s curves will be routinely reused Number of Controls The NUMBER OF CONTROLS screen allows you to enter the number of control groups t...

Page 90: ...CTL2 CTL3 CTL4 After choosing an identifier for CONTROL 1 press ENTER to choose the identifier for the next control Number of Control Replicates The NUMBER OF CONTROL REPLICATES entry screen is displayed if the number of control groups is greater than 0 E N T E R N U M B E R O F R E P L I C A T E S O F P C 0 2 The well ID associated with CONTROL 1 appears first Press ENTER to advance to the next c...

Page 91: ...NTER to advance to the next replicate or sample group Number of Samples The number of sample groups on the plate can be defined here and or it can be defined at run time if UTIL READ PROMPT FOR SAMPLE COUNT is set to YES See Selecting Read Options on page 99 for more information E N T E R N U M B E R O F S A M P L E S 2 4 Use the numeric keys to enter the number of sample groups on the plate The r...

Page 92: ...o 12 replicates The software automatically performs a check to ensure that the number of replicates multiplied by the number of samples does not exceed the number of undefined wells remaining on the plate Sample Location If MANUAL MAP GENERATION is selected and samples are defined the locations for each sample replicate must be specified S A M P L E 1 L O C A T I O N R E P 1 C 0 2 Use the numeric ...

Page 93: ...elect FORMULA The SELECT FORMULA TYPE screen will appear described on the next page Calculation Structure During data reduction formulas are processed in the order shown below The number of permitted formulas of each type is shown as well Blank Validation 0 1 Control Validation 0 4 Assay Validation 0 4 Transformations 0 1 Cutoff Formulas 0 1 Curve Fit Analysis if a curve fit method is defined To c...

Page 94: ... to classify results During data reduction results are evaluated against the cutoff formulas and each well is assigned a user specified label POS NEG or EQUIV TRANSformation formulas are applied to the raw data in preparation for further data reduction and or curve fit calculation VALidation formulas can be used to determine whether or not blanks and or controls are valid In addition Assay Validat...

Page 95: ...UNCTION options continue to press the appropriate SOFT KEY For example press the MATH SOFT KEY several times to see etc When the desired option appears press the right arrow key to select it and advance to the next editable field Press the left arrow key to move the cursor to the left Press CLEAR to delete the item above the cursor When a formula is complete press ENTER to continue Select MATH to ...

Page 96: ...OR MAP The available MAP options depend on the formula type and the current plate map MAP options resemble BLK x mean of the blank wells NC 1 the first NC well or OD every well FUNCTION The following functions can be used in formulas LOG10 Log Base 10 ALOG Anti Log ALOG10 Anti Log Base 10 LOG Log AB Absolute Value SQRT Square Root PWR Power EXAMPLES LOG10 Log Base 10 Log10 2 0 301029995 ALOG Anti ...

Page 97: ...ut it applies to the control replicates or control mean If the criteria are not met results are considered suspect and the message RESULTS INVALID BLANK OR CONTROL VALIDATION FAILED appears on results reports One blank validation formula can be defined Up to four control validation formulas can be defined Define the plate map via DEFINE MAP before creating blank or control validation formulas Blan...

Page 98: ...reen or use the Previous Screen key to move backward through the menu structure Assay Validation Formulas Assay Validation formula s establish a set of criteria used to determine whether or not an assay can be considered valid If the criteria are not met results are considered suspect and the message RESULTS INVALID Assay validation failed appears on results reports Up to 4 assay validation formul...

Page 99: ...nsformation formula may be defined per assay A transformation formula can be simple ex OD 2 100 to multiply all wells on the plate by 100 or more complex with the inclusion of a predefined Transformation Variable see TVAR below Simple Transformation Formulas Simple transformation formulas are typically applied to all wells on the plate For example To divide the OD in each well on the plate by 2 an...

Page 100: ...rmula definition screen will appear so that you can define a formula for use within the transformation formula Use the formula keys Math Other Map and Function to define the Transformation Variable TVAR Once the variable has been defined it can be used in a transformation formula The TVAR will be available as a MAP option when building the transformation formula Example The assay plate map has 2 b...

Page 101: ...ula definition screen is displayed Choose TRANS Enter the formula OD TVAR TVAR 100 using the MATH OTHER MAP and FUNCTION keys TVAR is available as MAP option Example In the case of competitive reactions converting absorbance data to percent B B0 can be OD STD1 100 This divides all the wells by STD1 presumably the 0 standard and multiplies the results by 100 To do this At the SCOPE VARIABLE selecti...

Page 102: ...erion for each of the PC replicates or PC x to indicate the average of the Positive Control replicates or a formula combining the two NC x 0 050 A greyzone around the cutoff value can be defined to indicate equivocal or indeterminate results Do not use the or mathematical symbols in a cutoff formula Tip Choose to print a Column Report to see the greyzone and cutoff values as well as the equations ...

Page 103: ...represents the sample wells EQUIV SMP CUTOFF CUTOFF GREYZONE and SMP CUTOFF CUTOFF GREYZONE POS SMP CUTOFF CUTOFF GREYZONE NEG SMP CUTOFF CUTOFF GREYZONE Examples 1 The cutoff between negative and positive calls should be calculated as the average of the negative controls plus the OD value of 0 500 Samples greater than the cutoff should be labeled as positive No greyzone is required For this examp...

Page 104: ...hould be labeled as negative All other samples should be considered equivocal For this example STD2 x the mean of the STD2 wells equals 2 000 OD The cutoff formula is simply STD2 x The greyzone is 10 POS is selected for SAMP CUTOFF 10 Calls are assigned to sample wells as follows EQUIV if the sample is greater than or equal to 1 800 and less than or equal to 2 200 POS if the sample is greater than...

Page 105: ... definable curve fitting parameters include Curve Fit Type Editing of Outliers Axis Identification Extrapolation of Unknowns Note These screens are displayed on the ELx800 in the order in which they appear in the assay If a closed variable i e an element of the assay definition that you cannot access or modify is being used in the assay the entry screen is omitted Curve Fit Type The ELx800 support...

Page 106: ... to a simpler form in which experimentally determined values are used for the responses at concentrations of zero and infinity Note Uses linear axis type only Cubic Spline C Spline curve fit A piecewise polynomial approximation consisting of joining a set of data points by a series of straight lines which is then smoothed by using a cubic fit 4 Parameter Logistic or 4 P A curve fitted to the stand...

Page 107: ...I E R S M A N U A L N O N E M A N U A L Select NONE to suppress the EDIT STANDARD OUTLIERS capability for this assay Choose MANUAL to enable the capability If AVERAGE STANDARDS is set to NO the individual standard replicates are available for editing If set to YES the standard groups are available for editing After the assay is run and reports are generated press REPORT from the Main Menu Press RE...

Page 108: ...e the curve to evaluate samples outside of the absorbance range defined by the standards E X T R A P O L A T E U N K N O W N S Y E S Y E S N O Select YES to enable extrapolation otherwise select NO On the printed reports extrapolated concentrations RSLT values are surrounded by e g 44 425 If extrapolation is chosen for the Point to Point curve fit unknown concentrations will be extrapolated linear...

Page 109: ...st all read at the same wavelength s Any curve fit type formulas or standard concentrations previously defined for each assay will be used when the assay is selected for a Panel Panel assays cannot reuse standard curves Only an Auto map is recognized by a Panel assay Custom locations entered using a Manual map will be overwritten by the Panel assay If the Panel runs in a 1 12 ACROSS configuration ...

Page 110: ...nclude in the panel 1 to 8 Press ENTER to continue The MAPPING DIRECTION selection screen will appear M A P P I N G D I R E C T I O N D O W N D O W N A C R O S S This option ensures that all assays will be mapped in the same direction Select DOWN or ACROSS After selecting the mapping direction of the assays choose which assays to include in the panel S E L E C T A S S A Y N U M B E R 2 2 N A M E H...

Page 111: ... total of 12 wells defined for controls blanks and standards In the Panel the mapping for Assay 1 begins in well A01 The user wants to run 6 samples in Assay 1 Assay 1 now fills wells A01 through B03 The mapping for Assay 8 can begin in well B04 or any well other than A01 to B03 The reader will chirp if you try to map into a well that is already assigned for use with the Panel The mapping for Assa...

Page 112: ...READ The SELECT ASSAY NUMBER screen will appear 2 Select an assay and then press ENTER See Selecting an Assay to Run on the next page for detailed instructions 3 If required enter the number of Samples Plate ID and or Sample ID Note The options to present these screens are configurable by selecting UTIL READ See Selecting Read Options on page 99 for more information E N T E R N U M B E R O F S A M...

Page 113: ...pts may be presented depending on preferences selected in UTIL READ whether or not the assay specifies manual mapping or if the assay was created or downloaded from BioTek s Extensions Define Reader Protocol software Prompts enabled via UTIL READ can include ENTER NUMBER OF SAMPLES PLATE ID and ENTER SAMPLE ID If the assay specifies manual mapping prompts for information will include the locations...

Page 114: ...the maximum number of wells remaining on the plate after any blank control or standard wells are mapped If you enter a value greater than the number of empty wells remaining on the plate the reader will chirp and automatically change the value to the maximum permissible number of samples Enter Plate ID If the PLATE ID prompt is presented enter a unique plate identifier to be stored in memory with ...

Page 115: ...9 The software will automatically increment each subsequent sample identification number by 1 The sample IDs will be assigned according to the plate map defined in the assay Prompts for Well Location If the assay specifies manual plate mapping and if PROMPT FOR SAMPLE COUNT is set to YES under UTIL READ sample well locations can be defined at run time S A M P L E 1 L O C A T I O N R E P 1 G 0 1 Th...

Page 116: ...R I E R A N D P R E S S R E A D K E Y Before reading the plate ensure that the printer is connected turned on and full of paper Place the plate in the carrier and press the READ key to initiate the plate read After the read is complete data reduction will be performed Calculating Results and then the reports will print Generating Reports To halt the read in progress press the STOP key ...

Page 117: ... L P R I N T R E P O R T R E S U L T M A P A S S A Y L I S T Select RESULT to print an exact copy of results from the plate reading the 8 most recent sets of plate data are stored in memory The form in which the results are presented is determined by the report settings Matrix Column Curve Fit specified under UTIL OUTPUT Select MAP to print a matrix showing the locations of the Blanks Standards Co...

Page 118: ...mory Press ENTER to select a plate and continue If a standard curve was generated and EDIT STANDARD OUTLIERS was set to manual in the assay definition the EDIT STD OUTLIERS prompt is presented otherwise the PRINT RESULTS prompt is presented Editing Standard Outliers If a standard curve was generated and if EDIT STANDARD OUTLIERS was set to MANUAL in the assay definition the option to edit outliers...

Page 119: ... more standard groups can be chosen for exclusion E D I T S T D 1 X Y E S Y E S N O Select YES to exclude the group from curve fit calculations Select NO to retain the group Press ENTER to advance to the next group Each curve fit type requires a minimum number of standards for curve generation 4 for 2 P 4 P cubic and cubic spline 3 for quadratic and 2 for linear and point to point Exercise caution...

Page 120: ...plate map S E L E C T A S S A Y N U M B E R 0 1 N A M E H B S A G Press Options to cycle through the list of available assays or enter the number of the desired assay Press ENTER to print the report Assay Report The Assay Report lists the assay definition parameters and their current settings S E L E C T A S S A Y N U M B E R 0 1 N A M E H B S A G Press Options to cycle through the list of availab...

Page 121: ... on the keypad The cursor is positioned under the first editable field and advances automatically To change the date format press the soft key beneath MMDDYY or DDMMYY The display automatically updates to reflect the new format Press ENTER to return to the EDIT SETUP INFORMATION menu 3 To change the current time and or the time format select TIME from the EDIT SETUP INFORMATION menu The TIME entry...

Page 122: ...ill appear E D I T S E T U P I N F O R M A T I O N D A T E T I M E F I L T E R M O R E 2 Select FILTER to display the wavelength for Filter 1 E N T E R F I L T E R 1 W A V E L E N G T H 4 0 5 3 Enter the correct wavelength for each of the five positions using the numeric keys The range is from 400 to 750 nm 340 to 750 nm for the ELx800NB 4 Continue to scroll through the five available filter posit...

Page 123: ...P O R T O U T P U T B O T H P R I N T C O M P U T E R B O T H 2 The current output option is displayed on the top line Select PRINT to send reports directly to a printer COMPUTER to send data out through the serial port or BOTH Note These options have no effect on data output if the instrument is being controlled by software such as Gen5 KC4 or KCjunior running on a host PC 3 Press ENTER to contin...

Page 124: ...atrix or BOTH See Appendix C for examples of Reports 7 Press ENTER to continue The SAMPLES IN COLUMN REPORT screen will appear S A M P L E S O N C O L R P T N O Y E S N O 8 Select YES to print results for all wells on the plate including samples 9 Select NO to limit the results information to blanks controls and standards 10 Press ENTER to continue The PRINT CURVE FIT screen will appear P R I N T ...

Page 125: ...N O P R O M P T S A M P L E C O U N T Y E S Y E S N O R E A D I N R A P I D M O D E Y E S Y E S N O If selected at read time PLATE ID prompts for microplate identification SAMPLE ID prompts for identification for each sample group SAMPLE COUNT prompts for the number of samples on the plate RAPID MODE prompts for the plate to be read in Rapid Mode or Normal Mode RAPID MODE reads a 96 well plate in ...

Page 126: ...100 Chapter 3 Operation BioTek Instruments Inc ...

Page 127: ...nded Qualification Schedule arranges tasks into Installation Operational and Performance Qualification categories Recommendations for Achieving Optimum Performance 102 Recommended Qualification Schedule 103 Qualification Procedures 104 System Test 105 Checksum Test 107 Absorbance Plate Test 108 Liquid Testing 114 Liquid Test 1 117 Liquid Test 2 119 Liquid Test 3 For UV Models Only 123 ...

Page 128: ...r most measurements Inaccuracy in pipetting has a large effect on measurements especially if smaller volumes of liquid are used For best results use at least 100 μl per well in a 96 well plate and 25 μl in a 384 well plate Dispensing solution into 384 well plates often traps air bubbles in the wells which may result in inaccurate readings A dual wavelength reading method usually eliminates these i...

Page 129: ... than shown below Table 1 Recommended Qualification Schedule Installation Qualification Operational Qualification Performance Qualification Tests Initially Initially Annually Monthly Quarterly System Self Test p 105 Checksum Test p 107 Absorbance Plate Test p 108 Liquid Test 1 p 117 Liquid Test 2 p 119 Liquid Test 3 p 123 Optional for 340 nm If you have an Absorbance Test Plate run Liquid Test 1 I...

Page 130: ...u reshipped the instrument or if regulatory requirements dictate that Performance Qualification Testing is necessary you should perform the following tests After the initial confirmation of operation you should perform the Absorbance Plate Test monthly and Liquid Testing semiannually The System Test described on the next page confirms that the light levels at all installed filter wavelengths and e...

Page 131: ...tructions 2 From the Main Menu select UTIL The SELECT UTILITY OPTION screen will appear S E L E C T U T I L I T Y O P T I O N T E S T S S E T U P O U T P U T R E A D 3 From the SELECT UTILITY OPTION screen select TESTS The SELECT TEST screen will appear S E L E C T T E S T S Y S T E M C H K S U M C A L P L A T E 4 Select SYSTEM to run the System Test or if you are using PC software Gen5 Select Sys...

Page 132: ..._______________________________________________________________ 11 14AM 04 10 06 SYSTEM SELF TEST Filter 405 Gain 36 57 Air 50286 Dark 1105 Delta 49181 Filter 450 Gain 36 57 Air 48848 Dark 1100 Delta 47738 Filter 490 Gain 8 53 Air 54482 Dark 1123 Delta 53359 Filter 630 Gain 10 24 Air 54596 Dark 1122 Delta 53474 Noise Max 1107 Noise Min 1103 Delta 4 AUTOCAL ANALYSIS Upper Left Corner x 9226 y 10746...

Page 133: ...hey will be displayed 1 Press UTIL at the Main Menu At the SELECT UTILITY OPTION screen press TESTS and then CHKSUM at the SELECT TEST screen to run the Checksum Test The information displayed will resemble the following 7 3 3 0 2 0 2 V E R S I O N 3 0 7 C O D E C H E C K S U M 1 7 A 5 The initial checksum test display will show the onboard base code software part number version number and checksu...

Page 134: ...is performed during the Absorbance Plate Test in readers equipped with monochromators Although the ELx800 does not have a monochromator you must enter values for the peak wavelength test when defining the 7 filter test plate in Gen5 KC4 or KCjunior software The peak wavelength test will not actually be performed on the ELx800 during the Absorbance Plate Test Mechanical Alignment The Test Plate has...

Page 135: ... filter calibration values entered are the same as those on the Test Plate Standards Certificate Ensure that the Test Plate is within its calibration certification period If these remedies are ineffective or if the Test Plate requires recalibration call your BioTek representative or BioTek Technical Support Repeatability This test ensures that the instrument meets its repeatability specification b...

Page 136: ...ignment Results WELLS B2 B12 G1 G11 DATA 0 000 0 000 0 000 0 000 TOLERANCE 0 015 0 015 0 015 0 015 RESULT PASS PASS PASS PASS Wavelength 405 nm Accuracy Results WELLS C1 E2 G3 H6 F5 D4 STANDARD 0 147 0 618 1 133 1 701 2 279 2 945 MIN VALUE 0 124 0 586 1 090 1 647 N A N A MAX VALUE 0 170 0 650 1 176 1 755 N A N A DATA 0 148 0 616 1 129 1 695 2 243 2 850 RESULT PASS PASS PASS PASS N A N A Repeatabil...

Page 137: ...isplay the second EDIT SETUP menu Select CALPLATE to access the CALIBRATION FILTER menu 3 Using the Standards Certificate packaged with the Test Plate select the filter wavelength E D I T S E T U P I N F O R M A T I O N R S 2 3 2 C A L P L A T E M O R E C A L I B R A T I O N F I L T E R 4 0 5 4 0 5 4 5 0 4 9 0 M O R E 4 Press ENTER The WELL LOCATION CALIBRATION VALUE screen appears W A V E L E N G...

Page 138: ...tics Universal Plate Test 2 Click New Data Sheet Click the Help button for guidance Running the Absorbance Plate Test IMPORTANT Before running the Absorbance Plate Test ensure that the reader is not running in Rapid Mode To check this select UTIL READ and then cycle through the prompts until READ IN RAPID MODE is displayed Choose NO for an accurate result To run the Absorbance Plate Test from the ...

Page 139: ...gnostics Run Universal Plate Test Click the Help button for guidance To run the Absorbance Plate Test in KCjunior 1 Select Utilities Diagnostics Universal Plate Test 2 Select the Universal Plate ID Wavelength Click the Help button for guidance Test Failures If any of the test parameters report as FAIL confirm that the standard values on the test plate Standards Certificate match the values on the ...

Page 140: ...s that must have proof of linearity at wavelengths lower than those attainable with the Absorbance Test Plate This test is optional since the ELx800 has good front end linearity throughout the specified wavelength range see page 123 BioTek offers a dye solution PN 7120779 25 ml or 7120782 125 ml that may be used in the stock solution formulation for Liquid Tests 1 and 2 or if you prefer you may us...

Page 141: ... boat 1 liter volumetric flask Preparation of Stock Solution 1 Weigh out 0 092 gram of FD C No 5 yellow dye powder into a weigh boat 2 Rinse the contents into a 1 liter volumetric flask 3 Add 0 5 ml of Tween 20 or 5 ml of BioTek s wetting agent 4 Make up to 1 liter with DI water cap and shake well This should create a solution with an absorbance of about 2 000 when using 200 µl in a flat bottom mi...

Page 142: ...ution No 1 into a 100 ml volumetric flask 2 Make up to 100 ml with DI water cap and shake well This should create a solution with an absorbance of about 2 000 when using 200 µl in a flat bottom microwell The OD value result will be proportional to the volume in the well and the amount of QC Check Solution No 1 used You can use a larger or smaller well volume or add more Check Solution or water to ...

Page 143: ... dilution 2 Pipette 200 μl of the concentrated solution into Column 1 of the plate 3 Pipette 200 µl of the diluted solution into Column 2 of the plate After pipetting the diluted test solution into the microplate and before reading the plate we strongly recommend shaking the plate for four minutes This will allow any air bubbles in the solution to settle and the meniscus to stabilize If a plate sh...

Page 144: ... 5 0 005 OD 8 The standard deviation for each set of readings should be less than the allowed deviation For example Absorbance readings of 1 950 1 948 1 955 1 952 and 1 950 will result in a mean of 1 951 and a standard deviation of 0 0026 The mean 1 951 multiplied by 0 5 1 951 0 005 0 0098 which when added to the 0 005 0 0098 0 005 0 0148 OD which is the allowable deviation Since the standard devi...

Page 145: ...st 1 Preparation of Dilutions 1 Set up a rack containing 10 tubes numbered consecutively 2 If you have not already done so prepare concentrated stock Test Solution A or B Refer to Table 3 when executing steps 3 and 4 Table 3 Test Tube Dilutions Tube Number 1 2 3 4 5 6 7 8 9 10 Volume of Concentrated Solution ml 20 18 16 14 12 10 8 6 4 2 Volume of 0 05 Tween Solution ml 0 2 4 6 8 10 12 14 16 18 Abs...

Page 146: ...ion to settle and the meniscus to stabilize If a plate shaker is not available wait 20 minutes after pipetting the diluted test solution before reading the plate Read Plate 1 Read the microplate prepared above five times using Normal Read Mode dual wavelength at 450 nm with 630 nm as the blank Note Do not discard the plate you will use it for the Alignment Test 2 Print the raw data or export it to...

Page 147: ...d for each concentration The well that shows the most variation for each concentration is selected for data reduction 2 For each mean below 2 000 OD calculate the allowed deviation using the repeatability specification for a 96 well format of 0 5 0 005 OD from 0 000 to 2 000 OD 405 nm 3 The standard deviation for each set of readings should be less than the allowed deviation For example Absorbance...

Page 148: ...n values for the H1 well to the same well in the A12 position The difference in the values for any two corresponding wells should be within the accuracy specification for the instrument For example If the mean of well A1 in the normal position is 1 902 where the specified accuracy is 1 0 0 010 OD then the expected range for the mean of the same well in the H12 position is 1 873 to 1 931 OD 1 902 1...

Page 149: ... Required Materials 340 nm filter installed in the reader New 96 well flat bottom microplates Corning Costar 3590 is recommended Calibrated hand pipette s Beakers and graduated cylinder Precision balance with readability to 0 01 g Buffer solution A or B Solution A 10x Concentrate Phosphate Buffered Saline PBS Required Materials Deionized water Ingredients shown in Table 4 D NADH powder D Nicotinam...

Page 150: ...horoughly This is the 10x Concentrate PBS Solution Solution B Sigma PBS Required Materials Deionized water Tween 20 polyoxyethylene 20 sorbitan monolaurate Sigma PBS tablets P4417 or equivalent D NADH Powder D Nicotinamide Adenine Dinucleotide Reduced Form Sigma bulk catalog number N 8129 or preweighed 10 mg vials Sigma number 340 110 Store the D NADH powder according to the guidelines on its pack...

Page 151: ...nt If using the 10x Concentrate PBS Solution mix one part of the concentrate with nine parts of DI water Use 10 ml of this solution as the diluent 4 Pipette the three solutions into a new 96 well microplate 150 µl of the 100 Test Solution into all wells of columns 1 and 2 150 µl of the 75 Test Solution into all wells of columns 3 and 4 150 µl of the 50 Test Solution into all wells of column 5 and ...

Page 152: ... the standard deviation for well A1 is less than 0 017 the well meets the test criteria Linearity Test 1 For each of the three dye concentrations calculate the mean absorbance for the wells containing that solution mean of wells A1 to H2 A3 to H4 and A5 to H6 2 Perform a regression analysis on the data to determine if there is adequate linearity Example using Microsoft Excel In a spreadsheet enter...

Page 153: ...Decontamination This chapter contains procedures for maintaining and decontaminating the ELx800 Maintenance 128 Routine Cleaning Procedure 128 Filter Storage and Handling 129 Replacing the Bulb 130 Decontamination Procedure 133 ...

Page 154: ...rument spray it with liquid or use a wet cloth Do not allow the cleaning solution to run into the interior of the instrument If this happens contact the BioTek Service Department Do not soak the keypad this will cause damage Moisten a clean cloth with deionized or distilled water and wipe the keypad Dry immediately with a clean dry cloth Do not apply lubricants to the microplate carrier or carrier...

Page 155: ...sed surfaces of the instrument 4 If detergent was used wipe all surfaces with a cloth moistened with water 5 Use a clean dry cloth to dry all wet surfaces Filter Storage and Handling To properly store interference filters for the ELx800 microplate reader during extended periods of non use package the filters in a light tight envelope or container away from high humidity This will ensure the longes...

Page 156: ...the instrument over on its back and remove the screws holding on the top cover see Figure 3 in Chapter 2 3 Turn the instrument over again and lift the top cover off 4 Use a Phillips head screwdriver to remove the optics arm cover 5 Loosen the hex screws that hold the lamp in place Do not remove the screws 6 Slide the bulb out backward and detach it from the connector at the back of the optics arm ...

Page 157: ...he Bulb 131 ELx800 Operator s Manual Optics arm cover Optics arm 1 aperture Lamp retention screws Lamp retention screws Bulb retainer Bulb assembly Optics arm Filter wheel cover Figure 20 Lamp assembly Lead wires ...

Page 158: ... completing the alignment turn the instrument off and on again to return the carrier to the home position Warning The alignment procedure requires you to observe the light path while the bulb is turned on To prevent possible vision impairment avoid looking directly at the bulb while it is on To align bulbs on UV and NB model readers 1 Use the lead wires to position the bulb to avoid damage to the ...

Page 159: ...recommendations of the Centers for Disease Control and Prevention CDC Neither BioTek nor the CDC assumes any liability for the adequacy of these solutions and methods Each laboratory must ensure that decontamination procedures are adequate for the Biohazard s they handle Wear prophylactic gloves when handling contaminated instruments Gloved hands should be considered contaminated at all times keep...

Page 160: ...ion to run into the interior of the instrument If this happens contact the BioTek Service Department Do not soak the keypad this will cause damage Important Turn off and unplug the instrument for all decontamination and cleaning operations 1 Turn off and unplug the instrument 2 Prepare an aqueous solution of 0 5 sodium hypochlorite NaClO or bleach As an alternative 70 isopropyl alcohol may be used...

Page 161: ... water Dry immediately with a clean dry cloth 5 Wipe the plate carrier and all exposed surfaces of the instrument 6 Wait 20 minutes Moisten a cloth with deionized or distilled water and wipe all surfaces of the instrument that have been cleaned with the bleach solution or alcohol 7 Use a clean dry cloth to dry all wet surfaces 8 Discard the used gloves and cloths using a Biohazard trash bag and an...

Page 162: ...136 Chapter 5 Maintenance and Decontamination BioTek Instruments Inc ...

Page 163: ...is appendix describes potential error codes that may be displayed on the ELx800 and suggests possible solutions for these problems Overview 138 System Test Description 138 Glossary of Terms 138 Error Codes 139 General Errors 140 Fatal Errors 146 ...

Page 164: ...runs an internal System Test each time it is powered on The reader will chirp repeatedly if the power on System Test results do not meet the internally coded Failure Mode Effects Analysis FMEA criteria established by BioTek A system test should then be initiated to try to retrieve an error code from the reader Glossary of Terms Air Blank A full light reading through a filter with no plate in the l...

Page 165: ...espond to keypad input See General Errors on the next few pages for more information Motor or Optical Sensor Channel 0 Carrier X Axis 0 Reference 1 Carrier Y Axis 1 Measurement A indicates a more serious error In this case turn off the instrument Upon restarting the instrument you should be able to use the keypad See Fatal Errors on page 146 for more information Contact BioTek s Technical Assistan...

Page 166: ...ses User aborted read from KC4 User aborted from another serial interface 0200 X axis carrier in out motor did not find the opto home sensor A motor was unable to move to its home position as registered by feedback from an optical sensor Probable Causes Defective or broken optical sensor Flex cable is disconnected or damaged X axis movement is limited so that the optical sensor cannot be interrupt...

Page 167: ...r cannot be interrupted 0202 Filter wheel did not find opto home sensor Probable Cause Failed lamp 0300 0301 Light beam not found During the X axis movement the light beam saturation transition max light to no light was never found during autocalibration Probable Causes Loose belt loose motor pulley or defective motor drive causing the carrier to ignore movement instructions Failed lamp Loose filt...

Page 168: ... axis failed to get to the same position when moved a known number of steps from the home position and back Probable Causes The belt has slipped because of incorrect tension a loose motor pulley or a loose belt clamp Defective motor drive 0402 Filter wheel failed positional verify lamp failed Probable Causes Optical trigger flag has moved or is loose Filter wheel is binding against the motor gear ...

Page 169: ...xternal signals may be penetrating the measurement chamber Ensure that the bottom and top shrouds as well as the filter wheel cover are correctly installed A faulty motherboard PN 7330400 7330410 or 7330415 or daughter board PN 7330404 or 7330414 Photodiode or power supply may be too close to the instrument Failure indicates excessive variation in the dark current background noise levels of the me...

Page 170: ...ce power up 0A01 0A05 Read time Air Blank out of range The blank full signal reading taken during the current read has changed significantly from the same reading taken during the power up self check Probable Cause Lamp is unstable Reboot system and or install new lamp Turn unit OFF when not in use to conserve lamp life 0B00 Invalid assay An assay number that is not programmed was selected 0C00 Pr...

Page 171: ...et number in the assay protocol Probable Causes Assay wavelength does not match the filter table Run system test Verify that wavelengths in the filter wheel match settings in the filter table see Chapter 2 0F00 0F05 Filter signal out of range The filter 1 5 has a signal that is out of range Probable Causes This indicates a light level problem with one or more filters A UV filter may have been inst...

Page 172: ...on data missing No Autocal data exists for the read location Probable Cause Autocalibration was not performed following assay definition download 1300 Motor not homed successfully If the motor is not correctly homed the instrument sends an error and exits the function Probable Causes Errors 0200 or 0300 have been ignored Refer to Probable Causes for these errors on pages 140 and 141 Fatal Errors F...

Page 173: ...C4 or KCjunior software This Appendix provides basic instructions for installing and configuring Gen5 KC4 or KCjunior software for control of the reader Overview 148 Controlling the Reader With Gen5 149 Controlling the Reader With KC4 152 Controlling the Reader With KCjunior 154 ...

Page 174: ...ures not available through the front panel may be defined and run This Appendix provides basic instructions for installation and configuration of BioTek s Gen5 KC4 or KCjunior software for control of the ELx800 Instructions are also provided for reading a plate using these software packages For more detailed instructions refer to the Gen5 Getting Started Guide or Help system or to the KC4 or KCjun...

Page 175: ...achines 2 Turn on both machines 3 Install Gen5 on the computer s hard drive and register the software with BioTek If you purchased Gen5 s Reader Diagnostics Utility required for the Absorbance Test Plate install this software on the computer s hard drive 4 Start Gen5 5 Login if prompted The default System Administrator password is admin 6 When the Welcome to Gen5 screen appears select System Menu ...

Page 176: ...pe was selected in step 9 Try a different COM port Check the serial cable connections Ensure that the cable is properly attached to the port defined in step 10 and is not a Null cable If this is suspected add another Null and try again Confirm that the reader has passed its Self Test The reader will not communicate if it fails an internal system test If the test still fails refer to the Troublesho...

Page 177: ...parameters as desired Click the Help button for assistance 9 When complete click OK to return to the Procedure dialog Click Validate if you would like Gen5 to verify the defined parameters If all parameters are valid you will receive confirmation If any parameters are invalid Gen5 will provide information for correcting the problem Refer also to the Troubleshooting section of the Help system 10 Cl...

Page 178: ...computer s hard drive and register the software with BioTek 4 Once installed start KC4 5 Select System Readers 6 Scroll through the list of Available Readers and select the appropriate ELx800 reader model Click the Port button and subsequent Setup button to define the following communications parameters Port Select the COM port used for the RS 232 serial cable connection Transmission Speed 9600 de...

Page 179: ...reports and more To read a plate using KC4 1 Select Data New Plate 2 If prompted to select a protocol select Empty Protocol and click OK If not prompted select Protocol New or use KC4 s Protocol Wizard to step through protocol creation 3 Select Protocol Reading The Reading parameters dialog will appear 4 Select a Reading Type 5 Define the Filters wavelengths at which the plate will be read 6 Selec...

Page 180: ...de or Help system for more detailed instructions 1 Power off the computer and the reader Connect the serial cable PN 75053 between the two machines 2 Power up both machines 3 Install KCjunior on the computer s hard drive and register the software with BioTek 4 Once installed start KCjunior 5 Select Setup then Reader1 or Reader 2 To select the reader and define the communications parameters choose ...

Page 181: ...iefly describe how to read a plate using KCjunior Refer to KCjunior s Help system and User s Guide early and often to learn how to create protocols assign well identifiers read plates print reports and more To read a plate using KCjunior 1 Click Read Plate from KCjunior s main screen The Read Plate Dialog will appear 2 If desired enter a Results ID and a Plate Description and then click Read Plate...

Page 182: ...156 Appendix A Computer Control BioTek Instruments Inc ...

Page 183: ...metry To capture and manipulate the raw data you must use Gen5 KC4 or KCjunior software This Appendix provides instructions for setting up and performing a 384 well plate read for data reduction in Gen5 KC4 and KCjunior Perform a 384 Well Plate Read in Gen5 158 Perform a 384 Well Plate Read in KC4 162 Perform a 384 Well Plate Read in KCjunior 165 ...

Page 184: ...pears select System Menu 3 At Gen5 s main screen System Reader Configuration to open the Reader Configuration dialog Figure 21 Reader Configuration dialog 4 Click the Add button to open the Reader Settings dialog Figure 22 Reader Settings dialog 5 Use the drop down list in Reader Type to select ELx800 6 Enter the appropriate Com Port The Baud Rate is set to the default transmission speed of 9600 f...

Page 185: ... return to the main screen If the test is not successful and you receive an error message refer to the Troubleshooting section of the Help system 8 At the main screen select File New Experiment The New Experiment dialog will appear Figure 23 New Experiment dialog 9 Click Default Protocol then click OK Gen5 will open the Experiment workspace which includes the Protocol menu and Plate screen Figure ...

Page 186: ...Tek Instruments Inc 10 Select Plate Read or click the Read Plate icon The Procedure dialog will appear Figure 25 Procedure dialog 11 Select 384 WELL PLATE for the Plate Type 12 Click Read to open the Read Step dialog Figure 26 Read Step dialog ...

Page 187: ...s 18 Click OK to save and close the Procedure The Plate screen will change to a 384 well plate layout and the Plate Reading dialog will appear Figure 27 Plate Reading dialog 19 Enter any desired information place a 384 well plate on the carrier and then click READ to begin the plate read Click OK when the Load Plate dialog appears The plate will be read To view the raw data results use the Data dr...

Page 188: ... the ELx800NB model to perform these instructions Other ELx800 models do not support 384 well geometry 1 Start KC4 Login if prompted 2 At KC4 s main screen select System Readers Figure 28 KC4 main menu System Readers 3 The Reader Selection dialog will appear Select the ELx800 NB Figure 29 Reader Selection dialog ...

Page 189: ...t Reader to establish communication with the reader then click Close If the test fails follow the instructions provided by KC4 6 At the main screen select Data New Plate The New Data File dialog will appear Figure 30 New Data File dialog 7 Select Empty Protocol and click OK 8 Select Protocol Reading from the main screen to open the Reading dialog Figure 31 Reading dialog ...

Page 190: ...or correcting the parameters 14 Select Data Read Plate The Plate Reading dialog will appear Figure 32 Plate Reading dialog 15 Enter any comments place the 384 well plate on the carrier and then click START READING to begin the plate read The plate will be read and then the raw data results will be displayed in the Results screen for one of the wavelengths tested To view the data for anther wavelen...

Page 191: ...ell geometry 1 Start KCjunior 2 At KCjunior s main screen choose Setup Reader 1 or Reader 2 Figure 33 Setup Reader dialog 3 The Reader Setup dialog will appear Select ELx800NB EL800NB for the Reader Type Figure 34 Reader Setup dialog 4 Define the communication parameters 5 Click Test Communications to establish communication with the reader then click OK If the test fails follow the directions pro...

Page 192: ... the Read Plate button in KCjunior s main screen The Read Plate Dialog will appear Figure 35 Read Plate dialog 7 If desired enter the Results ID and Plate Description then click Read Plate 8 The Protocol Definition dialog will appear Figure 36 Protocol Definition dialog ...

Page 193: ...ould like KCjunior to verify the defined parameters If any parameters are invalid or incomplete follow the instructions provided by KCjunior for correcting the parameters Figure 37 Example Read Method parameters 14 Place the 384 well plate on the plate carrier then click OK to read the plate Click OK when prompted to Place the plate on the carrier The plate will be read and then the raw data resul...

Page 194: ...168 Appendix B Using 384 Well Geometry BioTek Instruments Inc ...

Page 195: ... reports that can be generated and or printed from the ELx800 Refer to Printing Reports in Chapter 3 for details on how to print these reports In addition an Assay List Assay Definition Map and Result can be printed by choosing Report from the Main Menu screen ...

Page 196: ...170 Appendix C Report Format BioTek Instruments Inc Figure 38 Samples with calls on Matrix Report ...

Page 197: ... 171 ELx800 Operator s Manual Figure 39 Curve Fit Report ...

Page 198: ...172 Appendix C Report Format BioTek Instruments Inc Figure 40 Samples with calls on Column Report ...

Page 199: ... 173 ELx800 Operator s Manual Figure 41 Column Report without samples ...

Page 200: ...174 Appendix C Report Format BioTek Instruments Inc Figure 42 Panel Report ...

Page 201: ... 175 ELx800 Operator s Manual Figure 43 Assay Detail Report Sheet 1 of 2 ...

Page 202: ...9 ASSAY 19 20 ASSAY 20 21 ASSAY 21 22 ASSAY 22 23 ASSAY 23 24 ASSAY 24 25 ASSAY 25 26 ASSAY 26 27 ASSAY 27 28 ASSAY 28 29 ASSAY 29 30 ASSAY 30 31 ASSAY 31 32 ASSAY 32 33 ASSAY 33 34 ASSAY 34 35 ASSAY 35 36 ASSAY 36 37 ASSAY 37 38 ASSAY 38 39 ASSAY 39 40 ASSAY 40 41 ASSAY 41 42 ASSAY 42 43 ASSAY 43 44 ASSAY 44 45 ASSAY 45 46 ASSAY 46 47 ASSAY 47 48 ASSAY 48 49 ASSAY 49 50 ASSAY 50 51 ASSAY 51 52 AS...

Page 203: ... x 48 well plates x 96 well plates x x Edit Assay Name x Absorbance Values x x Formulas x Curve fits x Assays available 55 10 Results stored 8 1 Blanks and samples only in Map x Blanks standards controls and blanks in Map x Panel Assay x Computer Control capability x x Serial and Parallel output x x Customizable via Extensions x The EL800 version can be computer controlled and will function as an ...

Page 204: ...178 Appendix D Comparison of the ELx800 and the EL800 BioTek Instruments Inc ...

Page 205: ...tructions are provided so that users can see how it is possible to translate the kit wording into an ELx800 assay program For clarity only the user menu choices from the reader screens are shown Refer to Chapter 3 for details Important The ELx800 user is responsible for programming the reader properly according to their specific kit instructions and for verifying that the calculations are performe...

Page 206: ...ve on this ANA kit should be tested for the specific autoantibodies indicative of the various systemic rheumatic diseases Principle of the Assay Microwells are pre coated with purified antigens The prediluted controls and diluted patient samples are added to the wells and autoantibodies recognizing one or a combination of antigens bind during the first incubation After washing the wells to remove ...

Page 207: ...Control 1 bottle containing 1 8 ml of diluted stabilized human serum Ready to use ANA Conjugate 1 bottle containing 12 ml of peroxidase labeled antibody to human IgG Colored red ready to use TMB Substrate 1 bottle containing 14 ml TMB substrate Ready to use Stop Solution 1 bottle containing 14 ml of 3M Phosphoric acid Ready to use Additional Materials and Equipment Not Supplied Automatic Microplat...

Page 208: ...teria are not met the assay is invalid and the test should be repeated Calculation of the Sample Results Use the following formula to calculate the ANA result for each sample Control or sample OD x 10 Control or sample value U ml Cutoff control OD 9 Expected Values The normal range was determined on serum from 200 normal adult blood donors The cutoff control has been set at a point equivalent to t...

Page 209: ...the plate map press MAP AUTO Auto mapping is normally preferred because it fills in the well IDs logically and automatically after determining which direction to map and how many wells to fill DOWN Maps the wells down the column DOWN Locates replicates in a vertical orientation down the column A01 Begins mapping at well location A01 BLANK MAP AIR Choose to blank on AIR if no blank wells are requir...

Page 210: ...C replicates and PC x to indicate the average of the Positive Control replicates To create the plate transformation for CALC divide the ANA result by a cutoff standard FORMULA MORE TRANS VAR SCOPE VARIABLE SMP or OD Select OD to advance to the formula definition screen and define the transformation variable Tvar FORMULA CTL1 X Defining TRANS VAR CTL1 X isolates the OD value for CTL1 X for use in t...

Page 211: ...yndrome APS Anticardiolipin antibodies have also been associated with a range of clinical conditions including fetal loss endocarditis stroke heart attack and autoimmune haemolytic Principle of the Assay Microwells are precoated with cardiolipin and cofactor Standards controls and patient samples are added to the wells and autoantibodies recognizing cardiolipin bind during the first incubation Aft...

Page 212: ... Cardiolipin IgG Standards 5 bottles each containing 1 2 ml of diluted human serum with the following concentrations of anticardiolipin autoantibody 100 50 25 12 5 6 25 GPL U ml Ready to use The standard set is calibrated against the Louisville APL reference preparation Cardiolipin IgG Positive Control 1 bottle containing 1 2 ml of diluted human serum The expected value is given on the QC certific...

Page 213: ...mple dilution Quality Control and Results Quality Control For an assay to be valid all the following criteria must be met Standards and the positive and negative controls must be included in each run The values obtained for all the controls should be in the ranges specified on the QC Certificate The curve shape should be similar to the standard curve shown on the QC Certificate If the above criter...

Page 214: ... appears to be anomalous then the assay should be repeated Calculation of Autoantibody Levels in Controls and Samples Read the level of the anticardiolipin autoantibody in the controls and diluted samples directly from the calibration curve The control values should fall within the range given on the QC Certificate Note The standard values have been adjusted by a factor of 100 to account for a 1 1...

Page 215: ... normal pregnancy the levels are higher still at 17 0 IgM and 10 6 IgG Expected Values The normal range was determined on serum from 102 normal adult blood donors The ranges below are provided as a guide only ELISA assays are very sensitive and capable of detecting small differences in sample populations It is recommended that each laboratory determine its own normal range based on the population ...

Page 216: ...red At the define menu follow steps below STEP COMMENTS To program the reading method press METHOD WAVELENGTH Dual or Single Single MEASURE Wavelength s to use 450 PLATE TYPE 96 To program the plate map press MAP AUTO DOWN DOWN A01 BLANK MAP AIR NUMBER STDS 05 NUMBER STD REPLICATES 01 CONCENTRATIONS STD1 6 25 STD2 12 5 STD3 25 STD4 50 STD5 100 NUMBER CTLS 02 CONTROL 1 PC CONTROL 2 NC NUMBER OF REP...

Page 217: ...e value or transformed value see the previous example and cannot calculate based on concentration The technician must make the positive or negative determination visually based on the calculated concentration results Curve 4P As a general guideline choose linear if you expect a straight line result Choose 4P for all others unless otherwise specified by the kit instructions The reader will automati...

Page 218: ...192 Appendix E Instructions for Programming a New Assay BioTek Instruments Inc ...

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