Agilent 2100 Bioanalyzer 2100 Expert User’s Guide Edition May 2005

Notices

Manual Part number: G2946-90004

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Edition: May 2005

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Contents About this Manual ... 5 In this Manual... 6 Related Documents ... 8 How to Use this Manual ... 10 Quick Start ... 14 Looking at 2100 Expert ... 28 Introduction to the Key Features of the 2100 expert... 29 Starting 2100 Expert ... 31 2100 Expert Work Area ... 32 Closing 2100 Expert ... 43 Running and Evaluating Electrophoretic Assays ... 45 Principles of Nucleic Acid and Protein Analysis on a Chip... 46 Preparing and Running an Electrophoretic Assay... 50 Analyzing and Evaluating the Results of an Electrophoretic Assay ... 81 Result Flagging ... 152 Running and Evaluating Flow Cytometric Assays... 166 Principles of Flow Cytometric Measurements ... 167 Overview of Flow Cytometric Assays ... 176

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Preparing and Running a Flow Cytometric Assay ... 179 Analyzing and Evaluating the Results of a Flow Cytometric Assay... 211 Working with Chip Data and Assays... 258 2100 Expert Data Overview ... 259 Handling Assays... 262 Handling Chip Data ... 267 Organizing, Backing up, and Archiving 2100 Expert Data ... 269 Importing Data... 271 Exporting Data ... 277 Printing Reports ... 286 Configuring Tables... 293 Administering System Functions ... 299 Configuring 2100 expert... 300 Using Log Books ... 310 Running Instrument Diagnostics ... 316 How to Run Instrument Diagnostics Tests... 321 Performing Verifications ... 325 Products, Spare Parts, and Accessories... 334 Glossary ... 338

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About this Manual

Welcome to the User’s Guide for the Agilent 2100 expert software. This manual provides beginners and advanced users with information needed to successfully run electrophoretic and flow cytometric assays with the bioanalyzer. The 2100 expert software allows the control of the bioanalyzer (including diagnostic functions) and, in combination with a LabChip kit, the acquisition, interpretation and result presentation of data generated during the analysis of DNA, RNA, proteins, and cells.

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In this Manual This manual provides bioanalyzer users with the following information: • “About this Manual” on page 5 gives an overview of the subjects in this manual, and lists major innovations and improvements of the 2100 expert software. It also lists supplemental literature and shows you how to make efficient use of this manual. • “Quick Start” on page 14 is meant for experienced users. It briefly summarizes the necessary steps to prepare and run an assay. • “Looking at 2100 Expert” on page 28 shows how to get started with the 2100 expert software, and outlines its main operational possibilities. • “Running and Evaluating Electrophoretic Assays” on page 45 explains how electrophoretic measurements are made using the bioanalyzer, gives detailed descriptions of all steps necessary to run electrophoretic assays, and shows how to analyze and evaluate results using electropherograms and gel-like images. • “Running and Evaluating Flow Cytometric Assays” on page 166 explains how flow cytometric measurements are made using the bioanalyzer, gives detailed descriptions of all steps necessary to run flow cytometric assays, and shows how to analyze and evaluate results using histograms and dot plots. • “Working with Chip Data and Assays” on page 258 shows you what to do to open, save, import and export files, and how to print the results. • “Administering System Functions” on page 299 is your guideline for configuring the 2100 expert software. • “Running Instrument Diagnostics” on page 316 shows how to use the diagnostic tests to check the bioanalyzer hardware for proper functioning.

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• “Performing Verifications” on page 325 describes how you can validate your bioanalyzer system. • “Products, Spare Parts, and Accessories” on page 334 lists all parts and accessories-including reorder numbers-that are required for electrophoretic and flow cytometric measurements. • “Glossary” on page 338 explains terms in context with flow cytometry, electrophoresis, and terms specific to the bioanalyzer software and hardware. If you have any questions this manual cannot answer, please refer to the supplemental literature listed in “Related Documents” on page 8. If you still have questions, contact Agilent for additional support at: http://www.agilent.com/chem/labonachip

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Related Documents A collection of supplemental literature is given in the following. Bioanalyzer Manuals Publication Number

Title

G2938-90006

Agilent 2100 Bioanalyzer Installation and Safety Manual

G2946-90003

Agilent 2100 Bioanalyzer Maintenance and Troubleshooting Guide

CD-ROM Publication Number

Title

G2946-60002

Agilent 2100 Bioanalyzer – How to Use Multimedia CD-ROM

Reagent Kit Guides The Reagent Kit Guides give you information on how to perform specific assays, including sample and chip preparation. Publication Number

Title

G2938-90300

Kit Guide Binder English (including all Reagent Kit Guides)

G2938-90010

Reagent Kit Guide DNA 500 and DNA 1000 Assay

G2938-90020

Reagent Kit Guide DNA 7500 and DNA 12000 Assay

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Publication Number

Title

G2938-90030

Reagent Kit Guide RNA 6000 Nano Assay

G2938-90040

Reagent Kit Guide RNA 6000 Pico Assay

G2938-90050

Reagent Kit Guide Protein 200 Plus Assay

G2938-90060

Reagent Kit Guide Protein 50 Assay

G2938-90070

Reagent Kit Guide Cell Fluorescence Assays

G2938-90080

Reagent Kit Guide Cell Fluorescence Checkout Kit

Application Notes and Technical Notes Application Notes and Technical Notes are available from the Agilent 2100 Bioanalyzer Help Desk or from the lab-on-a-chip web pages: http://www.agilent.com/chem/labonachip Newly Published Documentation Follow this link to see if there is any new documentation: http://www.chem.agilent.com/scripts/Library.asp

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How to Use this Manual This manual uses convenient online navigation features and follows certain typographic conventions.

Online Navigation Use the interactive bookmarks in this frame to move to your desired topic.

Use Acrobat Reader’s navigation bar to move around within a topic.

Click here to go to the table of contents.

Click here to go to the index.

Here is the current page number. ▲ Displays the previous page.

▼ Displays the next page. Contents

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After you have chosen a topic with the bookmarks, use the buttons in Acrobat Reader’s toolbar to move around within the topic. Displays the next page.

Returns to the previous view. Click several times to undo more view changes.

Displays the previous page.

Displays the first page. Displays the last page.

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Returns to the next view. Click several times to redo more view changes.

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Layout Conventions The following typographic conventions are used in this manual: Highlight

Meaning

Italic

On-screen element Example: ... the OK button. Emphasis Example: Right-click the ... Term Example: Dot plots show events as dots. Reference to another document Example: Refer to the Agilent 2100 Bioanalyzer Troubleshooting and Maintenance Guide.

Blue

Cross-reference or hyperlink Examples: “Introduction to the Key Features of the 2100 expert” on page 29 http://www.agilent.com/chem/labonachip

Courier

Code Example: .. the command line parameter -port 2 ...

Courier bold

User input Example: Enter 50 MB.

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Safety Notices, Notes and Tips Safety notices, notes and tips in this document have the following meaning: WA R N I N G A warning notice denotes a hazard. It calls attention to an operating procedure, practice, or the like that, if not correctly performed or adhered to, could result in personal injury or death. Do not proceed beyond a warning notice until the indicated conditions are fully understood and met.

C A U T IO N A caution notice denotes a hazard. It calls attention to an operating procedure, practice, or the like that, if not correctly performed or adhered to, could result in damage to the product or loss of important data. Do not proceed beyond a caution notice until the indicated conditions are fully understood and met.

NOTE A note contains important, helpful, or additional information.

TIP A tip usually points out a timesaving feature.

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Quick Start

The following step-by-step instructions guide you through a measurement with the Agilent 2100 bioanalyzer. Preparing the Agilent 2100 Bioanalyzer 1 Ensure that the proper cartridge is installed in the bioanalyzer. You can identify the installed cartridge by the number engraved at the front. Engraved number Note that there are also electrode cartridges without an engraved number.

c = Electrode Cartridge for electrophoretic assays d = Pressure Cartridge for flow cytometric assays

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2 If you have to change the cartridge, follow the instructions in “Loading the Electrophoresis Chip into the Bioanalyzer” on page 58 or “Loading the Cell Chip into the Bioanalyzer” on page 189, respectively. Switching on the Agilent 2100 Bioanalyzer 1 Make sure the bioanalyzer is connected to line power and connected to the PC. 2 Turn on the line switch at the rear of the instrument. The status LED at the front of the bioanalyzer should light up. Lid Status LED

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The status LED shows you the current status of the instrument. Signal

Meaning

Green light

Instrument is switched on and ready for measurement.

Green blinking

Measurement is running.

Orange blinking

Instrument is busy (running self diagnostic, for example).

Red light

Instrument is not ready for measurement. Switch the instrument off and on again. If the problem persists, call Agilent service.

Running a Measurement 1 To start the 2100 expert software on the connected PC, go to your desktop and double-click the following icon:

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After startup of the software, you enter the Instrument context:

NOTE If you started 2100 expert for the first time after installation, you first need to activate the different software modules with your license keys. See Figure ,“How to Activate Software Licenses” for details.

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The Instrument tab shows you the status of the bioanalyzer: Icons

Meaning Bioanalyzer detected, lid is open.

Bioanalyzer detected. Lid is closed, but no chip is inserted.

No bioanalyzer has been detected. Check the COM Port setting (see figure under step 3), the RS 232 connection cable, the power cable, and the power switch. For details on how to set up the bioanalyzer and connect it to a PC, see Agilent 2100 Bioanalyzer Installation and Safety Guide. 2 Make sure that a bioanalyzer has been detected before continuing.

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3 Select an assay for the chip run. On the Instrument tab, click the Assays button. – OR – Click the Assays menu. Both will open a menu, allowing you to select an assay for the measurement. Note that you can also select File > Open File to Run. This opens a dialog box allowing you to load either an assay (.xsy) or a chip data file (.xad). 4 Prepare the samples and the chip. For detailed information on sample and chip preparation refer to: – Reagent Kit Guides that are available for each reagent kit – Application Notes that are available for specific kits and applications NOTE When preparing chip and samples, pay attention to the essential measurement practices described in “Essential Measurement Practices (Electrophoretic Assays)” on page 55 and “Essential Measurement Practices (Flow Cytometric Assays)” on page 187 or as described in the respective Reagent Kit Guide.

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5 Insert the chip in the Agilent 2100 bioanalyzer: a Open the lid. The status of the bioanalyzer is updated on the Instrument tab.

b Check that the cartridge is inserted properly and the chip selector is in the correct position (“1” for electrophoretic assays, “2” for flow cytometric assays). For details, please refer to “Loading the Electrophoresis Chip into the Bioanalyzer” on page 58 or “Loading the Cell Chip into the Bioanalyzer” on page 189, respectively.

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