Operator’s Manual - Pera Medikal HT Rev_D.pdf · PowerWave Operator’s Manual ix Document...

Operator’s Manual

Microplate Spectrophotometer

PowerWave™

PowerWaveTM Microplate Spectrophotometer

Operator's Manual

MAY 2006

2006

PART NUMBER 7281000

REVISION D

BIOTEK

INSTRUMENTS, INC.

ii Preface

Notices

BIOTEK INSTRUMENTS, INC.

Highland Park, P.O. Box 998 Winooski, Vermont 05404-0998 USA

All Rights Reserved

© 2006, BioTek Instruments, Inc. No part of this publication may be reproduced, transmitted, transcribed, stored in a retrieval system, or translated into any language without the written permission of BioTek Instruments, Inc.

Trademarks

BioTek is a registered trademark, and PowerWave™, PowerWave™ 340, PowerWave™ HT, PowerWave™ HT 340, Gen5™, KC4™, KCjunior™, and Bio-Stack™ are trademarks of BioTek Instruments, Inc. Bio-Cell™ is a trademark of BioTek Instruments and is patented under U.S. patent number 5,963,318. 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 by BioTek Instruments, Inc. Changes made to the information in this document will be incorporated in new editions of the publication. No responsibility is assumed by BioTek for the use or reliability of software or equipment that is not supplied by BioTek, or its affiliated dealers.

PowerWave Operator’s Manual iii

Contents Preface ............................................................................................................ I

Notices ................................................................................................................................ ii

All Rights Reserved...................................................................................................... ii

Trademarks.................................................................................................................. ii

Restrictions and Liabilities............................................................................................ ii

Contents..............................................................................................................................iii

Contacting BioTek Instruments, Inc. .................................................................................viii

Document Conventions...................................................................................................... ix

Revision History .................................................................................................................. x

Intended Use Statement ....................................................................................................xii

Specimen Preparation........................................................................................................xii

Quality Control ...................................................................................................................xii

Repackaging and Shipping ................................................................................................xii

Warnings...........................................................................................................................xiii

Hazards.............................................................................................................................xiii

Precautions .......................................................................................................................xiv

CE Mark Information ......................................................................................................... xv

Electromagnetic Interference and Susceptibility................................................................xvi

User Safety .......................................................................................................................xvi

Safety Symbols ................................................................................................................xvii

Chapter 1: Introduction.............................................................................. 1-1 Introducing the PowerWave .............................................................................................1-2

Variations .........................................................................................................................1-3

Internal Barcode Scanner.................................................................................................1-3

Compatibility With the Bio-Stack ......................................................................................1-4

Hardware Features...........................................................................................................1-5

Software ...........................................................................................................................1-5

Package Contents............................................................................................................1-6

Optional Accessories .......................................................................................................1-6

Specifications ...................................................................................................................1-7

Microplates................................................................................................................1-7

Speed of Reading .....................................................................................................1-7

Optical Specifications................................................................................................1-8

Optical Performance .................................................................................................1-8

Hardware and Environmental Specifications.............................................................1-9

iv Preface

Product Support & Service.............................................................................................1-10

Contacting the Technical Assistance Center...........................................................1-10

Returning Instruments for Service/Repair ...............................................................1-10

Chapter 2: Installation............................................................................... 2-1 Unpack and Inspect the Instrument..................................................................................2-2

Remove the Microplate Carrier Shipping Bracket ............................................................2-4

Select an Appropriate Environment..................................................................................2-5

Connect the Power Supply and Cord ...............................................................................2-5

Turn on the Reader ..........................................................................................................2-7

Connect the Host Computer.............................................................................................2-9

Install Software on the Host Computer...........................................................................2-10

Set Communication Parameters ....................................................................................2-10

Problems........................................................................................................................2-11

Repackaging the Instrument ..........................................................................................2-12

Before Repackaging the Instrument........................................................................2-12

Attach the Microplate Carrier Shipping Bracket ......................................................2-12

Repackage the Instrument ......................................................................................2-14

Chapter 3: Instrument Qualification ........................................................ 3-1 Recommendations for Achieving Optimum Performance.................................................3-2

Recommended Qualification Schedule ............................................................................3-3

Installation and Performance Qualification Procedures....................................................3-3

Routine Procedure ...........................................................................................................3-4

System Test ..............................................................................................................3-4

Absorbance Plate Test..............................................................................................3-8

Description.........................................................................................................3-8

Test Plate Certificates........................................................................................3-9

Entering the Absorbance Test Plate Data........................................................3-10

Running the Absorbance Plate Test ................................................................3-11

Results.............................................................................................................3-13

Liquid Testing.................................................................................................................3-15

Stock Solution Formulation .....................................................................................3-16

Procedure A.....................................................................................................3-16

Procedure B.....................................................................................................3-17

Liquid Test 1 ...........................................................................................................3-18

Channel-to-Channel Variation..........................................................................3-18

Accuracy Specification.....................................................................................3-19

PowerWave Operator’s Manual v

Liquid Test 2 ...........................................................................................................3-20

Linearity – Test A.............................................................................................3-21

Repeatability – Test B......................................................................................3-21

Channel-to-Channel Variation and Alignment – Test C ...................................3-22

Liquid Test 3 ...........................................................................................................3-24

Procedure A.....................................................................................................3-25

Procedure B.....................................................................................................3-25

Liquid Test 3 Procedure...................................................................................3-25

Repeatability – Test A......................................................................................3-26

Linearity – Test B.............................................................................................3-26

Appendix A: Decontamination and Cleaning.......................................... A-1 Purpose.................................................................................................................... A-2

Decontamination ...................................................................................................... A-3

Tools and Supplies ........................................................................................... A-3

Procedure ......................................................................................................... A-3

Cleaning................................................................................................................... A-4

Tools and Supplies ........................................................................................... A-4

Procedure ......................................................................................................... A-4

Appendix B: Computer Control................................................................ B-1 Overview.................................................................................................................. B-2

Controlling the Reader with Gen5 ............................................................................ B-3

Setting Up Gen5 ............................................................................................... B-3

Problems........................................................................................................... B-4

Getting Started with Gen5................................................................................. B-4

Controlling the Reader with KC4 .............................................................................. B-6

Setting Up KC4 ................................................................................................. B-6

Problems........................................................................................................... B-7

Getting Started with KC4 .................................................................................. B-7

KC4’s OLE Functions........................................................................................ B-8

Controlling the Reader with KCjunior ....................................................................... B-9

Setting Up KCjunior .......................................................................................... B-9

Problems......................................................................................................... B-10

Getting Started with KCjunior.......................................................................... B-10

Controlling the Reader Using ActiveX .................................................................... B-11

Controlling the Reader Using Serial Protocol ......................................................... B-11

vi Preface

Appendix C: Error Codes ......................................................................... C-1 Error Codes in Gen5, KC4, or KCjunior ................................................................... C-2

Error Codes During Operation of the Reader with the Bio-Stack ............................. C-2

Diagnostics .............................................................................................................. C-2

General Errors ......................................................................................................... C-3

Fatal Errors ............................................................................................................ C-11

Appendix D: Instrument Dimensions ...................................................... D-1 Instrument Dimensions for Robotic Interface ........................................................... D-2

Appendix E: Barcode Scanner..................................................................E-1 Internal Plate Barcode Scanner ............................................................................... E-2

Enabling Barcode Scanning in KC4 ......................................................................... E-2

Enabling Barcode Scanning in KCjunior .................................................................. E-3

Specifications for Barcode Labels ............................................................................ E-3

Positioning the Labels on the Plates ........................................................................ E-6

PowerWave Operator’s Manual vii

List of Illustrations

Figure 1-1: The PowerWave With the Bio-Stack ...................................................................1-4

Figure 2-1: Unpacking the PowerWave ..................................................................................2-3

Figure 2-2: Removal and Storage of the Carrier Shipping Bracket and Screws .....................2-4

Figure 2-3: Serial Port and 24 VDC Power Supply Inlet ........................................................2-6

Figure 2-4: Power On/Off Switch and Carrier Eject Button....................................................2-8

Figure 2-5: Attaching the Carrier Shipping Bracket and Screws ..........................................2-13

Figure 2-6: Packing the PowerWave.....................................................................................2-15

Figure 3-1a: Sample Output for the System Self-Test (Sheet 1 of 2) .......................................3-6

Figure 3-1b: Sample Output for the System Self-Test (Sheet 2 of 2) .......................................3-7

Figure 3-2: Sample Standards Certificate for the Absorbance Test Plate ...............................3-9

Figure 3-3: Sample Output for the Absorbance Plate Test....................................................3-12

Figure 3-4: Sample Peak Wavelength Certificate Showing Wavelength of Peak in the Interval Between 580 and 590 nm ...........................................................3-14

Figure D-1: Microplate Carrier Position, Mounting Holes, and Instrument Dimensions ....... D-3

Figure E-1a: Sample of One BioTek Barcode Label From Roll (PN 7121094)....................... E-4

Figure E-1b: Section of Label for Attachment to Microplate ................................................... E-4

Figure E-2: Barcode Label Artwork With Recommended Dimensions ................................. E-5

Figure E-3: Position of Barcode Label (Opposite Well A1) on the Microplate ..................... E-6

List of Tables

Table 2-1: Serial Pinout Description......................................................................................2-9

Table 3-1: Recommended Qualification Schedule.................................................................3-3

Table 3-2: Typical Enzyme-Substrate Combinations and Stopping Solutions ....................3-15

Table 3-3: Stock Solution Formulation for Liquid Test Nos. 1 and 2 .................................3-16

Table 3-4: Test Tube Dilutions ............................................................................................3-20

Table 3-5: Phosphate Buffered Saline 10X Concentrate Solution.......................................3-24

Table B-1: Serial Cable Pin-Out Description ...................................................................... B-11

viii Preface

Contacting BioTek Instruments, Inc.

BIOTEKINSTRUMENTS, INC.

Highland Park, P.O. Box 998 Winooski, Vermont 05404-0998 USA

Customer Service and Sales Internet: www.biotek.com Phone: 888-451-5171 (toll free in the U.S.) 802-655-4740 (outside the U.S.) Fax: 802-655-7941 E-Mail: [email protected]

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: [email protected]

European Coordination Center BioTek Instruments GmbH Kocherwaldstrasse 34 D-74177 Bad Friedrichshall Germany

Internet: www.biotek.de Phone: +49 (0) 7136 968 0 Fax: +49 (0) 7136 968 111 E-Mail: [email protected]

mailto:[email protected]

PowerWave Operator’s Manual ix

Document Conventions

This manual uses the following typographic conventions.

Example Description

!

This icon calls attention to important safety notes.

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.

Note Bold text is primarily used for emphasis.

iii This icon calls attention to important information.

x Preface

Revision History

Revision Date Changes

A 12/01 First Issue.

B 2/02 For the Optical Performance Specifications, changed the maximum allowable Gain on Optics from 6.0 to 10.0 (page 1-5).

Replaced Figures 3-1a and 3-1b, Sample Output for the System Test (pages 3-4 and 3-5).

Updated technical support contact information (pages iii, 1-6, and 1-7).

Made editorial changes.

C 8/03 Updated TAC information in Preface and Ch. 1, and Customer Service and European contact information, Hazards, Standards, Safety Symbols, Intended Use Statement, and Warranty in Preface.

Updated Introduction, Variations, Hardware Features, Software Features, Package Contents, Optional Accessories, and Specifications, Ch. 1.

Enhanced Unpacking/Repackaging sections, Ch. 2; added Fig. 2-5, 2-6.

Revised Ch. 3 sections: System Test, Universal Plate Test, Liquid Testing.

Enhanced Decontamination section; added Cleaning section in App. A.

Enhanced section on KC4™, added section on KCjunior™ in App. B, and replaced previous serial control section with a reference to Bio-Tek’s serial communication protocol specification 7266201-SP.

Replaced Error Codes tables in App. C with more detailed tables from the PowerWave™ HT Service Manual.

Added information about the optional barcode scanner in Ch. 1, and added new App. E, Barcode Scanner.

Referenced new versions of PowerWave™, PowerWave™ 340, and PowerWave™ HT 340 throughout manual.

Included references to KCjunior as an additional primary operating software and substitute for KC4.

Included information about the compatibility of the PowerWave with the Bio-Stack™ Automated Microplate Stacking System, and brief paragraphs concerning installation, serial cable connections, error codes, and alignment when operating the PowerWave with the Bio-Stack; referenced the Bio-Stack Operator’s Manual.

D 5/06 Updated the manual primarily to support introduction of Gen5™ Software.

General: Added Gen5 references and instructions wherever KC4™ and KCjunior™ references and instructions were present. Changed ‘Bio-Tek’ to ‘BioTek,’ ‘Bio-Stack™ Automated Microplate Stacking System’ to ‘Bio-Stack Microplate Stacker,’and ‘Abs’ to ‘OD’. Removed ‘Scanning’ from ‘PowerWave™ Microplate Scanning Spectrophotometer’. (Continued on the following page.)

PowerWave Operator’s Manual xi

Revision History (Cont’d)

Revision Date Changes

D 5/06 (Continued from previous page)

Cover: Replaced cover with new design from Marketing (includes updated photo of PowerWave™ and new BioTek logo).

Preface: Updated contact information, Warnings, Hazards, Precautions and Safety Symbols, and removed Warranty and Registration Card.

Chapter 1, Introduction: Added clarification (in Internal Barcode Scanner section) that some older models of the reader may include the scanner. Updated Package Contents, Optional Accessories, and replaced previous Technical Support pages with a Product Support and Service page.

Chapter 2, Installation: Rearranged installation steps to better reflect actual practice.

Chapter 3, Performance Verification/Qualification Tests: Added Gen5™ instructions for the Self Test and Absorbance Plate Test. In Recommended Qualification Schedule, moved Absorbance Plate Test and Liquid Tests from IQ to initial/annual OQ, changed PQ semiannual frequency to quarterly, and clarified criteria for running Liquid Tests 1, 2, or 3. Changed ‘Universal’ to ‘Absorbance’ in ‘Universal Test Plate’ and ‘Universal Plate Test’. In Liquid Test 1, added BioTek wetting agent (7773002) to list of ingredients. In Liquid Test 3, changed Sigma® ‘P 3563 packets’ to ‘PBS tablets (#4417, or equivalent).’ In Liquid Tests 1 and 3, changed ‘Analytical balance’ to ‘Precision balance.’ In all Liquid Tests, added ‘Corning” to “Costar’ (microplates), and added note to shake plate or wait after pipetting and before reading the plate.

Appendix A, Decontamination and Cleaning: Corrected dilution mixtures for bleach on page A-3 by changing ‘20:1’ ratio for commercial bleach to ‘1:20’, and ‘10:1’ ratio for household bleach to ‘1:10’.

Appendix B, Computer Control: Added new section, “Controlling the Reader with Gen5.”

xii Preface

Intended Use Statement

• The PowerWave™ is an eight-channel, automated, benchtop, general-purpose Microplate Spectrophotometer that performs optical density measurements of samples in a microplate format. The user must evaluate this instrument with PC-based software in conjunction with the specific assay. This evaluation must include the confirmation that performance characteristics for the specific assay are met.

• This system is designed for use with PC-based software only. BioTek’s Gen5™, KC4™, or KCjunior™ software will provide the user with instrument control and data reduction. Alternatively, the ActiveX component will provide programmers with tools to control the instrument for use with independently developed control software.

• The PowerWave can operate with standard robotic systems, such as BioTek’s Bio-Stack™ Microplate Stacker.

• In the European Union: This product may only be used for Research and Development and other non-clinical uses.

• In all other jurisdictions: This product may be used for Research and Development and in vitro Diagnostic Use.

Specimen Preparation

Samples should be obtained, treated, and stored following instructions and recommendations determined by the user’s laboratory.

Quality Control

It is considered good laboratory practice to run laboratory samples according to instructions and specific recommendations included in the package insert or standard laboratory protocol for the test to be conducted. Failure to conduct Quality Control checks could result in erroneous test data.

Repackaging and Shipping

iii If you need to ship the reader to BioTek for service or repair, contact BioTek for a Return

Materials Authorization (RMA) number, and be sure to use the original packing. Other forms of commercially available packing are not recommended and can void the warranty. If the original packing materials have been damaged or lost, contact BioTek to ask for replacement packing.

PowerWave Operator’s Manual xiii

Warnings

! Operate the instrument on a level surface away from excessive humidity.

Bright sunlight or strong incandescent light can reduce the linear performance range and affect the instrument’s readings.

Measurement values may be affected by extraneous particles (such as dust) in the microplate wells. A clean work area is necessary to ensure accurate readings.

When operated in a safe environment according to the instructions in this document, there are no known hazards associated with the instrument. However, the operator should be aware of certain situations that could result in serious injury; these may vary depending on the instrument model. See Hazards and Precautions.

Hazards

Warning! Power Rating. The instrument’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 the utility ground, creating a severe shock hazard. Always connect the power cord directly to a three-prong receptacle with a functional ground.

Warning! Internal Voltage. Always turn off the power switch and unplug the power cord before cleaning the outer surface of the instrument.

Warning! Potential Biohazards. Some assays or specimens may pose a biohazard. Adequate safety precautions should be taken as outlined in the assay’s package insert. Always wear safety glasses and appropriate protective equipment, such as chemically resistant rubber gloves and apron.

Warning! Liquids. Avoid spilling liquids on the reader; fluid seepage into internal components creates a potential for shock hazard. Wipe up all spills immediately. Do not operate the instrument if internal components have been exposed to fluid.

Warning! Unspecified Use. Failure to operate this equipment according to the guidelines and safeguards specified in this manual could result in a hazardous condition.

xiv Preface

Precautions

The following precautions are provided to help avoid damage to the instrument:

Caution: Service. The system should be serviced by BioTek authorized service personnel. Only qualified technical personnel should perform troubleshooting and service procedures on internal components.

Caution: Environmental Conditions. Do not expose the system to temperature extremes. For proper operation, ambient temperatures should remain between 18-40°C. Performance may be adversely affected if temperatures fluctuate above or below this range.

Caution: Sodium Hypochlorite. Do not expose any part of the instrument to the recommended diluted sodium hypochlorite solution (bleach) for more than 20 minutes. Prolonged contact may damage the instrument surfaces. Be certain to rinse and thoroughly wipe all surfaces.

Caution: Power Supply. Only use the power supply shipped with the instrument. Operate this power supply within the range of line voltages listed on it..

Caution: Carrier Shipping Bracket. The microplate carrier shipping bracket must be removed prior to operating the instrument.

Caution: Disposal. 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).”

Caution: Warranty. Failure to follow preventive maintenance protocols may void the warranty. See Appendix A, Decontamination and Cleaning for preventive maintenance procedures.

PowerWave Operator’s Manual xv

Based on the testing described below, this instrument bears the CE mark.

EMC EC DIRECTIVE 89/336/EEC Electromagnetic Compatibility

• Emissions – CLASS A

The system has been type tested by an independent, accredited testing laboratory and found to meet the requirements of EN 61326-1:1998 for Radiated Emissions and Line Conducted Emissions. Verification of compliance was conducted to the limits and methods of the following:

CISPR 16-1:1993 and CISPR 16-2:1999

• Immunity

The system has been type tested by an independent, accredited testing laboratory and found to meet the requirements of EN 61326-1:1998 for Immunity. Verification of compliance was conducted to the limits and methods of the following:

EN 61000-4-2 (1998) Electrostatic Discharge

EN 61000-4-3 (1998) Radiated EM Fields

EN 61000-4-4 (1995) Electrical Fast Transient/Burst

EN 61000-4-5 (1995) Surge Immunity

EN 61000-4-6 (1996) Conducted Disturbances

EN 61000-4-11 (1994) Voltage Dips, Short Interruptions and Variations

EC Directive 73/23/EEC Low Voltage (Safety)

The system has been type tested by an independent testing laboratory and was found to meet the requirements of EC Directive 73/23/EEC for Low Voltage. Verification of compliance was conducted to the limits and methods of the following:

• EN 61010-1 (2001)

“Safety requirement for electrical equipment for measurement, control and laboratory use, 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).”

xvi Preface

Electromagnetic Interference and Susceptibility

USA FCC CLASS A

Warning: Changes or modifications to this unit not expressly approved by the manufacturer could void the user's authority to operate the equipment.

This equipment has been tested and found to comply with the limits for a Class A digital device, pursuant to Part 15 of the FCC Rules.

These limits are designed to provide reasonable protection against harmful interference when the equipment is operated in a commercial environment. Like all similar equipment, this equipment generates, uses, and can radiate radio frequency energy and, if not installed and used in accordance with the instruction manual, may cause harmful interference to radio communications. Operation of this equipment in a residential area is likely to cause interference, in which case the user will be required to correct the interference at his own 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 prescrites dans le Reglement sur le brouillage radioelectrique edicte par le ministere des Communications du Canada.

User Safety

This device has been type tested by an independent laboratory and found to meet the requirements of the

following:

• Underwriters Laboratories UL 61010A-1 1st Edition, 2002

"Electrical Equipment for Laboratory Use; Part 1: General Requirements"

• Canadian Standards Association CAN/CSA C22.2 No. 1010.1-92

"Safety Requirements for Electrical Equipment for Measurement, Control and Laboratory Use; Part 1: General Requirements"

PowerWave Operator’s Manual xvii

Safety Symbols

Some of the following symbols may appear on the instrument.

Alternating current Courant alternatif Wechselstrom Corriente alterna Corrente alternata

Direct current Courant continu Gleichstrom Corriente continua Corrente continua

Both direct and alternating current Courant continu 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 dem Netz) Conectado Chiuso

Off (Supply) Arrêt (alimentation) Aus (Trennung vom Netz) Desconectado Aperto (sconnessione dalla rete di alimentazione)

xviii Preface

Caution (refer to accompanying documents) Attention (voir documents d'accompanement) Achtung siehe Begleitpapiere Atención (vease los documentos incluidos) Attenzione, consultare la doc annessa

Warning, risk of electric shock Attention, risque de choc électrique Gefährliche elektrische Schlag Precaución, riesgo de sacudida eléctrica Attenzione, rischio di scossa elettrica

Warning, risk of crushing or pinching Attention, risque d’écrasement et pincement Warnen, Gefahr des Zerquetschens und Klemmen Precaución, riesgo del machacamiento y sejeción Attenzione, rischio di schiacciare ed intrappolarsi

Warning, hot surface Attention, surface chaude Vorsicht, 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-Diagnostikum Dispositivo médico de diagnóstico in vitro Dispositivo medico diagnostico in vitro

Separate collection for electrical and electronic equipment Les équipements électriques et électroniques font l’objet d’une collecte selective Getrennte Sammlung von Elektro- und Elektronikgeräten Recogida selectiva de aparatos eléctricos y electrónicos Raccolta separata delle apparecchiature elettriche ed elettroniche

Chapter 1 Introduction

This chapter introduces the PowerWave™ and describes its hardware and software features and technical

specifications. Instructions on how to contact BioTek for Product Support & Service are included on page

1-10.

This chapter includes the following sections:

Introducing the PowerWave .................................................................................................................1-2

Variations..............................................................................................................................................1-3

Internal Barcode Scanner......................................................................................................................1-3

Compatibility With the Bio-Stack ........................................................................................................1-4

Hardware Features................................................................................................................................1-5

Software................................................................................................................................................1-5

Package Contents..................................................................................................................................1-6

Optional Accessories ............................................................................................................................1-6

Specifications........................................................................................................................................1-7

Product Support & Service .................................................................................................................1-10

1-2 Chapter 1 - Introduction

Introducing the PowerWave

Robust, reliable, and designed to operate 24 hours a day, seven days a week, BioTek’s PowerWave™ offers tunable wavelength selection and wavelength scanning without the need for interference filters. The eight-channel reader is completely computer controlled via BioTek’s intuitive and user-friendly Gen5™, KC4™, or KCjunior™ software. Its key features include the following:

• A variety of read methods including endpoint, kinetic, multiwavelength, and spectral scanning.

• A monochromator for continuous wavelength selection from

200 to 999 nm in 1 nm increments in the PowerWave™ and PowerWave™ HT models

340 to 999 nm in 1 nm increments in the PowerWave™ 340 and PowerWave™ HT 340 models

• A xenon flash lamp for both UV and visible light absorbance measurements.

• Superior optical specifications, with an extended dynamic range of up to 4.000 OD.

• Ability to read standard 96- and 384-well (HT models) microplates and BioTek’s patented Bio-Cell™ quartz vessel for 1 cm measurements.

• Three reading speeds: normal, rapid, and the ultra fast sweep mode.

• A unique 4-Zone™ temperature control from 4º over ambient to 50ºC that ensures superior temperature uniformity necessary for kinetic assays.

• Low, medium, high, and variable plate shake speeds with adjustable durations.

• Robot accessible carrier. Compatible with BioTek’s optional Bio-Stack™ Microplate Stacker (see page 1-4, Compatibility With the Bio-Stack).

PowerWave Operator's Manual 1-3

Variations

BioTek’s PowerWave is available in the following models:

• PowerWave™: Reads 96-well microplates at wavelength selections from 200 to 999 nm.

• PowerWave™ 340: Reads 96-well microplates at wavelength selections from 340 to 999 nm.

• PowerWave™ HT: Reads 96- and 384-well microplates at wavelength selections from 200 to 999 nm.

• PowerWave™ HT 340: Reads 96- and 384-well microplates at wavelength selections from 340 to 999 nm.

All models include incubation and are compatible with the Bio-Stack™ Microplate Stacker described on the following page.

Internal Barcode Scanner

Some of the earlier models of the PowerWave may include an internal barcode scanner for scanning labels on microplates before the read cycle.

BioTek’s KC4™ and KCjunior™ software include barcode scanner options that must be selected in order to activate the scanner. If your PowerWave model includes the barcode scanner, please refer to Appendix E, Barcode Scanner, for instructions for selecting the barcode option in KC4 or KCjunior, and for specifications for label formats.


Compatibility With the Bio-Stack

The PowerWave™ is compatible with BioTek’s optional Bio-Stack™ Microplate Stacker (see Figure 1-1 below).

Designed to operate with the PowerWave via a PC running Gen5™ or KC4™ software, the Bio-Stack can rapidly and systematically transfer microplates one at a time to and from the reader, and includes:

• Two removable stacks (one input and one output) that can hold up to 30 microplates each.

• Optional automatic restacking of plates to maintain correct sequencing.

• The ability to continue processing a stack of plates following the aborting/failure of one plate.

• The ability to pause processing to add more plates to the input stack or to remove some from the output stack.

If you have purchased the Bio-Stack to operate with the PowerWave, refer to the Bio-Stack Operator’s Manual for instructions on setup and operation of the reader with the Bio-Stack. If you are interested in purchasing the Bio-Stack, contact your local BioTek dealer for more information.

Figure 1-1: The PowerWave with the Bio-Stack


Hardware Features

• Xenon flash lamp

• Eight optics channels, with an additional reference channel

• Monochromator with a range of

200 to 999 nm in the PowerWave™ and PowerWave™ HT models

340 to 999 nm in the PowerWave™ 340 and Power Wave™ HT 340 models

• Low, medium, high, and variable plate shake frequencies with adjustable durations

• Capability of reading 96- and 384-well (HT models) microplates and up to 8 Bio-Cells™

• 24-volt external power supply compatible with 100-240 V~ ± 10% @50-60 Hz

• One serial COM port (25-pin male connector)

• 4-Zone™ incubation

Software

This instrument is controlled using computer control commands sent via the RS-232 interface. BioTek’s Gen5™ software is the primary operating software for the PowerWave, however, BioTek’s KC4™ or KCjunior™ software may be used instead of Gen5. Gen5 or KC4 software is required when using the Bio-Stack™ Microplate Stacker with the PowerWave. Additionally, an ActiveX component is available to allow external control of the reader within other automated environments. Documentation on the serial communication protocol is available upon request.


Package Contents

• PowerWave™, PowerWave™ 340, PowerWave™ HT, or PowerWave™ HT 340 Microplate Spectrophotometer

• Power supply (PN 76053)

• Power cord (PN varies according to country of use)

• Serial cable (PN 75053)

• Operator’s Manual (PN 7281000)

• Unpacking (PN 7281002) and packing instructions (PN 7281003)

• Set of shipping materials (PN 7283000) and 2-ml plastic bag (PN 98085)

• Shipping document (PN 94075) that includes a Warranty Statement and Certificate of Compliance and Calibration

• Declaration of Conformity (PN 7281004)

• Gen5™ Software (PN 5320200)

Optional Accessories

• PowerWave™ Service Manual (PN 7281006)

• 7-Filter Absorbance Test Plate (PN 7260522)

• Bio-Cell™ Quartz vessel for 1 cm wavelength fixed pathlength absorbance measurements (PN 7272051)

• Bio-Cell adaptor plate for containing up to eight Bio-Cells (PN 7270512)

• PowerWave Installation/Operational/Performance Qualification (IQ/OQ/PQ) package (7280520)

• Liquid Test Solutions:

BioTek Wetting Agent (PN 7773002)

BioTek QC Check Solution No. 1 for liquid testing (PN 7120779 for 25 ml; PN 7120782 for 125 ml)

• Gen5™ Reader Diagnostics Utility (PN 5320201)

• KC4™ and KCjunior™ software (PNs and versions listed on biotek.com)

• Bio-Stack™ Microplate Stacker (PN BIOSTACK)

http://www.biotek.com/products/software.php


Specifications

• Microplates

All models accommodate standard 96-well microplates, and up to 8 Bio-Cells™.

PowerWave™ HT models also accommodate standard 384-well microplates.

• Speed of Reading

The actual plate read time and accuracy are dependent on the method of reading:

Normal mode is the slowest of the three available modes. After positioning the well over the beam, the instrument waits 100 milliseconds before taking the measurement (8-flash data collection). Note: The 100 ms delay is to allow for more complete fluid settling.

Rapid mode is faster than Normal mode because the instrument does not wait before taking the measurement (8-flash data collection).

Sweep is the fastest of the three available modes. The plate carrier sweeps each row past the optics channel without stopping, and collects data with a single flash at each well as it goes by.

The following read times are based on a single or dual wavelength measurement. Actual reading speeds may vary, depending upon the reading wavelength selected. Each wavelength has a unique location within the monochromator, and the different locations require varying amounts of time to position.

96-Well Read Timing 630 nm 630/450 nm

Normal Read Mode Single Dual

Endpoint 16 to 25 sec. 26 to 44 sec.

Rapid Read Mode Single Dual

Endpoint 16 sec. 26 sec.

Sweep Read Mode Single Dual


Kinetics:

All three read modes are available in Kinetics mode. Single Wavelength reads are limited to the following minimum times.

- 20 seconds from A1 to A1 in Normal mode, single wavelength, depending upon density of solution.

- 11 seconds from A1 to A1 in Rapid mode, single wavelength.

- 5 seconds from A1 to A1 in Sweep mode, single wavelength.


384-Well Read Timing 630 nm 630/450 nm

Normal Read Mode Single Dual

Endpoint 32 to 67 sec. 57 to 129 sec.

Rapid Read Mode Single Dual

Endpoint 28 sec. 49 to 51 sec.

Sweep Read Mode Single Dual


Kinetics:

All three read modes are available in Kinetics mode. Single Wavelength reads are limited to the following minimum times.

- 66 seconds from A1 to A1 in Normal mode, single wavelength, depending upon density of solution.

- 23 seconds from A1 to A1 in Rapid mode, single wavelength.

- 11 seconds from A1 to A1 in Sweep mode, single wavelength.

• Optical Specifications

λ range: 200 to 999 nm (PowerWave™, PowerWave™ HT) 340 to 999 nm (PowerWave™ 340, PowerWave™ HT 340)

λ accuracy: ± 2 nm

λ repeatability: ± 0.2 nm

λ bandpass: 5 nm

• Optical Performance (flat-bottom and round-bottom full-well plates)

Absorbance Measurement Range: 0.000 to 4.000 OD

Accuracy:

0.000 to 2.000 OD ± 1.0% ± 0.010 OD Normal and Rapid modes, all plates


2.500 to 3.000 OD ± 3.0% ± 0.010 OD Normal 96-well plates only

0.000 to 1.000 OD ± 1.0% ± 0.010 OD Sweep mode, all plates


Linearity:

0.000 to 2.000 OD ± 1.0% Normal and Rapid modes, 96-well plates

0.000 to 2.000 OD ± 2.0% Normal and Rapid modes, 384-well plates

2.000 to 2.500 OD ± 3.0% Normal and Rapid modes, all plates

2.500 to 3.000 OD ± 3.0% Normal mode, 96-well plates only 0.000 to 1.000 OD ± 1.0% Sweep mode, all plates

Repeatability:

0.000 to 2.000 OD ± 1.0% ± 0.005 OD Normal and Rapid modes, 96-well plates

0.000 to 2.000 OD ± 2.0% ± 0.010 OD Normal and Rapid modes, 384-well plates


2.500 to 3.000 OD ± 3.0% ± 0.005 OD Normal mode, 96-well plates only

0.000 to 1.000 OD ± 2.0% ± 0.010 OD Sweep mode, all plates

For the above performance, the Gain on Optics test should be below 10.0.

• Hardware and Environmental Specifications

Display: None

Light Source: Xenon flash light source

- 10 W max. average power - Life: 1 billion flashes

Dimensions: 16.0" deep x 8.5" wide x 8.5" high

(40.6 cm deep x 21.6 cm wide x 21.6 cm high)

Weight: 24 lb. (10.9 kg)

Environment: Operational temperature 18°-40°C

Humidity: 10% to 80%, non-condensing

Power Source: 24-volt external power supply compatible with

100-240 V~ ± 10% @50-60 Hz

Power Consumption: 100 VA max

Incubation option: Temperature Control: 4°C over ambient to 50°C

Temperature Variation: ± 0.5°C across the plate @ 37°C

(250 µl per well with the plate sealed)


Product Support & Service

A superior support staff backs all of BioTek’s products. If your instrument(s) or software ever fails to function perfectly, if you have questions about how to use or maintain it, or if you need to send an instrument to BioTek for service or repair, please contact our Technical Assistance Center (TAC).

Contacting the Technical Assistance Center

Our Technical Assistance Center is open from 8:30 AM to 5:30 PM (EST), Monday through Friday, excluding standard U.S. holidays. You can send a fax or an e-mail any time.

Phone: 800-242-4685 (in the U.S.) or 802-655-4740 (outside the U.S.)

Fax: 802-655-3399

E-Mail: [email protected]

Please be prepared to provide 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

• For troubleshooting assistance or instruments needing repair, the specific steps that produce your problem, and any error codes displayed (see also Appendix C, Error Codes).

Returning Instruments for Service/Repair

If you need to return an instrument to BioTek for service or repair, please contact the TAC for a Return Materials Authorization (RMA) number before shipping the instrument. Repackage the instrument properly (see Chapter 2, Installation), write the RMA number on the shipping box, and ship to this address:

BioTek Instruments, Inc.

Technical Assistance Center 100 Tigan Street Highland Park Winooski, Vermont 05404 USA

mailto:[email protected]

Chapter 2 Installation

This chapter includes instructions for unpacking and setting up the PowerWave™, connecting to a PC, and

repackaging the instrument.

This chapter includes the following sections:

Unpack and Inspect the Instrument ......................................................................................................2-2

Remove the Microplate Carrier Shipping Bracket................................................................................2-4

Select an Appropriate Environment......................................................................................................2-5

Connect the Power Supply and Cord....................................................................................................2-5

Turn on the Reader ...............................................................................................................................2-7

Connect the Host Computer..................................................................................................................2-9

Install Software on the Host Computer...............................................................................................2-10

Set Communication Parameters..........................................................................................................2-10

Problems .............................................................................................................................................2-11

Repackaging the Instrument ...............................................................................................................2-12

Before Repackaging the Instrument ............................................................................................2-12

Attach the Microplate Carrier Shipping Bracket .........................................................................2-12

Repackage the Instrument ...........................................................................................................2-14

2-2 Chapter 2 - Installation

Unpack and Inspect the Instrument

iii Important! Save all packaging materials. If you need to ship the PowerWave™ to BioTek for repair or replacement, you must use the original packing materials. Using other forms of commercially available packing materials, or failing to follow the repackaging instructions, may void your warranty. If the original packing materials have been damaged, replacements are available from BioTek (PN 7283000).

During this process, inspect the packaging, reader, and accessories for shipping damage. If the reader is damaged, notify the carrier and your manufacturer’s representative. Keep the shipping cartons and the packing materials for the carrier’s inspection. The manufacturer will arrange for repair or replacement of your reader immediately, before the shipping-related claim is settled.

Perform these steps to unpack and inspect the reader (refer to Figure 2-1 on the following page):

1. Carefully open the top of the box.

2. Remove the accessories and set them aside.

3. Remove the vertical cardboard insert and the top foam caps.

4. Carefully lift the reader out of the box (and out of the bottom foam caps), and place it on a level surface.

5. Remove the reader from its plastic bag. Place all shipping materials back into the box.


Figure 2-1: Unpacking the PowerWave™

Foam caps (4): 2 top/2 bottom

26 x 32 2-ml plastic bag


Remove the Microplate Carrier Shipping Bracket

iii The PowerWave™ model is shipped with a microplate carrier shipping bracket and

shipping bracket screws that must be removed before the reader is used, and saved in case the instrument needs to be repackaged for shipment. See Figure 2-2 below.

1. Pull down the door to the carrier compartment on the front of the PowerWave.

2. Using a phillips screwdriver, remove the three screws (PN 19186) holding the shipping bracket (PN 7282014) to the mounting block and carrier.

3. Screw the shipping bracket to the back of the reader for storage.

Figure 2-2: Removal and storage of the carrier shipping bracket and screws

Carrier

Shipping bracket and 3 screws

Mounting block


Select an Appropriate Environment

For optimal operation, install the PowerWave™ on a level surface in an area where ambient temperatures between 18°C and 40°C can be maintained. The reader is sensitive to extreme environmental conditions. Conditions to avoid are:

• Excessive humidity: Condensation directly on the sensitive electronic circuits can cause the instrument to fail internal self-checks.

• Excessive ambient light: Bright sunlight or strong incandescent light can reduce the linear performance range and affect the instrument’s readings.

• Dust: Optical density readings may be affected by extraneous particles (such as dust) in the microplate wells. A clean work area is necessary to ensure accurate readings.

Connect the Power Supply and Cord

!

Warning! Power Rating. The PowerWave power supply 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 the utility ground, creating a severe shock hazard. Always connect the power cord directly to a three-prong receptacle with a functional ground.

Refer to Figure 2-3 on page 2-6 for the following instructions.

1. Connect the power cord to the external 24-volt power supply.

2. Locate the power supply inlet on the rear of the PowerWave, below the serial port. See Figure 2-3.

3. Plug the power supply outlet plug into the power supply inlet. Tighten the plug barrel.

4. Plug the 3-prong end of the power cord into an appropriate power receptacle.


Figure 2-3: Serial port and 24 VDC power supply inlet

Power supply inlet

Serial port


Turn on the Reader

iii The PowerWave™ does not need to be connected to a PC for the following test.

1. Locate the power on/off switch on the front of the PowerWave, below the carrier eject button. See Figure 2-4 on the following page. Turn on the power. The reader will perform an internal Self-Test and carrier homing sequence.

2. Verify that the following occur while the reader performs the Self-Test:

• The carrier should eject outside the PowerWave, then retract to its home position inside the reader before it ejects again.

• The LED light should remain illuminated while the power is on.

3. Press the carrier eject button. The carrier should retract and the door should close. Press it again – the carrier should eject.

4. If the test completes successfully, proceed to the next section, Connect the Host Computer.

5. If the test fails, the reader will “beep” continuously. Press the carrier eject button to stop the beeping, and proceed to the following sections on pages 2-9 and 2-10 for attachment of the serial cable, installation of Gen5™, KC4™, or KCjunior™, and setting the communication parameters. Run the Self-Test to retrieve an error code (see the Problems section on page 2-11); if the Self-Test still fails, contact BioTek.


Figure 2-4: Power on/off switch and carrier eject button

Carrier eject button On/off switch

Microplate A1 position


Connect the Host Computer

iii If you have purchased BioTek’s Bio-Stack™ Microplate Stacker for operation with the PowerWave™, you will need a PC equipped with two RS-232 serial ports and any version of Gen5™ or the appropriate version of KC4™ software. Refer to the Bio-Stack Operator’s Manual for more information.

The serial port on the rear panel of the PowerWave is a DTE configuration with a 25-pin (male) D-sub connector. Refer to Figure 2-3 on page 2-6 for the location of the serial port, and to Table 2-1 below for a description of the serial/RS-232 pin connection of the reader.

1. Turn off the reader. If the computer is on, turn it off.

2. Using the supplied serial cable, attach one end of the cable to a serial port on the computer.

3. Attach the other end of the cable to the serial port on the rear of the PowerWave.

4. Tighten the securing screws on both ends of the cable.

Table 2-1

Serial Pinout Description

Serial Pin Description

Pin Signal Pin Signal 1 NC 14 NC 2 TX 15 NC 3 RX 16 NC 4 RTS 17 NC 5 CTS 18 NC 6 DSR 19 NC 7 GND 20 DTR 8 DCD 21 NC 9 NC 22 RI 10 NC 23 NC 11 NC 24 NC 12 NC 25 NC 13 NC


Install Software on the Host Computer

The PowerWave™ is controlled by Gen5™, KC4™, or KCjunior™ software running on a host PC. To install Gen5, KC4, or KCjunior:

1. Ensure that the appropriate serial cable has been connected between the reader and controlling computer as instructed on the previous page.

2. Turn on the computer.

3. Install Gen5, KC4, or KCjunior software on the computer’s hard drive. Refer to Gen5’s Getting Started Guide or Help system or to the KC4 or KCjunior User’s Guides for instructions.

iii If you purchased Gen5’s Reader Diagnostics Utility, PN 5320201 (required for the Absorbance Test Plate), install this software on the computer’s hard drive.

Set Communication Parameters

Before serial communication can be initiated between the PowerWave and the host PC running Gen5, KC4, or KCjunior, the communication parameters must match on both devices. The reader’s default communication parameters are:

• Baud Rate: 9600

• Data Bits: 8

• Parity: None

• Stop Bits: 2

See Appendix B, Computer Control, for instructions for defining the communication parameters in Gen5, KC4, or KCjunior.

iii Although the baud rate may be changed through KC4 or KCjunior, the factory setting is 9600; therefore, changing this rate is not recommended. In Gen5, the baud rate for the PowerWave is set at the default 9600 and cannot be changed.

The Data Bits, Parity and Stop Bits cannot be changed.


Problems

Each time the reader is turned on, it performs a Self-Test. If the test fails, note any error codes that are displayed in Gen5™, KC4™, or KCjunior™, and refer to Appendix C, Error Codes, for a comprehensive list of the codes and their probable causes. If the problem cannot be resolved, contact BioTek Technical Support.

The system Self-Test and other diagnostics tests may be run in Gen5, KC4, or KCjunior. See Chapter 3, Instrument Qualification, for instructions on running diagnostic tests and for a recommended Qualification Schedule that includes Installation, Operational and Performance Qualification tasks.


Repackaging the Instrument

Before Repackaging the Instrument

1. Decontaminate the reader prior to shipping (see Appendix A, Decontamination and Cleaning).

2. Replace the carrier shipping bracket and the three screws according to the instructions below.

Attach the Microplate Carrier Shipping Bracket

iii The microplate carrier shipping bracket and three shipping bracket screws must be re-attached to the carrier before the PowerWave™ can be shipped. See Figure 2-5 on the following page.

1. If the carrier has not been returned to its home position inside the reader, press the carrier eject button to retract the carrier.

2. Turn off the PowerWave, and unplug the 24 VDC power supply from the power outlet and from the power supply connector on the back of the reader. Remove the serial cable from the reader.

3. Using a screwdriver, remove the carrier shipping bracket and screws from the back of the reader.

4. Pull down the door to the carrier compartment.

5. Install the carrier shipping bracket to the front of the carrier and mounting block.


Figure 2-5: Attaching the carrier shipping bracket and screws

Mounting block

Carrier

Shipping bracket and 3 screws


Repackage the Instrument

iii Important! Use the instrument’s original shipping container and packaging material. This shipping system was designed to be used no more than five times. If the container is damaged and/or has been used more than five times, contact BioTek for a new set of shipping materials, and ask for PN 7283000. The shipping box, vertical cardboard insert (for accessories), four foam caps, etc. are included as a whole set under this part number and cannot be ordered separately.

Ensure that the PowerWave™ carrier shipping bracket and three shipping screws have been attached to the reader’s carrier and mounting block as instructed in the preceding section, Attach the Microplate Carrier Shipping Bracket. Refer to Figure 2-6 on the following page when performing these steps:

1. Place two foam caps into the bottom of the shipping container. Note the orientation of the foam caps in the box as illustrated in Figure 2-6.

2. Place the reader inside the original 26 x 32 2-ml plastic bag and carefully lower the reader into the two foam caps in the bottom of the box. Note the orientation of the reader in the box.

3. Place two foam caps over the reader.

4. Bundle the power cord and place it into the accessories box as shown.

5. Close the top of the box and secure it with shipping tape.

6. A Return Materials Authorization (RMA) number must be obtained before returning equipment for service. Contact BioTek’s Technical Assistance Center for this number, then write “RMA” and the RMA number in large, clear letters on the outside of the shipping container.

7. Return the box to BioTek at the address provided in the Product Support & Service section of Chapter 1.


Figure 2-6: Packing the PowerWave™

Power cord

Top foam caps (2)

26 x 32 2-ml plastic bag

Bottom foam caps (2)

Chapter 3 Instrument Qualification

This chapter discusses the tasks and procedures necessary for verifying and qualifying instrument performance on an ongoing basis. A convenient Recommended Qualification Schedule arranges tasks into Installation, Operational and Performance Qualification categories.

This chapter contains the following sections:

Recommendations for Achieving Optimum Performance....................................................................3-2

Recommended Qualification Schedule.................................................................................................3-3

Installation and Performance Qualification Procedures .......................................................................3-3

Routine Procedure ................................................................................................................................3-4

System Test ...................................................................................................................................3-4

Absorbance Plate Test ..........................................................................................................................3-8

Description ....................................................................................................................................3-8

Test Plates Certificates ..................................................................................................................3-9

Entering the Absorbance Test Plate Data ....................................................................................3-10

Running the Absorbance Plate Test.............................................................................................3-11

Results .........................................................................................................................................3-13

Liquid Testing.....................................................................................................................................3-15

Stock Solution Formulation.........................................................................................................3-16

Liquid Test 1 ...............................................................................................................................3-18

Liquid Test 2 ...............................................................................................................................3-20

Liquid Test 3 ...............................................................................................................................3-24

3-2 Chapter 3 - Instrument Qualification

Recommendations for Achieving Optimum Performance

Note: An Installation/Operational/Performance Qualification (IQ/OQ/PQ) package (PN 7280520) for the PowerWave™ is available for purchase. Contact your local dealer for more information.

• Microplates should be perfectly clean and free of dust or bottom scratches. Use new microplates from sealed packages. Do not allow dust to settle on the surface of the solution; use microplate covers when not reading the plate. Filter solutions to remove particulates that could cause erroneous readings.

• Although the PowerWave supports most flat, U-bottom, and V-bottom microplates, the reader achieves optimum performance with optically clear, flat-bottomed wells.

• Non-uniformity in the optical density of the well bottoms 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 results, use at least 100 µl per well in a 96-well plate and 25 µl in a 384-well plate (PowerWave™ HT models).

• 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 inaccuracies; however, for best results, remove the air bubbles by degassing the plate in a vacuum chamber before reading.

• The inclination of the meniscus can cause loss of accuracy in some solutions, especially with small volumes. Agitate the microplate before reading to help bring this problem within acceptable limits. Use Tween 20, if possible (or some other wetting agent) to normalize the meniscus. Some solutions develop menisci over a period of several minutes. This effect varies with the brand of microplate and the solution composition. As the center of the meniscus drops and shortens the light path, the density readings change. The meniscus shape will stabilize over time.

PowerWave Operator’s Manual 3-3

Recommended Qualification Schedule

The schedule shown in Table 3-1 below defines the factory-recommended intervals for performance testing for a microplate reader used for one shift seven days a week.

Note: The risk factors associated with your tests may require that the Operational and Performance Qualification procedures be performed more or less frequently than shown below.

Table 3-1 Recommended Qualification Schedule

Installation Qualification

Operational Qualification

Performance Qualification

Initially Initially/ Annually Monthly Quarterly

System Self-Test, p. 3-4

Absorbance Plate Test, p. 3-8

Liquid Test 1*, p. 3-18

Liquid Test 2*, p. 3-20

Liquid Test 3**, p. 3-24

* If you have an Absorbance Test Plate, run Liquid Test 1. If you do not have a Test Plate, run Liquid Test 2.

** Liquid Test 3 is optional; it is provided for sites requiring verification at wavelengths lower than those attainable with the Absorbance Test Plate.

Installation and Performance Qualification Procedures

Tests outlined in this section may be used to confirm initial and ongoing performance of the PowerWave™.

Your PowerWave reader was fully tested at BioTek prior to shipment and should operate properly upon initial setup. If it is suspected that problems may have occurred during shipment, if you reshipped the reader, or if regulatory requirements dictate that Performance Qualification is necessary, the following tests should be performed.

• System Self-Test: Verifies proper gains, bulb operation, low electronic noise, and optional incubator functionality. This test also prints the reader Serial Number and basecode software part number and version number.

• Absorbance Plate Test: Uses BioTek Absorbance Test Plate to confirm the mechanical alignment, optical accuracy/linearity, repeatability, channel-to-channel variation, and wavelength accuracy of the instrument.

• Liquid Testing: Uses liquid solutions in a microplate to confirm mechanical alignment, optical accuracy/linearity, repeatability, and channel-to-channel variation of the instrument.


Routine Procedure

iii Ensure that you have defined the communication parameters in Gen5™, KC4™, or KCjunior™ for the PowerWave™ (see Appendix B), or you will not be able to perform the following tests.

Refer to the Recommended Qualification Schedule on the previous page for the factory-recommended intervals for performing the System Test, the Absorbance Plate Test, and the Liquid Tests.

System Test

iii Important! Do not turn on the instrument until the carrier shipping bracket has been removed.

Note: The desired wavelengths must be entered into the Gen5, KC4, or KCjunior wavelength table and downloaded to the reader before running the test.

• To adjust the wavelength table using Gen5, select System > Reader Configuration. At the ‘Reader Configuration’ dialog, highlight the PowerWave, then click View/Modify > Setup. Click the Absorbance tab and enter the recommended wavelengths. Click Send Wavelengths to download them to the reader.

• To adjust the wavelength table using KC4, select System > Readers. Highlight the correct PowerWave model, then click Filters/Wavelengths. Enter the recommended wavelengths and click Send Wavelengths to download them to the reader.

• To adjust the wavelength table using KCjunior, create a new protocol (for example, click the New Protocol button or select Protocols > New Protocol). Set the Read

Method Type to MultiWavelength. Enter the recommended wavelengths, then click Calibrate Reader.

Each time it is powered on, the PowerWave™ automatically runs the System Test. The instrument will “beep” repeatedly if the System Test results do not meet the internally coded Failure Mode Effects Analysis (FMEA) criteria established by BioTek. If this happens, press the carrier eject button to stop the beep and then attempt to run the System Test using Gen5, KC4, or KCjunior to retrieve an error code from the instrument. See Appendix C for a list of possible error codes.


• To run the System Test using Gen5, select System > Diagnostics > Run System

Test. If the Instrument Selection dialog appears (Gen5 and Gen5 Secure only), select PowerWave, then click OK. The test will run and results will appear in Gen5 in a pass/fail format.

• To run the System Test using KC4™, select System > Diagnostics > Run Optics

Test. Enter the Reader Serial Number, then click Start. The test will run and results will appear in KC4 in a pass/fail format.

• To run the System Test using KCjunior™, select Utilities > Diagnostics > Reader

System Test. Enter the Reader Serial Number, then click Run Test. The test will run and results will appear in KCjunior in a pass/fail format.

The System Test results should be printed for documenting periodic testing and for troubleshooting purposes. See the sample System Test report in Figure 3-1 on the following page.


Figure 3-1a: Sample output for the System Self-Test (Sheet 1 of 2) (Note: The format varies depending on the software used to run the test.)

Test Date/Time: 02/06/2006 12:56:49 PM Reader: 123456 Operator: System Administrator Comments: System Test performed during IQ Operator ID:______________________________________________________________ Notes:____________________________________________________________________ __________________________________________________________________________ SYSTEM SELF TEST 7280201 Version 1.20 128790 Lambda: 200 Gain: 3.28 Resets: 1 Channel: Ref 1 2 3 4 5 6 7 8 Air: 20506 35689 37068 35775 39588 36708 32962 35604 34634 Dark: 9911 10002 10031 9880 9980 9904 9930 9869 9974 Delta: 10595 25687 27037 25895 29608 26804 23032 25735 24660 Lambda: 405 Gain: 2.53 Resets: 2 Channel: Ref 1 2 3 4 5 6 7 8 Air: 20072 35592 39355 36205 38232 37636 33564 36682 36095 Dark: 9914 9960 9975 9900 9945 9915 9928 9895 9948 Delta: 10158 25632 29380 26305 28287 27721 23636 26787 26147 Lambda: 490 Gain: 1.72 Resets: 2 Channel: Ref 1 2 3 4 5 6 7 8 Air: 20280 35642 39779 36310 38486 37962 33494 36949 36219 Dark: 9919 9949 9960 9909 9939 9919 9928 9905 9942 Delta: 10361 25693 29819 26401 28547 28043 23566 27044 26277 Lambda: 550 Gain: 2.51 Resets: 1 Channel: Ref 1 2 3 4 5 6 7 8 Air: 20248 35530 39702 36099 38344 37862 33339 36781 36154 Dark: 9914 9985 10007 9891 9967 9910 9930 9881 9963 Delta: 10334 25545 29695 26208 28377 27952 23409 26900 26191 Lambda: 630 Gain: 3.94 Resets: 1 Channel: Ref 1 2 3 4 5 6 7 8 Air: 20230 35302 39381 35837 38113 37659 33193 36503 35852 Dark: 9907 10018 10053 9870 9990 9900 9931 9854 9984 Delta: 10323 25284 29328 25967 28123 27759 23262 26649 25868 Lambda: 999 Gain: 5.33 Resets: 1 Channel: Ref 1 2 3 4 5 6 7 8 Air: 19977 34186 39376 35131 37500 36779 32458 36029 35249 Dark: 9900 10050 10095 9849 10010 9888 9934 9831 10004 Delta: 10077 24136 29281 25282 27490 26891 22524 26198 25245 Channel: Ref 1 2 3 4 5 6 7 8 Noise Max: 9878 9980 10018 9847 9923 9893 9919 9836 9964 Noise Min: 9877 9978 10017 9846 9922 9891 9917 9835 9962 Delta: 1 2 1 1 1 2 2 1 2


Figure 3-1b: Sample output for the System Self-Test (Sheet 2 of 2) (Note: The format varies depending on the software used to run the test.)

INCUBATOR SELF TEST Temperature Setpoint: 37.0 Current Average: 37.1 A/D Test: PASS Zone 1: 37.1 Min: 37.0 Max: 37.1 Range: PASS Thermistor: PASS Zone 2: 37.0 Min: 37.0 Max: 37.1 Range: PASS Thermistor: PASS Zone 3: 37.1 Min: 37.1 Max: 37.1 Range: PASS Thermistor: PASS Zone 4: 37.1 Min: 37.0 Max: 37.1 Range: PASS Thermistor: PASS AUTOCAL ANALYSIS Upper Left Corner: x= 1676 y= 9220 Lower Left Corner: x= 2448 y= 8944 Lower Right Corner: x= 11140 y= 8912 Upper Right Corner: x= 10364 y= 9172 Delta 1: 1676 - 2448= -772 Delta 2: 10364 -11140= -776 Delta 3: 9172 - 9220= -48 Delta 4: 8912 - 8944= -32 Middle Sensor: x= 13852 Tested: 13852 Delta: +0 SYSTEM TEST PASS


Absorbance Plate Test

iii Gen5’s Reader Diagnostics Utility must be installed on the host PC in order to run the Absorbance Plate Test in Gen5™.

Description

This test uses BioTek’s 7-Filter Absorbance Test Plate (PN 7260522) to confirm the mechanical alignment, optical density accuracy/linearity, repeatability, channel-to-channel variation and wavelength accuracy of the PowerWave™.

The Absorbance Plate Test (also referred to as the ‘Universal Plate Test’ in KC4™ or KCjunior™) compares the reader’s optical density and wavelength measurements to NIST-traceable values.

Note: An alternate method that may be used to determine accuracy/linearity, repeatability, and alignment is Liquid Test 2, described on page 3-20.

The Absorbance Plate Test confirms the following:

• Mechanical Alignment. Mechanical alignment of the plate carrier and standard microplates is confirmed by the four-corner positional accuracy check.

• Accuracy/Linearity. Accuracy of the optical density readings at specific wavelengths is confirmed by comparing the actual readings with those supplied with the Absorbance Test Plate Standards Certificate. Linearity of the optical density readings is confirmed by default if the optical density readings are accurate.

• Repeatability. Repeatability of readings is confirmed by reading each filter twice and comparing the two readings.

• Channel-to-Channel Variation. Ensures that selected channels read the same value for a filter as their paired channel when the plate is “turned around” 180° in the plate carrier.

• Wavelength Accuracy (Peak Wavelength/Peak Absorbance). To check the wavelength accuracy, use BioTek's 7-Filter Absorbance Test Plate (PN 7260522). This Absorbance Test Plate provides a multiband test filter in location C6, for the “Peak Wavelength” or “Peak Absorbance” Test.

Note: BioTek’s 6-Filter Absorbance Test Plate (PN 9000547) does not have a filter in

the C6 location. This test plate can be used on any BioTek reader; however, if this

plate is used, the Peak Wavelength/Peak Absorbance Test will not be performed.


Test Plates Certificates

To run the Absorbance Plate Test on the PowerWave™, you will need BioTek's 7-Filter Absorbance Test Plate (PN 7260522), with its accompanying certificates.

• The Standards Certificate contains standard OD values for the filters at several different wavelengths. See the sample Standards Certificate in Figure 3-2 below.

• The Peak Wavelength Certificate contains one or more “Peak Wavelength” values for the glass filter in position C6 on the plate. Each value has a valid test range associated with it. For example, a Peak Wavelength value may be 586 nm with a test range of 580 to 590 nm (or tolerance values of –6/ +4). See the sample Peak Wavelength Certificate in Figure 3-4 on page 3-14.

Before the Absorbance Plate Test can be performed, the standard OD values and the peak wavelength value(s) must be entered into Gen5™, KC4™, or KCjunior™. This only has to be done once.

Instructions for entering the Absorbance Test Plate data and for running the test are provided for Gen5, KC4, and KCjunior on the following pages.

Figure 3-2: Sample Standards Certificate for the Absorbance Test Plate

SAMPLE


iii Note: The instructions provided below and on the following page are guidelines. Refer to the Absorbance Plate Test sections of the Gen5™ Help system or to the Universal Plate Test section of the KC4™ or KCjunior™ User’s Guides for detailed instructions when entering the Absorbance Test Plate data and running the Absorbance Plate Test.

Entering the Absorbance Test Plate Data

Before the Absorbance Plate Test can be performed, the wavelengths’ settings and the calibration data for each wavelength selected must be initially entered into Gen5, KC4, or KCjunior. Use the Standards Certificate and Peak Wavelength Certificate included with the Absorbance Test Plate for the following:

• In Gen5, select System > Diagnostics > Test Plates > Add/Modify Plates, then click Add. Click the Help button for guidance when setting the wavelengths and entering the OD and peak wavelength value(s).

• In KC4, select System > Diagnostics > Define Universal Plates, then click Add. Click the Help button for guidance when setting the wavelengths and entering the OD and peak wavelength value(s).

• In KCjunior, select Utilities > Diagnostics > Universal Plate Test, then click New

Data Sheet. Click the Help button for guidance when setting the wavelengths and entering the OD and peak wavelength value(s).

The wavelengths and corresponding calibration data that have been entered will now be available in Gen5, KC4, or KCjunior each time the Absorbance Plate Test is performed.


Running the Absorbance Plate Test

iii Notes: Gen5’s Reader Diagnostics Utility must be installed before you can run the Absorbance Plate Test in Gen5™.

The desired wavelengths must be entered into Gen5’s, KC4’s, or KCjunior’s wavelength table and downloaded to the reader before running the test.

• Place the Absorbance Test Plate in the carrier so that well A1 is in the right rear corner of the carrier. (See Appendix D, Instrument Dimensions for orientation.)

• To run the test in Gen5, select System > Diagnostics > Test Plates > Run. Select the appropriate reader, if the Instrument Selection dialog appears. Select the appropriate test plate, if the Select Test Plate dialog appears. Enter the reader Serial Number and User name (required information), then click Start Test.

• To run the test in KC4™, select System > Diagnostics > Run Universal Plate Test. Select the appropriate Universal plate, enter the reader Serial Number, then click Run

Test.

• To run the test in KCjunior™, select Utilities > Diagnostics > Universal Plate Test. Select a Universal Plate ID (Wavelength), enter the Reader Serial Number, then click Run Test.

• For wavelength accuracy: While performing the steps above, click Perform peak

wavelength test in Gen5 or KCjunior, or click Peak absorbance search in KC4.

When the test is complete, print the results. A sample test report is provided in Figure 3-3 on the following page.


Figure 3-3: Sample output for the Absorbance Plate Test (Note: The format varies depending on the software used to run the test.)

Universal Test Plate Analysis Wavelength: 405 Alignment Results A1= 0.000 PASS A12= 0.000 PASS H1= 0.000 PASS H12= 0.000 PASS Accuracy Results C01 D04 E02 F05 G03 H06 STANDARD 0.147 2.945 0.618 2.279 1.133 1.701 DATA 0.139 2.914 0.613 2.265 1.128 1.694 RESULT PASS PASS PASS PASS PASS PASS F12 E09 D11 C08 B10 A07 STANDARD 0.147 2.945 0.618 2.279 1.133 1.701 DATA 0.145 2.910 0.613 2.264 1.125 1.692 RESULT PASS PASS PASS PASS PASS PASS Repeatability Results C01 D04 E02 F05 G03 H06 READ1 0.139 2.914 0.613 2.265 1.128 1.694 READ2 0.141 2.913 0.616 2.265 1.128 1.694 RESULT PASS PASS PASS PASS PASS PASS F12 E09 D11 C08 B10 A07 READ1 0.145 2.910 0.613 2.264 1.125 1.692 READ2 0.140 2.914 0.614 2.265 1.126 1.692 RESULT PASS PASS PASS PASS PASS PASS Turnaround Results C01 D04 E02 F05 G03 H06 READ1 0.139 2.914 0.613 2.265 1.128 1.694 F12 E09 D11 C08 B10 A07 READ2 0.145 2.910 0.613 2.264 1.125 1.692 RESULT PASS PASS PASS PASS PASS PASS Spectral Scan Results 580nm= 1.933 586nm= 2.892 581nm= 1.956 587nm= 2.753 582nm= 2.059 588nm= 2.516 583nm= 2.253 589nm= 2.334 584nm= 2.511 590nm= 2.186 585nm= 2.785 Test Plate Standard=587nm Calculated Peak=586nm PASS


Results

iii Note: For the following Accuracy, Linearity, Repeatability, and Channel-to-Channel

Variation tests, there may not be a pass/fail indication for filter values that are beyond the specified accuracy range of the instrument.

The Absorbance Test Plate Analysis contains results for the following:

• Mechanical Alignment: This portion of the test measures the alignment of the microplate carrier with the optical path. A reading greater than 0.015 OD represents an out-of-alignment condition. Wells A01, A12, H01, and H12 are the only valid alignment holes for the reader on the PN 7260522 Absorbance Test Plate.

• Accuracy/Linearity: Accuracy is a measure of the absorbance (optical density) of Absorbance Test Plate wells C01, D04, E02, F05, G03, and H06 as compared with known standard values contained in the Standards Certificate that accompanies each Test Plate.

Linearity of the optical density readings is confirmed by default if the optical density readings are accurate. To further verify this, perform a regression analysis on the Test Plate OD values in a program such as Microsoft Excel as follows:

In Microsoft Excel, open a spreadsheet and label one column “Assigned” and the next column “Observed.”

Note: Analyze one wavelength at a time.

1. Enter the Assigned OD data for each glass filter in the first column from the

Standards Certificate provided with the Test Plate.

2. Next, enter the Observed OD values for the same glass filters in the adjacent

column.

3. Under Tools, select Data Analysis, and then Regression. (If the Data Analysis

command is not available on the Tools menu, you may need to install the

Analysis ToolPak in Microsoft Excel. Consult Microsoft Excel Help for

assistance.)

4. Use the Regression “Input” box to enter the Assigned values as the “Input Y

Range” and the Observed OD as the “Input X Range.”

5. Click the OK box and the Summary Output sheet will be displayed. An R

Square value of at least 0.99 is expected and the results will often be 0.999 or

better.

• Repeatability: Repeatability is a measure of the instrument’s ability to read the same well with minimum variation between the two reads with the well in the same location.


• Channel-to-Channel Variation: This test ensures that selected channels read the same value for a filter as their paired channel when the plate is “turned around” 180° in the plate carrier. The channel/well “pairs” for the turnaround test are: C01/F12; D04/E09; E02/D11; F05/C08; G03/B10; H06/A07.

• Wavelength Accuracy (Peak Wavelength/Peak Absorbance): If ‘Perform peak wavelength test’ (Gen5™, KCjunior™) or ‘Peak absorbance search’ (KC4™) is enabled as part of the Absorbance Plate Test, the C6 filter will be scanned across a specified wavelength range in 1-nm increments. The wavelength of the maximum absorbance will be compared with the peak wavelength value(s) entered in Gen5, KC4, or KCjunior, which comes from the Peak Wavelength Certificate supplied with the test plate (see the sample in Figure 3-4 below; note that the format of your Certificate may be different). The accuracy of the wavelength should be ± 3 nm (± 2 nm instrument, ± 1 nm filter allowance).

For example, if the test range is 580 to 590 nm, the Certificate value is 587 nm, and the reader reports a peak value of 590 nm, then the reader meets specifications. If the reader reports 591 nm, then the reader does not meet specifications.

Figure 3-4: Sample Peak Wavelength Certificate showing wavelength of peak in the interval between 580 and 590 nm


Liquid Testing

Liquid testing tests the reader in ways that the Absorbance Test Plate cannot. The test plate will indicate the absolute amount of light absorbed, which will effectively test the electronics. The liquid test will help detect optical defects such as dirt on the lenses or contamination that can contribute to errant readings.

• If you have the Absorbance Test Plate, you will only need to perform Liquid Test 1 for routine testing.

• If you do not have the Absorbance Test Plate, you can test the accuracy/linearity, repeatability, and alignment of the reader by performing Liquid Test 2.

• If you must test the reader’s performance at 340 nm, perform Liquid Test 3.

• 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 use the dye solution described in Table 3-3. The purpose of the formulation is to create a solution that absorbs light in a well-defined manner at ~2.000 OD full strength when dispensed at 200 µl in a flat-bottom microplate well.

• Alternatively, any solution that gives a stable color will suffice. (This includes substrates incubated with an enzyme preparation and then stopped with an acidic or basic solution.) Some enzyme/substrate combinations that may be used as alternates to the described dye are shown in Table 3-2 below.

Table 3-2 Typical Enzyme-Substrate Combinations and Stopping Solutions

Enzyme Substrate Stopping Solution

Alkaline Phosphate o-nitrophenyl phosphate 3N sodium hydroxide

beta-Galactosidase o-nitrophenyl -beta-D galactopyranoside 1M sodium carbonate

Peroxidase 2,2'-Azino di-ethylbenzothiazoline-sulfonic acid (ABTS)

citrate-phosphate buffer, pH 2.8

Peroxidase o-phenylenediamine 0.03N sulfuric acid


Stock Solution Formulation

The stock solution for Liquid Tests No. 1 and No. 2 may be formulated from the chemicals listed below, or by diluting a dye solution available from BioTek. See Procedure A or B outlined below and on the following page for details.

Procedure A

Required Materials:

• Deionized water

• FD&C Yellow #5 dye powder (typically 90% pure)

• Tween 20 (polyoxyethylene (20) sorbitan monolaurate, a wetting agent) or BioTek wetting agent, PN 7773002

• Precision balance

• 1-liter volumetric flask

Table 3-3 Stock Solution Formulation for Liquid Test Nos. 1 and 2

FD&C Yellow #5 dye powder 0.092 g

Tween 20 0.5 ml

Deionized water to bring volume to: 1000 ml

Preparation of Stock Solution:

1. Weigh out 0.092 gram of FD&C Yellow #5 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. Mix up to 1 liter with DI water; cap and shake well.

This should create a solution with an absorbance of about 2.000 OD when using 200 µl in a flat-bottom microwell. The OD value will be proportional to the volume in the well and the amount of FD&C Yellow #5 dye used. You can use a larger or smaller well volume, or add more dye or water to adjust the solution.

iii Note: Too small a well volume may result in increased pipetting-related errors.


Procedure B

Required Materials:

• BioTek QC Check Solution No. 1 (PN 7120779, 25 ml; or 7120782, 125 ml)

• Deionized water

• 5-ml Class A volumetric pipette

• 100-ml volumetric flask

Preparation of Stock Solution:

1. Pipette a 5-ml aliquot of BioTek QC Check Solution No. 1 into a 100-ml volumetric

flask.

2. Mix up to 100 ml with DI water; cap and shake well.

This should create a solution with an absorbance of about 2.000 OD when using 200 µl in a flat-bottom microwell. The OD value 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 adjust the stock solution.

iii Note: Too small a well volume may result in increased pipetting-related errors.


Liquid Test 1

Note: A 96-well, flat-bottom microplate is required for this test (Corning Costar #3590 is recommended). Use a new microplate; any fingerprints or scratches may cause variations in the readings.

1. Using a freshly prepared stock solution (see Procedure A on page 3-16 or Procedure B on page 3-17), prepare a 1:2 dilution using deionized water (one part stock, one part deionized water; the resulting solution is a 1:2 dilution). The concentrated stock solution should have an optical density of approximately 2.000 OD or lower.

2. Pipette 200 µl of the concentrated solution into the first column of wells in the new, 96-well microplate.

3. Pipette 200 µl of the diluted solution into the second column of wells.

Important: After pipetting the diluted test solution into the microplate and before reading the plate, we strongly recommend shaking the plate at Variable speed for four minutes. This will allow any air bubbles in the solution to settle and the meniscus to stabilize. Alternatively, wait 20 minutes after pipetting the diluted test solution before reading the plate.

4. Read the microplate five times at 405 nm using normal reading mode, single wavelength, no blanking (“Normal” plate position).

5. Without delay, rotate the microplate 180° so that well A1 is now in the H12 position. Read the plate five more times (“Turnaround” plate position).

The ten sets of raw plate data can be exported to an Excel spreadsheet using Gen5™, KC4™, or KCjunior™. The mathematical computations described below may then be performed and the template kept for future data reduction.

Channel-to-Channel Variation:

6. Calculate the mean value for each physical well location in columns 1 and 2 for the five plates read in the Normal position, and then again for the five plates read in the Turnaround position. This will result in 32 mean values.

7. Perform a mathematical comparison of the mean values for each microwell in its Normal and Turnaround positions (A1/H12, A2/H11, B1/G12, B2/G11, and so on). In order to pass this test, the differences in the compared mean values must be within the accuracy specification for the instrument.


For example:

If the mean value for 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 its Turnaround (H12) position is 1.873 to 1.931 OD.

1.902 * 0.01 + 0.010 = 0.029; 1.902 – 0.029 = 1.873; 1.902 + 0.029 = 1.931

Note: If any set of mean values is out of the expected range, review the other three sets of mean values for the same channel pair. For example, if the A1/H12 comparison fails (the wells are not within the expected range of each other), review the comparisons of A2/H11, H1/A12, and H2/A11. If two or more sets of mean values for a channel pair are out of the expected range, there is a problem with one of the eight read channels. If only one of the four mean values results in a failure, check the well for debris and the plate for scratches or fingerprints.

Accuracy Specification:

For comparison in this test, the following accuracy specifications are applied, using Normal/Standard read mode and a 96-well microplate.

± 1.0% ± 0.010 OD from 0.000 to 2.000 OD

± 3.0% ± 0.010 OD from 2.000 OD to 3.000 OD


Liquid Test 2

Note: A 96-well, flat-bottom microplate is required for this test (Corning Costar #3590 is recommended). Use a new microplate; any fingerprints or scratches may cause variations in the turnaround reading.

Preparation of Dilutions:

1. Set up a rack containing 10 tubes, numbered consecutively.

2. Prepare a concentrated stock test solution insert using either Procedure A on page 3-16, or Procedure B on page 3-17

3. Create a percentage dilution series, beginning with 100% of the original concentrated stock solution in the first tube, 90% of the original solution in the second tube, 80% in the third tube, all the way to 10% in the last tube. Use Class A volumetric pipettes for better accuracy.

4. Dilute, using amounts of the remaining 0.05% solution of deionized water and Tween 20, as shown in Table 3-4 below.

Table 3-4 Test Tube Dilutions

Tube Number 1 2 3 4 5 6 7 8 9 10 Volume of original 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

Absorbance expected if original solution is 2.000 OD at 200 µl

2.0 1.8 1.6 1.4 1.2 1.0 0.8 0.6 0.4 0.2

Plate Preparation:

5. Pipette 200 µl of the concentrated solution from tube 1 into each well of the first column, A1 to H1, of a new flat-bottom microplate (Corning Costar® #3590 is recommended). Next, pipette 200 µl from each of the remaining tubes into the wells of the corresponding column of the microplate (tube 2 into wells A2 to H2, etc.).

Note: The choice of dilutions and the absorbance of the original solution can be varied. Use Table 3-4 as a model for calculating the expected absorbances of a series of dilutions, given a different absorbance of the original solution.



Linearity – Test A

1. Read the microplate prepared above using a normal mode dual wavelength at 450 nm with 630 nm as the blank. Repeat the read four times for a total of five reads.

2. The plate data can be exported to an Excel spreadsheet using Gen5™, KC4™, or KCjunior™. The mathematical computations described below may then be performed and the template kept for future data reduction.

3. Calculate the mean absorbance for each well, and average the means for each concentration.

4. Perform a regression analysis on the data to determine if there is adequate linearity.

For example:

In Microsoft Excel, under Tools, select Data Analysis and then Regression. (Prior to

opening the regression analysis tool, enter the expected results of the dilutions in a

row of cells to use in the analysis.) Use the Regression “Input” box to enter the

expected values as the “Input Y Range” and the mean absorbance for each

concentration as the “Input X Range.”

Note: If the Data Analysis command is not available on the Tools menu, you may need to install the Analysis ToolPak in Microsoft Excel. Consult Microsoft Excel Help for assistance.

Expected results:

Since it is somewhat difficult to achieve high pipetting accuracy when conducting linear dilutions, an R Square value of at least 0.99 is considered adequate.

Repeatability – Test B

1. The well that shows the most variation for each concentration is selected for data reduction. Calculate the mean and standard deviation for the five readings taken above for the selected wells.

2. For a mean below 2.000 OD, calculate the allowed deviation using the repeatability specification for a 96-well format of ± 1.0% ± 0.005 OD. If above 2.000 OD, apply the ± 3.0% ± 0.005 specification.


3. The standard deviation 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 1.0% (1.951 * 0.010) = 0.0195, which, when added to the 0.005

(0.0195 + 0.005) = 0.0245 OD, which is the allowable deviation. Since the standard

deviation is less than this value, the reader meets the test criteria.

Repeatability Specification:

± 1.0% ± 0.005 OD from 0.000 to 2.000 OD

± 3.0% ± 0.005 OD from 2.000 OD to 3.000 OD

Channel-to-Channel Variation and Alignment – Test C

1. Using the plate prepared for Test A above, conduct a turnaround test by reading the plate with the A1 well in the H12 position five times. This test results in four comparisons of each channel to its corresponding channel, two in column 1, and two in column 2.

2. Calculate the means of the wells in columns 1 and 2 in the normal plate position (data is from Test A) and in the turnaround position (from Step 1 above). Compare the mean reading for well A1 to its mean reading when in the H12 position.

3. Compare the mean values for the other wells to their corresponding mean values with the well in the turnaround position. (Compare B1 to G12, C1 to F12, D1 to E12, E1 to D12, F1 to C12, G1 to B12, H1 to A12, A2 to H11, and B2 to G11, etc.). 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% = 0.019 + 0.010 = 0.029, which is

added and subtracted from 1.902 for the range.)

If any set of well values is out of the expected range, review the other three sets for

the same channel pair. Thus, if A1 and H12 are not within range of each other, review

the compliance of H1 to A12, A2 to H11, and H2 to A11.


This will confirm that there is a problem in one of the eight read channels, or indicate

that the result of one set of wells was in error. If any two sets of well values for a

channel pair are out of the allowed accuracy range, there may be contamination on, or

a problem with, one of the lenses.

4. If the four corner wells are within the accuracy range, the reader is also in alignment.


Liquid Test 3

Note: A 96-well, flat-bottom microplate is required for this test (Corning Costar #3590 is

recommended). Use a new microplate; any fingerprints or scratches may cause variations in the

turnaround reading.

Required Materials:

• Deionized water

• Pipettes

• Corning Costar #3590 flat-bottom microplate

• Beakers and graduated cylinder

• Precision balance

• Tween 20 (polyoxyethylene (20) sorbitan monolaurate)

• Phosphate Buffered Saline with Tween 20 (PBS Buffer Solution). Use Sigma PBS tablets (#P4417, or equivalent), which will be adequate for one liter of PBS solution each (Procedure B) or prepare a 10x concentrate per Table 3-5 (Procedure A).

• -NADH Powder ( -Nicotinamide Adenine Dinucleotide, Reduced Form) Sigma bulk catalog number N 8129, or pre-weighed 10-mg vials, Sigma number 340-110

Stock Buffer Solution Formulation:

Table 3-5 Phosphate Buffered Saline 10X Concentrate Solution

KH2PO4 anhydrous 0.2 g

NaCl 8.0 g

Na2HPO4 anhydrous 1.15 g

KCl 0.2 g

Tween® 20 0.5 ml

Add deionized water to bring to 100 ml


Procedure A:

Mix 45 ml of deionized water with 5 ml of the concentrated PBS solution (from Table 3-5)

in a beaker. Add 10 mg of the -NADH powder and mix thoroughly. This solution will be

referred to as the high-level (or 100%) test solution.

Procedure B:

Add 50 ml of a PBS solution (prepared from the Sigma tablets) to a beaker. Add 10 mg of

the -NADH powder and mix thoroughly. This is the alternate high-level test solution.

Liquid Test 3 Procedure:

1. Check the absorbance of a sample of either high-level test solution created in

Procedure A or B at 340 nm on the microplate reader. This solution will have an

optical density (absorbance) of approximately 0.700 to 1.000 OD. This value is not

critical, but it should be within this absorbance range. If low, adjust up by adding

-NADH powder until the solution is at least at the lower end of this range. Do not

adjust if slightly high.

2. Carefully prepare a mid-level (75%) test solution by diluting 15 ml of the high-level

test solution with 5 ml of the Sigma PBS solution. (If using the 10X-concentrate PBS

solution, mix one part of the concentrate with nine parts of deionized water to obtain a

low-level buffer similar to the Sigma PBS. Then use 5 ml of this solution as the

diluent.)

3. Carefully prepare a low-level test (50%) test solution by diluting 10 ml of the

high-level test solution with 10 ml of the Sigma PBS solution. (If using the

10X-concentrate PBS solution, mix one part of the concentrate with nine parts of

deionized water to obtain a low-level buffer similar to the Sigma PBS. Then use 10 ml

of this solution as the diluent.)

4. Pipette the three solutions into the new, 96-well microplate:

150 µl of the high-level (100%) solution into all wells of columns 1 and 2

150 µl of the mid-level (75%) solution into all wells of columns 3 and 4

150 µl of the low-level (50%) solution into all wells of columns 5 and 6



5. Read the microplate five times using Normal mode, single wavelength at 340 nm, no

blanking (or blank on air).

The five sets of raw plate data can be exported to an Excel spreadsheet using Gen5™, KC4™ or KCjunior™. The mathematical computations described below may then be performed and the template kept for future data reduction.

Repeatability – Test A

1. For each well, calculate the mean and standard deviation of the five readings.

2. For each mean calculated in Step 1, calculate the allowed deviation using the repeatability specification for a 96-well format of ± 1.0% ± 0.005 OD (Mean * 0.01 + 0.005).

3. For each well, compare the standard deviation calculated in Step 1 with the allowed deviation calculated in Step 2. The standard deviation should be less than the allowed deviation.

For example:

Five readings in well A1 of 0.802, 0.802, 0.799, 0.798, and 0.801 will result in a

mean of 0.8004, and a standard deviation of 0.0018. The mean multiplied by 1.0%

(0.8004 * 0.010) = 0.008, which, when added to the 0.005 (0.008 + 0.005) = 0.013,

which is the allowable deviation for well A1. Since the standard deviation for well

A1 is less than 0.013, the reader meets the test criteria.

Linearity – Test B

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.

For example:

In a Microsoft Excel spreadsheet, enter the three mean values in ascending order and

label the column as the Y values. Enter 0.50, 0.75, and 1.00 and label the column as

the X values.


Select Tools > Data Analysis > Regression to open the Regression dialog. Set “Input

Y Range” to reference the above-mentioned Y values and set “Input X Range” to

reference the above-mentioned X values. Click OK to perform the analysis, the results

of which will be output in a separate sheet.

Note: If the Data Analysis command is not available on the Tools menu, you may need to install the Analysis ToolPak in Microsoft Excel. Consult Microsoft Excel Help for assistance.

Expected results:

Since it is somewhat difficult to achieve high pipetting accuracy when conducting linear dilutions, an R Square value of at least 0.99 is considered adequate.

Appendix A Decontamination and Cleaning

This appendix contains the procedures for decontaminating and cleaning the PowerWave™.

This appendix includes the following sections:

Purpose ................................................................................................................................................ A-2

Decontamination.................................................................................................................................. A-3

Tools and Supplies ....................................................................................................................... A-3

Procedure...................................................................................................................................... A-3

Cleaning............................................................................................................................................... A-4

Tools and Supplies ....................................................................................................................... A-4

Procedure...................................................................................................................................... A-4

A-2 Appendix A - Decontamination and Cleaning

Purpose

Any laboratory instrument that has been used for research or clinical analysis is considered a biohazard and requires decontamination prior to handling. Intact skin is generally considered an effective barrier against infectious organisms; however, small abrasions and cuts may not always be visible.

Decontamination minimizes the risk to all who come in contact with the instrument during shipping, handling, and servicing. Decontamination is required by the U.S. Department of Transportation regulations.

Persons performing the decontamination process must be familiar with the basic setup and operation of the instrument.

iii BioTek Instruments, Inc. recommends the use of the following decontamination solutions and methods based on our knowledge of the instrument and 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 gloved hands away from eyes, mouth, and nose. Eating or drinking while decontaminating instruments is not advised.

!

Mucous membranes are considered prime entry routes for infectious agents. Wear eye protection and a surgical mask when there is a possibility of aerosol contamination. Intact skin is generally considered an effective barrier against infectious organisms; however, small abrasions and cuts may not always be visible. Wear protective gloves when performing the decontamination procedure.

PowerWave Operator’s Manual A-3

iii Important! Turn off and unplug the reader for all decontamination or cleaning operations.

iii Important! Do not immerse the instrument, spray with liquids, or use a “wet” cloth. Do not allow liquid to run into the interior of the instrument. If this happens, contact the BioTek Service Department.

Decontamination

Tools and Supplies

• Sodium hypochlorite (NaClO, or bleach)

• Deionized or distilled water

• Clean, lint-free cotton cloths

• Protective gloves

• Biohazard trash bags

• Lab coat

• Safety glasses

• Surgical mask

• 125 ml beakers

Procedure

!

Warning! The bleach solution is caustic; wear gloves and eye protection when handling the solution.

1. Turn on the PowerWave™ and press the carrier eject button to eject the carrier.

2. Turn off and unplug the instrument from the power supply.

3. Prepare an aqueous solution of 0.5% sodium hypochlorite (NaClO, or bleach).

Be sure to check the % NaClO of the bleach you are using; this information is printed on the side of the bottle. Commercial bleach is typically 10% NaClO; if this is the case, prepare a 1:20 dilution. Household bleach is typically 5% NaClO; if this is the case, prepare a 1:10 dilution.

A-4 Appendix A - Decontamination and Cleaning

4. Moisten a clean, lint-free cloth with the bleach solution. Do not soak the cloth.

5. Wipe the plate carrier and all exposed surfaces of the instrument.

6. Allow the instrument to dry for 20 minutes for thorough decontamination by the bleach.

7. Moisten a cloth with deionized or distilled water and wipe all surfaces of the instrument that have been cleaned with the bleach solution.

8. Use a clean, dry lint-free cloth to dry all wet surfaces.

9. Discard the used gloves and cloths, using a Biohazard trash bag and an approved Biohazard container.

Cleaning

Tools and Supplies

• Mild detergent

• Deionized or distilled water

• Clean, lint-free cotton cloths

Procedure

1. Turn on the PowerWave™ and press the carrier eject button to eject the carrier.

2. Turn off and unplug the reader from the power supply.

3. Moisten a clean, lint-free cloth with water, or with water and the mild detergent. Do not soak the cloth.

4. Wipe the plate carrier and all exposed surfaces of the instrument.

5. If detergent was used, wipe all surfaces with a cloth moistened with water.

6. Use a clean, dry lint-free cloth to dry all wet surfaces.

Appendix B Computer Control

The PowerWave™ may only be controlled by host software installed on a PC connected to the reader via the

computer’s serial port. This appendix describes the features of computer control and how to configure the

computer to control the reader.


Overview ............................................................................................................................................. B-2

Controlling the Reader with Gen5....................................................................................................... B-3

Setting Up Gen5 ........................................................................................................................... B-3

Problems....................................................................................................................................... B-4

Getting Started with Gen5 ............................................................................................................ B-4

Controlling the Reader with KC4 ........................................................................................................ B-6

Setting Up KC4 ............................................................................................................................ B-6

Problems....................................................................................................................................... B-7

Getting Started with KC4 ............................................................................................................. B-7

KC4’s OLE functions ................................................................................................................... B-8

Controlling the Reader with KCjunior ................................................................................................ B-9

Setting Up KCjunior..................................................................................................................... B-9

Problems..................................................................................................................................... B-10

Getting Started with KCjunior.................................................................................................... B-10

Controlling the Reader Using ActiveX.............................................................................................. B-11

Controlling the Reader Using Serial Protocol ................................................................................... B-11

B-2 Appendix B – Computer Control

Overview

The PowerWave™ is a completely computer controlled instrument. Gen5™ is the primary operating software for the PowerWave, however, BioTek’s KC4™ or KCjunior™ software may be used instead of Gen5.

For users requiring a custom interface to the PowerWave, there are other methods of computer controlling the reader. A typical example of this requirement is the need to integrate the PowerWave into an automated system.

This appendix describes all the options available to the user for computer control of the PowerWave and includes:

• Controlling the Reader with Gen5, page B-3

• Controlling the Reader with KC4, page B-6

• Controlling the Reader with KCjunior, page B-9

• Controlling the Reader Using ActiveX, page B-11

• Controlling the Reader Using Serial Protocol, page B-11

Note: If you are operating the PowerWave with the Bio-Stack™ Microplate Stacker, you will need a PC equipped with two serial ports, and any version of Gen5 or the appropriate version of KC4 software installed on the PC. Refer to Chapter 3, Installation, in the Bio-Stack Operator’s Manual for instructions.

PowerWave Operator’s Manual B-3

Controlling the Reader with Gen5

iii Before installing Gen5™, verify that your computer meets the minimum system requirements specified in Gen5’s Help system or Getting Started Guide.

Setting Up Gen5

The PowerWave™ can be operated via BioTek’s Gen5 software installed on a host computer. The following instructions briefly show you how to set up Gen5 for operation of the reader. Refer to Gen5’s Getting Started Guide or Help system for more detailed instructions.

1. Turn off the computer and the reader. Connect the appropriate serial cable

(PN 75053) between the two machines. (See Table B-1 on page B-11 for a pin-out

description of the cable.)

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. Open Gen5.

5. Login if prompted (Gen5™ Secure). The default Administrator password is admin.

6. When the ‘Welcome to Gen5’ screen appears, select System Menu.

7. From Gen5’s main screen, select System > Reader Configuration to open the

‘Reader Configuration’ dialog.

8. Click the Add button to open the ‘Reader Settings’ dialog.

• Gen5 and Gen5 Secure: Up to two readers may be added in Gen5.

9. Use the drop-down list in Reader Type to select Powerwave.

10. Enter the appropriate Com Port.

11. The Baud Rate is set at 9600 and cannot be changed.

12. Click Test Comm. Gen5 will attempt to communicate with the reader.

• If you receive “The Reader is communicating!” message, click OK, and then

click OK again to save the settings. Click Close at the Reader Configuration

dialog to return to the main screen.


• If the test is not successful and you receive an error message, refer to the

Problems section below or to the Troubleshooting section of Gen5’s Help

system for assistance.

• Gen5™ Secure only: An ‘Audit Trail’ dialog will appear after exiting Reader

Configuration, whenever you add, modify, or delete a reader. If desired, enter

any comments, then click Close.

Problems

If Gen5™ fails to communicate with the reader, try the following troubleshooting suggestions:

• Confirm that the correct Reader Type 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 Troubleshooting section in Gen5’s Help system for further

assistance.

Getting Started with Gen5

The following instructions briefly show you how to define and perform a “Quick Read” in Gen5

(File > New Experiment > Default Protocol). It’s called “Quick” because you can perform a

reading without having to take the time to create a new protocol.

If the reading is part of an experiment or assay that you will perform numerous times, you will need

to create a new protocol (File > New Protocol).

Refer to Gen5’s Help system early and often, to learn how to create protocols, assign well

identifiers, read plates, print reports, perform data reduction, and more.

To perform a “Quick Read”:

1. At Gen5’s Welcome screen, select System Menu, then at the main screen, select File

> New Experiment. (Alternative: select Read a Plate at the Welcome screen, then

proceed to step 4 on the following page.)

2. Click Default Protocol, then click OK. Gen5 will open the Experiment workspace,

which includes the Protocol menu tree and Plate screen.


3. Select Plate > Read or click the ‘Read Plate’ icon. The ‘Procedure’ dialog will open.

• Gen5™ and Gen5™ Secure: If more than one reader was added in Gen5, the

‘Instrument Selection’ dialog will appear instead of the Procedure dialog.

Select PowerWave, then click OK. The Procedure dialog will then appear.

4. Select a Plate Type.

5. Click Read to open the ‘Read Step’ dialog.

6. Select a Read Type.

7. Define the wavelength(s) at which the plate will be read.

8. Define other reading 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. Click OK again to save and close the Procedure dialog. The ‘Plate Reading’ dialog

will open.

11. Enter any desired information, place the plate on the carrier, 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 drop-down arrow in the Plate

screen to select one wavelength. The results will be displayed for the selected

wavelength. Repeat, for other wavelengths.

• To analyze, manipulate, or print results, Protocol parameters should be

defined. Refer to Gen5’s Help system for instructions.

Note: Gen5™ Reader Control does not support data reduction.


Controlling the Reader with KC4

iii Before installing KC4™, verify that your computer meets the minimum system requirements specified in KC4’s User’s Guide or Help system.

Setting Up KC4

The PowerWave™ can be operated via BioTek’s KC4 software installed on a host computer. The following instructions briefly show you how to set up KC4 for operation of the reader. Refer to KC4’s User’s Guide or Help system for more detailed instructions.

1. Turn off the computer and the reader. Connect the appropriate serial cable (PN 75053) between the two machines. (See Table B-1 on page B-11 for a pin-out description of the cable.)


3. Install KC4 on the computer’s hard drive and register the software with BioTek.

4. Once installed, open KC4. Log in, if prompted.

5. Select System > Readers to open the ‘Reader Selection’ dialog.

6. Scroll through the list of Available Readers and select the appropriate PowerWave model.

7. Click the Port button (and subsequent Setup button) to define the following communications parameters:

• Port: COMn (select the COM (serial) port used for the RS-232 cable

connection).

• Transmission Speed: 9600

• Data Bits: 8

• Parity: No

• Stop Bits: 2

8. Click OK to close the ‘Port Setup’ dialog, then click OK to close the ‘Port Selection’ dialog.

9. Click the Current Reader button to attempt to establish communication with the reader, using the currently defined communication parameters.


• If the test passes, click Close to save the settings and close the dialog.

• If the test fails, KC4 will provide appropriate instructions for resolving any

problems. See also the Problems section below.

Problems

If KC4™ fails to communicate with the PowerWave™ and it displays a serial communications

error, check the cable plug-in location to ensure that it matches the setup choices and is not a Null

cable. If this is suspected, add another Null and try again.

If an ‘Incorrect Reader Model Connected’ dialog is displayed, click OK to clear the screen and

select System > Readers > Available Readers. Verify that the reader selected is correct.

Getting Started with KC4

The following instructions briefly describe how to read a plate using KC4. Refer to KC4’s Help

system and the 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 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 (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. Select the appropriate Plate Type.

7. Define other reading parameters as necessary. Click the Help button for assistance.

8. When complete, click OK.

9. Select Data > Read Plate. The ‘Plate Reading’ dialog will appear.


10. Enter any comments, place the plate on the carrier, then click START READING to

begin the plate read.

• The plate will be read and then the raw data results will display in KC4.

• To analyze, manipulate, or print results, Protocol parameters should be

defined. Refer to the KC4 User’s Guide or Help system for instructions.

KC4’s OLE functions

KC4™ can be used as an OLE server to control the reader. This means that KC4 provides an OLE

interface allowing other applications (the OLE clients) to use most of its features.

For example, the client applications can use KC4 to:

• Perform a read

• Perform data reduction

• Export results

• Print a report

KC4 also provides basic reader commands, such as “open door,” “close door,” etc.

Because of its OLE automation capabilities, KC4 can be interfaced with any application written in

C++, Visual Basic, or any application offering a VBA (Visual Basic for Applications) syntax, such

as Excel. The user should refer to the Microsoft® OLE/ActiveX documentation and must have a

good knowledge of OLE mechanisms. Contact BioTek for more information.


Controlling the Reader with KCjunior

iii Before installing KCjunior™, verify that your computer meets the minimum system requirements specified in KCjunior’s User’s Guide or Help system.

Setting Up KCjunior

The PowerWave™ can be operated via BioTek’s KCjunior software installed on a host computer. The following instructions briefly show you how to set up KCjunior for operation of the reader. Refer to KCjunior’s User’s Guide or Help system for more detailed instructions.

1. Turn off the computer and the reader. Connect the appropriate serial cable (PN 75053) between the two machines. (See Table B-1 on page B-11 for a pin-out description of the cable.)


3. Install KCjunior on the computer’s hard drive and register the software with BioTek.

4. Once installed, open KCjunior.

5. Select Setup > Reader 1 or Reader 2 to open the ‘Reader Setup’ dialog.

6. Scroll through the Reader Type list and select the appropriate PowerWave model. Define the communications parameters as follows:

• Com Port: COMn (select the COM (serial) port used for the RS-232 cable

connection).

• Baud Rate: 9600

• Data Bits: 8

• Parity: None

• Stop Bits: 2

• EOT Character: Keep the default number.

7. Click the Test Communications button to attempt to establish communication with the reader, using the currently defined communication parameters. If a ‘Serial Write Error’ dialog is displayed, an incorrect COM port may have been selected. Select a different port and then repeat this step.

• If the test passes, at ‘The communications test was successful’ dialog, click

OK, then click OK again to save the settings and close the Reader Setup

dialog.


• If the test fails, follow the directions provided by KCjunior, then click Test

Communications again. See the Problems section below.

Problems

If KCjunior™ fails to communicate with the reader and displays a serial communications error,

check the cable plug-in location to make sure it matches the setup choices in Table B-1 on the

following page and is not a Null cable. If this is suspected, add another Null and try again.

Getting Started with KCjunior

The following instructions briefly describe how to read a plate using KCjunior. Refer to KCjunior’s

Help system and the 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.

The ‘Protocol Definition’ dialog will appear.

3. Select a Read Method Type.

4. Define the Wavelength(s) at which the plate will be read.

5. Select the appropriate Plate Geometry.

6. Define other reading parameters as necessary. Click the Help button for assistance.

7. When complete, click OK to return to the Read Plate dialog. If desired, enter a Plate

ID.

8. Place the plate on the carrier, and then click OK to start the plate read.

• The plate will be read and then the raw data results will display in KCjunior.

Print the raw data by selecting Results > Print Results.

• To analyze or manipulate results, a Protocol should be defined. Refer to

KCjunior's Help system or User's Guide for instructions.


Controlling the Reader Using ActiveX

The BioTek reader ActiveX® component allows an experienced programmer to create a container

application to control all reading parameters and obtain optical density values from the reader. For

more information about ActiveX, contact your BioTek dealer.

Controlling the Reader Using Serial Protocol

If you wish to serially control a PowerWave™ Reader, the serial communication protocol has been

documented in BioTek specification 7266201-SP, which is available free of charge. Contact your

BioTek dealer and ask for this part number.

Table B-1

Serial Cable Pin-Out Description

Serial Cable Pin-Out*

PC (9-pin female) Reader (25-pin female)

1+6 4

2 2

3 3

5 7

7 5+6+8

8 20

Shell Shell (shielding)

* For a 25-pin PC connection using BioTek serial cable (PN 75053) plus 9pM-25pF adapter (PN 49755).

Appendix C Error Codes

This chapter describes Error Codes that may appear on the controlling PC using Gen5™, KC4™, or

KCjunior™ software. If an error is displayed, call BioTek’s Technical Assistance Center. See Chapter 1,

Product Support & Service for contact information.


Error Codes in Gen5, KC4, or KCjunior ............................................................................................. C-2

Error Codes During Operation of the Reader with the Bio-Stack ....................................................... C-2

Diagnostics .......................................................................................................................................... C-2

General Errors...................................................................................................................................... C-3

Fatal Errors ........................................................................................................................................ C-11

C-2 Appendix C – Error Codes

Error Codes in Gen5, KC4, or KCjunior

An error code is displayed in Gen5™, KC4™, or KCjunior™ software as a four-digit identifier. The first digit will be 0, 1, 2, 3, or A.

• 0, 1, 2, or 3 denote a noncritical error, which means that it is still possible for the PowerWave™ to communicate with the controlling software and run the reader system test. See General Errors on pages C-3 through C-10.

• A denotes a more serious error with the memory or processing, which requires that the PowerWave be turned off and then powered up before any diagnostics can be performed. If the error reappears, call BioTek TAC for troubleshooting assistance (see Chapter 1). See Fatal Errors on page C-11.

Error Codes During Operation of the Reader with the Bio-Stack

Error codes may be displayed in Gen5 or KC4 software during operation of the PowerWave with the Bio-Stack™ Microplate Stacker. Error codes displayed in KC4 during operation with the Bio-Stack that are prefixed with a B-, indicate either errors generated by the Bio-Stack or problems with communication between the Bio-Stack and the PowerWave. Errors detected by Gen5 following calls to Gen5 functions by the Bio-Stack™ PC Control Software are displayed in a negative value format, for example: -8, -100, -2010. Refer to the Troubleshooting and Error Codes section of your Bio-Stack Operator’s Manual for a list of these types of error codes and their descriptions.

Diagnostics

If any error code is displayed in Gen5, KC4, or KCjunior, a System Self-Test should be conducted for diagnostic purposes. The following are brief instructions for performing the Self Test:

• To run a System Test from Gen5, select System > Diagnostics > Run System Test. If the Instrument Selection dialog appears (Gen5 and Gen5 Secure), select Powerwave, then click OK.

• To run a System Test from KC4, select System > Diagnostics > Run Optics Test. Enter the Reader Serial Number, then click Start.

• To run a System Test from KCjunior, select Utilities > Diagnostics > Reader System

Test. Enter the Reader Serial Number, then click Run Test.

Refer to Gen5’s Help system or to the KC4 and KCjunior User’s Guides for more detailed instructions.

PowerWave Operator’s Manual C-3

General Errors

Code Description and Probable Causes

0200

X-axis opto sensor failed to transition.

This error indicates that a motor was not able to move to its “home” position, as registered by feedback from an optical sensor, or it failed to transition after moving away from the home position.

Probable causes:

• Dirty x-axis rail or dry bearings are causing too much friction.

• Defective or broken optical sensor.

• Defective motor controller PCB.

0201 Order sorting (bandpass) filter wheel did not home.

Probable causes:

• Filter wheel is loose.

• Filter wheel is obstructed by too close proximity to the motor gear.


• Defective motor, motor controller PCB, or cable.

0202 Y-axis opto sensor failed to transition.

This error indicates that a motor was not able to move to its “home” position, as registered by feedback from an optical sensor, or it failed to transition after moving away from the home position.

Probable causes:

• Dirty y-axis rail or dry bearings are causing too much friction.


• Defective motor controller PCB.

Note: In cases where a sensor is not functioning, the motor will drive the axis to its mechanical stop and generate substantial noise.


General Errors (Cont’d)


0303 Monochromator did not find home.

During the instrument initialization, the monochromator is homed by rotating the monochromator mirror until the white light (full light) is detected. This requires a fully functional flash lamp/detection system.

Probable causes:

• Defective analog PCB.

• Defective flash lamp and or flash lamp power supply (inconsistent flashes) (high probability).

• Defective motor/power PCB.

• Defective monochromator (low probability).

0400

Carrier x-axis failed positional verify.

Motor x-axis failed to reach the same position when moved a known number of steps from the home position and back.

Probable causes:

• Dirty rail or dry bearings are causing too much friction.

0401

Order sorting (bandpass) filter wheel failed positional verify.

• Filter wheel obstructed by motor gear.

• Motor gear loose on motor shaft.

0402

Carrier y-axis failed positional verify.

• Dirty rail or dry bearings are causing too much friction.

0403

Monochromator failed to find the zero order position (white light).

The order sorting (bandpass) filter wheel is homed and moved to the open hole position. The monochromator is moved until the optical system detects saturation (home). It is then moved to a known number of steps away from home and then moved back the same number of steps, expecting to see light saturation point. The error is indicating the saturation did not clear or appear.

Probable causes:

• Flash lamp is missing flashes or is not flashing.

• The optic system does not detect the saturation.

• Defective monochromator.




0500 Measurement or reference channel is saturated during a spectral scan.

This error indicates the light signal level in one of the channels reached 65,535 counts during Lambda calibration within the spectral scan.

0501 - 0508 Measurement or reference channel is saturated during a spectral scan.

This error indicates the light signal level in the channel indicated by the last digit (1 through 8) of the error code reached 65,535 counts during the spectral scan or calibration.

Probable causes:

• The lamp is not properly aligned and there is too much light.

• The A/D reference voltage is not at the 4.5 V.

• The analog PCB is defective.

• Order sorting filter has degraded.

0503 Monochromator failed positional verify due to saturation.

This error indicates that, during initialization, the monochromator failed positional verify, or channel 3 failed during calibration or spectral scan.

0511 – 0568 Measurement channel A/D signal saturated.

This error indicates the light signal level reached 65,535 counts for one of the lambda values in the table after calibration, prior to a read, or during a read or optics test.

0600 Gain out of range for the target air readings. Reference channel = hot channel.

This error indicates that the measurement channel signal gain is out of range necessary to ensure the reader’s performance to specifications. During reader calibration, the gain selected is 36.56.

Probable causes:

• Flash lamp

• Monochromator

• Lamp power supply




0701 - 0708

Channel failed noise test greater than 20 counts during optics test.

This error indicates significant variations in background electronic noise were detected, when blocking the light and increasing the gain to maximum.

Probable causes:

• Electrical noise may be penetrating the measurement chamber. The bottom and top shrouds are part of the electrical shielding.

• The coaxial cable ground may be floating or disconnected.

• There may be an ambient light leak. Ensure that the plate carrier door is properly closed.

• Analog PCB failure; noisy photo-detector.

• Internal electronic noise may be caused by a faulty analog PCB or faulty internal grounding.

0801- 0808

Channel failed noise offset < 10 and > 2000 during optics test.

This error indicates that background electronic signal detected is outside of acceptable limits at maximum gain when blocking the light.

Probable causes:

• The photo-detector is not connected or is defective, yielding a noise reading of zero.

• The photo-detector is too noisy and is defective.

0901 - 0908 Channel dark range is < 100 or > 20000 during calibration, or < 100 during a filter test.

0911 - 0968 Measurement channel dark range is < 100 during a read in enhanced mode (64 flashes), or prior to a read or optics test.

The reference channel dark current value has changed since the last optics test measurement by more than 10%, or the dark value is less than 100. The last number in the error code is the channel number used during the failure.

Probable causes:

• The photo-detector is more sensitive to temperature changes.

• Ambient light leakage during the read.




0A00 - 0A68 Measurement channel air / blank out of range prior to a read.

This error is indicating the air reading at the time of the plate read was 50% less than the air reading at the time of the optic test. The last number of the error code represents the channel at the time of failure.

0A10 Reference channel air / blank out of range for the first wavelength in a scan, or filter / reference channel air / blank out of range prior to a read.

This error is indicating the air reading at the time of the plate read was 50% less than the air reading at the time of the optic test. The last number of the error code represents the channel at the time of failure.

Probable causes:

• Flash lamp has missed flashes during the read.

• Dirty optics or spilled substance on the optics.

0F00 – 0F08 Channel Delta out of range during calibration.

The Delta of the air / dark is out of range during the calibration at a wavelength reference channel < 500 or measurement channel < 8000.

0F10 Reference channel out of range during a spectral scan. Reference channel < 500 during a spectral scan and only checking the first wavelength.

0F10 - 0F60 Reference channel out of range during a read. Reference channel < 500 during a read.

This error indicates that the reading has failed. The last number of the error code represents the channel.

0F10 - 0F68 Channel out of range prior to a read. Reference channel < 500 or measurement channel < 8000 during an optics test or prior to a read. Reference channel out of range (< 50% or > 200%).

Probable causes:

• The flash lamp may be out of alignment.

• The order sorting (bandpass) filter is degraded, and does not allow enough light energy to pass through.

• Damaged reference channel optic spray.

• The reference channel photodiode detection circuit is defective.




1100 or 1101 Failed configuration checksum test for reader protocol or system configuration prior to a read or optics test. Last digit can be either a 0 or 1.

Probable causes:

• The flash memory on the PCB is defective or corrupt. The basecode software and/or assays may need to be re-downloaded.

1200 Lambda calibration data missing prior to a spectral scan, or autocalibration data is missing.

The instrument calibration values are not loaded into the flash memory.

Probable causes:

• The PCB was changed and the flash memory does not have the calibration values loaded.

• Failure in Main PCB memory. Contact BioTek TAC for more information.

1201 – 1206 Lambda table calibration data missing from reader.

This error occurs when the controlling PC requests the Lambda wavelength calibration data, and one of the wavelengths does not have calibration data in memory (not calibrated). The last digit represents the Lambda.

1300 Carrier not homed in the x-axis.

This error is only seen if an error 0200 is ignored. See the probable causes for 0200.

1301 Order sorting (bandpass) filter wheel not homed.


1302 Carrier not homed in the y-axis.


1501 – 1504

Temperature zone out of range (the last digit is the zone number failing).

1511 - 1514

Thermistor failed – resistance out of range (the last digit is the zone number).

1520 A/D converter failed; incubator PCB defective.




1600 Computer control assay definition error.

This error will occur for the following definitions:

• Well set

• Wave scan

• Checksum at the protocol sent from computer

• Plate geometry

• Filter(s)

• Features available

• Mono

1700 Kinetic interval too short for selected options, or kinetic interval = 0.

This error indicates that the kinetic interval in the current assay is too short.

Increase the kinetic interval.

1900 Memory allocation failed.

This error is typically used only for software development purposes. If it occurs, however, try turning the instrument off and then on again after a wait of 30 seconds. If the error persists, contact BioTek Technical Support.

1C00

A/D calibration standby signal on the analog board never went low, or A/D calibration standby line went low but never transitioned to a high.

This error indicates there is a failure with the absorbance analog board initialization, or the cable to the PCB is defective.

2000 Barcode scanner did not see 10 characters from barcode.

Probable causes:

• Barcode positioned incorrectly on plate.

• Insufficient carbon black in barcode label.

• Barcode label not in Code 39 format.




2100 Invalid parameter value selected.

This error can occur only during computer control, indicating that one of the following invalid assay configurations was sent to the instrument:

• Temperature out of range

• Wavelength not in ASCII format

• Incorrect plate geometry

• Incorrect row range or column range selected

• Kinetic interval = 0

• Incorrect range or order selected for start wavelength / end wavelength

• Incorrect well selected for scanning

2400 Middle sensor position incorrect.

This error occurs when homing to the middle sensor and the optical flag is in a different position since the last autocalibration was performed.

2500 or 2502 Sweep mode read missed well location; last digit is the motor number.

2800 - 2803 Motor currently in use; last digit indicates motor.

2F00 Results data being is sent not acknowledged by host PC.

This error indicates the handshaking between the host PC software and the reader did not complete. This is a lost data condition.

3200 - 3201 Never saw A/D ready transition.

This error indicates there is a failure with the absorbance analog board or the cable to the PCB is defective.


Fatal Errors

Fatal errors indicate conditions that require immediate attention. If a fatal error is displayed, contact BioTek’s Technical Assistance Center for further instructions (refer to the Product Support & Service section in Chapter 1).

Code Description

A100 Task control block not available.

A200 Read already in progress.

A300 Motors not available.

A301 Real time clock not available

A302 Display not available

A303 Flash not available

A400 Failed code checksum test on power-up.

A600 Data flash write timed out.

A700 Data flash readback did not match write.

A800 Code flash write timed out.

A900 Memory allocation heap corrupted.

Appendix D Instrument Dimensions

This appendix contains information concerning the instrument dimensions, and location of the microplate

carrier and mounting holes.


Instrument Dimensions for Robotic Interface ..................................................................................... D-2

D-2 Appendix D - Instrument Dimensions

Instrument Dimensions for Robotic Interface

Figure D-1 on the following page shows the location of the microplate carrier in reference to the exterior surfaces of the PowerWave™, and the mounting holes on the bottom. The illustration should facilitate system setup with a robotic unit, such as the Bio-Stack™ Microplate Stacker.

If you have purchased the Bio-Stack to operate with the PowerWave, special alignment hardware is included in the Bio-Stack accessory package for correct positioning of the PowerWave with the Bio-Stack. Refer to Chapter 3, Installation, in the Bio-Stack Operator’s Manual for instructions for alignment of the PowerWave with the Bio-Stack.

PowerWave Operator’s Manual D-3

Figure D-1: Microplate carrier position, mounting holes, and instrument dimensions

D-4 Appendix D - Instrument Dimensions

Appendix E Barcode Scanner

This appendix contains instructions for enabling the barcode scanner option in KC4™ or KCjunior™ (for

older models of the reader that are equipped with the scanner), and specifications for barcode label format

and position on the plate.


Internal Plate Barcode Scanner............................................................................................................ E-2

Enabling Barcode Scanning in KC4............................................................................................. E-2

Enabling Barcode Scanning in KCjunior ..................................................................................... E-3

Specifications for Barcode Labels ................................................................................................ E-3

Positioning the Labels on the Plates ............................................................................................. E-6

E-2 Appendix E - Barcode Scanner

Internal Plate Barcode Scanner

The barcode scanner is located inside the carrier compartment of the PowerWave™, on the left side, for models equipped with the scanner. As the carrier retracts inside the instrument for a plate read, the scanner is activated just long enough to scan the barcode label that has been attached to the plate, and is then deactivated before the plate is read.

The barcode scanner must first be enabled by selecting the scanner options in KC4™ or KCjunior™, as described below and on the following page.

Barcode labels that are attached to the microplates must meet certain requirements for format and quality and must be properly placed upon the microplate in order to read by the scanner. See page E-3 for label specifications, and pages E-4 through E-6 for illustrations of sample labels and proper location of labels on the microplates.

Enabling Barcode Scanning in KC4

To enable the barcode scanner option in KC4, follow the instructions below:

1. Select Protocol > Reading. At the ‘Reading’ dialog, check the Read Barcode option. (Refer to the KC4 sections in Appendix B, Computer Control, or to the KC4 User’s Guide for instructions on defining the other reading parameters, such as Reading Type, Wavelengths, etc.)

Note: If for any reason the barcode cannot be read, KC4 will prompt the operator to enter the barcode manually.

2. Click OK.

Once barcode scanning has been enabled in KC4, the barcode may be used as a file name. To select this option:

1. Select Protocol > Options > Raw Data.

2. Click on Automatic Save after reading, then Use barcode as filename. Click on the Help feature or refer to the KC4 User’s Guide for instructions on defining the other Save parameters such as the Save Directory, etc.

3. Click OK.

PowerWave Operator’s Manual E-3

Enabling Barcode Scanning in KCjunior

To enable the barcode scanner option in KCjunior™, follow the instructions below:

1. Select Setup > Configuration. At the ‘KCjunior Configuration’ dialog, click the General tab.

2. Click Use reader’s barcode scanner for Plate ID (if supported by reader). (Refer to the Help feature or the KCjunior User’s Guide for instructions on defining other parameters in the General, Well Ids, Reporting Options, or Pathnames tabs in the KCjunior Configuration dialog.)

3. Click OK.

4. If the read is unsuccessful, KCjunior will ask you to retry or enter the plate ID manually.

5. If the barcode returned from the reader is preceded by an unnecessary identification character, and you wish to have KCjunior remove it, select Setup > Configuration >

General, click Remove first character of barcode (typically defines the barcode

standard).

Specifications for Barcode Labels

Barcode labels are available for purchase from BioTek, or they may be created using barcode software and label products that meet the following specifications:

• Code 39 is the recommended format (“symbology”) for the labels.

• Industry standards: The labels should be made in accordance with Automation Identification Manufactures (AIM) uniform symbol specification for all codes supported. Label decodability should meet ANSI Specification X3, 182-199 “Bar-Code Print Quality Guideline” for a rating of A/05/880.

• The labels should be high-resolution, printed on good-quality copier machines or laserjets that have adequate amounts of carbon black in their toner. The scanner operates in the infrared range, and print that contains high levels of carbon black is readily detected in that range.

BioTek offers pre-printed, self-adhesive barcode labels (PN 7121094), in rolls containing 500 labels, that meet the above specifications for format, industry standards, and quality. Each label features a perforated section that may be easily attached to the side of a standard 96- or 384-well microplate. Figure E-1 on the following page is an actual-size reproduction of one of the BioTek labels and its perforated section.

Figure E-2 on page E-5 contains barcode label artwork that may be submitted to a print vendor for creation of labels that meet the code symbology and industry standards.


Figure E-1a: Sample of one BioTek barcode label from roll (PN 7121094)

nnnnnnnnnn

nnnnn

nnnnn

nnnnn

Figure E-1b: Section of label for attachment to microplate

PowerWave Operator’s Manual E-5

.05

1.12

2.28

.20(DIECUT,DO NOTCUTRELEASELINER)

.34 .11 GUARD BAR 2X.10 MINBLACK

.30 QUIET ZONE 2X

.20 MINWHITE

.75

.05

.62 (DIECUT)

.93

nnnn

nnn

nnn

nnnn

n

nnnn

n

Figure E-2: Barcode label artwork with recommended dimensions


Positioning the Labels on the Plates

The purchased or custom-made barcode label must be properly attached to a position on the side of the microplate opposite well A1, as illustrated below in Figure E-3 below.

1. Use a microplate with flat, vertical sidewalls so that the label will lie flat upon the surface.

2. Place the label on the area of the microplate specified below, and ensure that the bottom edge of the barcode (not the bottom edge of the label) is positioned approximately 7.0 mm up from the bottom of the plate.

3. Refer to Appendix D, Instrument Dimensions for correct orientation of well A1 to the reader when placing the microplate in the carrier.

Figure E-3: Position of barcode label (opposite well A1) on the microplate

Well A1 position

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