Microplate Systems. Absorbance Microplate Readers 366. Performance Validation and Software Tools 368. Microplate Washer 370

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1 /microplate/ Absorbance Microplate Readers 366 Performance Validation and Software Tools 368 Microplate Washer 370 Singleplex Assays and Reagents 371 Microplate Systems

2 Absorbance Microplate Readers /microplate/ Absorbance Microplate Readers Bio-Rad offers absorbance microplate readers, assays, software, accessories, and reagents that support a wide range of applications. Microplate System Selection Guide Model Plate Format Spectral Range Wavelength Selection Software imark microplate nm 6 preinstalled filters, Onboard or optional Microplate Manager 6 absorbance reader 8-position filter wheel software, via USB2, PC, or Mac xmark microplate absorbance 6 1,536; pixel intensity 200 1,000 nm Monochromator Microplate Manager 6 software spectrophotometer maps for 6 96 (included), via USB2, PC, or Mac For more information, request bulletin 5669 (a folder that includes bulletins ). See Also Protein assays: pages imark Microplate Absorbance Reader The imark microplate absorbance reader offers fast reading speeds, extensive onboard software capabilities, a preprogrammed validation protocol (for the Checkmark reader performance verification kit), and a built-in printer in a compact, lightweight unit. The instrument can be used as a stand-alone reader or controlled by a PC or Mac with Microplate Manager 6 software. The wavelength range of the imark reader is nm, permitting many applications such as end-point assays, ELISAs with colorimetric substrates, and protein assays (Bradford and Lowry). Kinetic reading can also be programmed with onboard software or controlled by Microplate Manager 6 software. The imark reader s onboard software is well suited for life science, clinical, and other research laboratories. The software includes security log-in features that allow locking and unlocking of key protocol settings by the administrator, generous memory capacity for protocol creation and storage, comprehensive curve-fit selections, and extensive reporting functions. Onboard software includes preprogrammed protocols for Bio-Rad s Checkmark reader performance verification kit, to validate the IQ/OQ/PQ of the imark reader in compliance with GLP. Other life science and specialized protocols are also included, such as those for the Bio-Rad TSE screening kits and a growing number of food testing and clinical kits. The imark reader comes with six standard absorbance filters: 415, 450, 490, 595, 655, and 750 nm. There are five additional filter options: 405, 540, 550, 570, and 630 nm. Bio-Rad also supplies custom filters at 5 nm intervals from 400 to 750 nm on request. A wide-format internal thermal graphical printer saves counter space and simplifies stand-alone operation. imark reader features include: Flexible configurations with ability to read flat-, U-, or V-bottom microplates or 8- or 12-well strip plates Automatic calibration before each reading Broad wavelength range for varied applications Variable-speed plate-shaking capability Easy access to 8-position filter wheel; 6 standard filters provided USB2 port for external computer control (with Microplate Manager software, PC or Mac) Multilingual interface and LCD display (English, Japanese, Chinese, and Russian) Data and protocol presentation on LCD display Onboard data storage of protocols, standard curves, and graphs Self-diagnostic capabilities to detect lamp burnout at startup Motorized door for plate loading Instant printing of templates, protocols, and results For more information, request bulletin Microplate Systems

3 Absorbance Microplate Readers /microplate/ Specifications Wavelength range nm Photometric range OD Linearity 1.0% from OD, 2.0% from OD Accuracy ±1.0% or from OD at 490 nm Precision 1.0% or OD from OD, 1.5% from OD Resolution OD Filter wheel capacity 8 Preinstalled filters 6 Printing Built-in thermal line-dot printer Plate shaking 3 speeds: low, medium, high; duration: sec Read time 6 sec at single wavelength, 10 sec at dual wavelengths Data output Onboard graphical thermal printer and USB2 interface with PC or Mac data stations Data storage Calendar/clock function, 64 assay protocols Multilanguage support 4 languages, LCD indication supported, printout report supported Dimensions (W x D x H) 34.6 x 37.7 x 16.4 cm (13.6 x 14.8 x 6.5") Weight 5.5 kg (12 lb) imark Microplate Absorbance Reader, 100/240 V, includes 6 filters (415, 450, 490, 595, 655, 750 nm), 5, built-in plate shaker, onboard software and thermal printer, one roll printer paper, USB2 and power cables imark Microplate Absorbance Reader With Microplate Manager 6 Software, for PC and MAC 5, xmark Microplate Absorbance Spectrophotometer With its unique imaging capabilities, monochromator design, and spectral scanning feature (which eliminates the need for filters), the xmark microplate absorbance spectrophotometer can find the best wavelength within a broad range for any photometric application. The xmark spectrophotometer is also flexible enough to read all standard microplates, from 6- to 1,536-well formats. Using Microplate Manager 6 software, the xmark spectrophotometer can perform a wide range of end point and kinetic applications with spectral scanning within low UV to infrared wavelengths. Features include: Monochromator-based system for maximum optical efficiency and flexibility in photometry reading Spectral scanning to define the best absorbance wavelength for assays High-performance optics for low- or high-density formats Imaging capabilities for pixel intensity data in a single well or entire plate Capability of scanning custom plates Flexible control and data analysis with PC or Mac software Complete reproducibility of assays using experiment protocols from Microplate Manager 6 software Built-in incubator with programmable temperature control Built-in plate shaker with variable mixing direction and adjustable speed For more information, request bulletin Microplate Systems 367

4 Performance Validation and Software Tools /microplate/ Specifications Optical Wavelength range 200 1,000 nm Photometric methods Single and dual wavelength Monochromator step size 1 nm Bandwidth 5 nm Wavelength accuracy, minimum ±2.0 nm Wavelength repeatability ±0.2 nm Imaging resolution 0.5, 0.75, 1.0 mm Performance Indication range OD Resolution OD Linearity 2.0%, 0 to 3.0 OD at 405 nm Accuracy ±1.0% or 0.015, OD Reproducibility 96-well plate: 1.0% or 0.005, OD at 405 nm Stability and drift at OD = 1 at 490 nm Read time 8 sec/96-well/single wavelength; 15 sec/384-well/single wavelength; 35 sec/1,536-well/single wavelength Physical Compatible plates 6- to 1,536-well and custom microplates; maximum plate height: 21 mm Incubator set point Temperature range: 25 45ºC; accuracy: ±0.5ºC; uniformity: 0.5ºC Well-to-well uniformity 2.0% or 0.005, 0 to 3.0 OD at 405 nm Plate shaking Linear or orbital plate-shaking capability Computer interface USB2 Dimensions (W x D x H) 54.4 x 44.0 x 21.9 cm (21.4 x 17.3 x 8.6") Weight 27.5 kg (61 lb) xmark Microplate Absorbance Spectrophotometer, PC or Mac, built-in incubator, plate shaker, 18, Microplate Manager 6 software, USB2 and power cables Plate Adaptor, for using Terasaki plate with xmark microplate absorbance spectrophotometer Performance Validation and Software Tools Checkmark Reader Performance Verification and IQ/OQ Kits The Checkmark reader performance verification kit consists of an absorbance calibration plate and Checkmark software. This kit is an IQ/OQ tool that can be used to validate instrument performance parameters of Bio-Rad s microplate absorbance readers. The Checkmark check plate is compatible with all Bio-Rad microplate absorbance readers and is NIST-certified to confirm calibration traceability. Checkmark software allows automated testing of Bio-Rad microplate absorbance readers and generates reports specific to instrument type and serial numbers to ensure GLP compliance. 368 Microplate Systems

5 Performance Validation and Software Tools /microplate/ Performance verification parameters include: Accuracy Precision Linearity Plate positioning (96-well plate format only) Cross talk (best-case and worst-case) Linearity Test 2.5 Filter check 0 OD and overflow (100% and 0% transmittance) Spectral blocking Checkmark software is compatible with Windows XP and Windows Vista operating systems. For more information, request bulletin OD y = x R 2 = Nominal value Filter check using the Checkmark reader performance verification kit Checkmark Reader Performance Verification Kit, includes absorbance calibration plate, Checkmark software 2, IQ/OQ Kit for imark Microplate Absorbance Reader, includes , IQ/OQ protocols 2, IQ/OQ Kit for xmark Microplate Absorbance Spectrophotometer, includes , IQ/OQ protocols 2, Microplate Manager 6 Software Bio-Rad s imark microplate reader and xmark microplate spectrophotometer can be utilized to their full potential with Microplate Manager 6 software. This comprehensive software package allows powerful, versatile colorimetric and turbidimetric analyses and report analysis for raw data, qualitative and quantitative calculations based on absorbance limits, matrix of equations, normalization, and curve fit. Microplate Manager software expands your range of data analysis options to include kinetics and screening, and adds the functionality of flexible template creation for any standard microplate format up to 1,536 wells and for 60- and 70-well Terasaki plates. Based on user-defined templates and parameters, Microplate Manager 6 software reads the plate, generates an array of values, performs data analysis, and creates reports. Key features include: Flexible, intuitive, and efficient software for reading microplates and assay analysis in any research setting Unique access to instrument controls, allowing additional options for optimizing microplate assays Complete reproducibility of the assay workflow, plate reading parameters, calculations, and reports Exact and controllable data with template design flexibility and intuitive guides for designating plate wells, mapping dilutions, and multiple sample series Can deliver comprehensive and configurable reports that contain every step of the workflow and analysis, including descriptive notes Custom reporting function provides one-button screening for predefined assays such as those for TSE screening For more information, request bulletin Microplate Systems 369

6 Microplate Washer /microplate/ Plot of standard curve in Microplate Manager 6 software. Secure and automated reporting for TSE screening. System Requirements PC Macintosh Operating system Windows 2000 or Windows XP Mac OS or Mac OS X 10.5 Processor/model Pentium 333 MHz PowerPC G3 or better RAM 256 MB 256 MB Hard disk space 3 GB 3 GB Monitor 17" or larger 17" or larger USB2 port Required for interface with microplate readers Required for interface with microplate readers Printer Optional Optional Microplate Manager 6 Software, PC and Mac, for end-point and kinetic data analysis with imark microplate 1, absorbance reader and xmark microplate spectrophotometer Microplate Washer Model 1575 Immunowash Microplate Washer The Model 1575 Immunowash microplate washer offers complete control of needle position to perform special wash routines and maximize wash efficiency. This washer includes a wide choice of wash sequences to facilitate protocol creation and storage, and it is compatible with 96-well microplates that have flat-, U-, or V-bottom wells and strips. Other features include: Programmable needle positions (horizontal and vertical) to an accuracy of 0.1 mm for bottom washing, crosswise aspiration, and overflow washing Dispenser speed control Plate-shaking option to help minimize bubbles and adherence of liquid to well sides Waste bottle sensor to detect high waste liquid levels Up to 75 programmable washing sequences Easily removable 8- or 12-way manifolds Easily accessible manifold interior for maintenance Removable and autoclavable plate carrier Aerosol protection cover Integrated vacuum and dispensing pumps to ensure accurate and quiet washing and to eliminate the need for external pumps For more information, request bulletins 2135 and Microplate Systems

7 Singleplex Assays and Reagents /microplate/ Specifications Voltage VAC, 50/60 Hz Residual well volume <2 µl Waste bottle volume 2,000 ml Soak time sec in strip mode 0 59 min in plate mode Available manifolds 8- and 12-way Storage capacity 75 kits Compatible plate types Any flat-, U-, or V-bottom 96-well plate Operating temperature C Dimensions (W x D x H) 32.5 x 40.5 x 19.3 cm (12.8 x 15.9 x 7.6") Weight 9.6 kg (21 lb) Model 1575 Immunowash Microplate Washer, V, includes bottle and tubing set, 6, port manifold, aerosol protection cover Model 1575 Immunowash 12-Channel Manifold Model 1575 Immunowash Standard Maintenance Kit Singleplex Assays and Reagents ELISA Reagents Enzyme-Antibody Conjugates Bio-Rad offers enzyme-antibody conjugates and ELISA soluble substrate systems suitable for use in all microtitration enzyme immunoassays. Antibodies are available conjugated to HRP or AP enzymes. Affinity purification procedures have been used to isolate these specific antibodies and to remove nonspecific antibodies and other serum proteins that can interfere in assay procedures. A mouse isotyping kit is also available. Protein A conjugates (page 247) offer an alternative method to detect immunoglobulins from a variety of species. Substrate Systems Bio-Rad s AP substrate kit contains easy-to-use P-nitrophenyl phosphate (pnpp) tablets and dilution buffer concentrate to prepare 500 ml of working solution. Benefits of the AP system include an extremely linear reaction rate and low background values for accurate quantitation by ELISA. There are two convenient substrate systems for detection of HRP-labeled antibodies. Both the 2,2 -azino-di(3- ethylbenzthiazoline)-6-sulfonic acid (ABTS) peroxidase substrate kit and the 3,3,5,5 -tetramethylbenzidine (TMB) peroxidase substrate kit contain liquid reagents that are mixed just prior to use. No weighing or dissolving of hazardous chemicals is necessary. The single-component TMB peroxidase EIA substrate kit makes microplate assays convenient and fast. See Also Blotting-grade conjugates: page 247. Comparison of ELISA Substrate Systems AP HRP ELISA Type pnpp ABTS TMB Qualitative Stop reagent Sodium hydroxide Oxalic acid Sulfuric acid Color Yellow Blue-green Yellow Detection wavelength 405 nm 415 nm 450 nm Quantitative (Endpoint) Stop reagent Sodium hydroxide Oxalic acid Sulfuric acid Color Yellow Blue-green Yellow Detection wavelength 405 nm 415 nm 450 nm Quantitative (Kinetic) Color Yellow Blue-green Blue Detection wavelength 405 nm 415 nm 655 nm Substrate performance: ++++, excellent; +++, good; ++, fair. Microplate Systems 371

8 Singleplex Assays and Reagents /microplate/ Enzyme-Antibody Conjugates Goat Anti-Mouse IgG (H + L)-HRP Conjugate, 2 ml Goat Anti-Human IgG (g)-hrp Conjugate, 1 ml Goat Anti-Rabbit IgG (H + L)-HRP Conjugate, 1 ml Rabbit Anti-Goat IgG (H + L)-HRP Conjugate, 1 ml Rabbit Anti-Sheep IgG (H + L)-HRP Conjugate, 1 ml Rabbit Anti-Goat IgG (H + L)-AP Conjugate, 1 ml Substrate Systems AP Substrate Kit, contains 100 pnpp tablets and 100 ml 5x concentrate diethanolamine buffer HRP Substrate Kit, contains 180 ml ABTS, 20 ml hydrogen peroxide TMB Peroxidase EIA Substrate Kit, contains 180 ml TMB solution, 20 ml hydrogen peroxide TMB Peroxidase EIA Substrate Kit, contains 900 ml TMB solution, 100 ml hydrogen peroxide Single-Component TMB Peroxidase EIA Substrate Kit, contains 100 ml TMB solution Single-Component TMB Peroxidase ELISA Substrate Kit, contains 1 L TMB solution Mouse Antibody Isotyping Kit The Mouse Typer isotyping kit provides a convenient ELISA method for identifying the class, subclass, and light-chain type of mouse IgGs in tissue culture supernatant and ascites fluid. The kit contains sufficient reagents to perform 100 typings (800 tests). The Mouse Typer kit allows screening of up to 12 different antibody samples per plate. Each sample is reacted with 8 subclass-specific antisera to determine the heavy- and light-chain types of the monoclonal antibody Mouse Typer Isotyping Kit, includes mouse isotyping panel, affinity purified goat anti-rabbit IgG (H + L) Mouse Typer Isotyping Panel, includes 10 ml each ultrapure rabbit anti-mouse subclass-specific antiserum to mouse IgG 1, IgG 2a, IgG 2b, IgG 3, IgM, IgA, k-chain, and l-chain 372 Microplate Systems