Agilent Technologies 3100 OFFGEL Fractionator User manual

Agilent Technologies
Agilent 3100
OFFGEL Fractionator
User Manual

OFFGEL User Manual
Notices
© Agilent Technologies, Inc. 2010
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Manual Part Number
G3100-90001
Edition
04/2010
Printed in Germany
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Safety Notices
CAUTION
A CAUTION notice denotes a haz-
ard. It calls attention to an operat-
ing procedure, practice, or the like
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adhered to, could result in damage
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A WARNING notice denotes a
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Microsoft ®is a U.S. registered trademark
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Software Revision
This guide is valid for B.06.30 or higher revi-
sions of the Agilent 3100 OFFGEL Fraction-
ator Firmware, where the last two digits of
the Firmware revision refer to minor revi-
sions of the Firmware that do not affect the
technical accuracy of this guide.

OFFGEL User Manual 3
In This Guide…
This book describes how to operate the Agilent 3100 OFFGEL
Fractionator including the browser based user interface.
Chapter Overview
1 Product overview
This chapter gives an overview of the Agilent 3100 OFFGEL
Fractionator. It describes the intended use and the
fractionation principles of the instrument, highlights its
features and outlines the different instrument components.
2 Site requirements and specifications
This chapter describes information about site requirements that
have to be met so that the Agilent 3100 OFFGEL Fractionator
performs within specifications.
Control PC
3100 OFFGEL Fractionator

4OFFGEL User Manual
3 PC, Browser and LAN Requirements
This chapter describes information about requirements for your
PC, browser and LAN that have to be met for the control of the
Agilent 3100 OFFGEL Fractionator.
4 Installation
This chapter describes how to install the Agilent 3100 OFFGEL
Fractionator.
5Operation
This chapter details how to operate the Agilent 3100 OFFGEL
Fractionator. It familiarizes the user with all control functions
from the internet browser interface. It describes the necessary
steps to perform a fractionation and set up a LAN connection to
a PC.
6 Maintenance
This chapter describes cleaning procedures that should be used
to do maintenance on your instrument at regular intervals or
before every use. In addition, it provides instructions on how
update the firmware.
7Troubleshooting
This chapter provides support for troubleshooting the
instrument and the application.
8Safety
This chapter contains safety information relevant to the Agilent
3100 OFFGEL Fractionator.
9 Appendix
This chapter contains details on non-condensing condition as
well as ordering information for consumables and spare parts.

OFFGEL User Manual 5
Related Documents
The Agilent 3100 OFFGEL Fractionator can be used with
different consumable kits which each are supplied with a Kit
Manual:
•5969-1582 Kit Manual

6OFFGEL User Manual

Contents
OFFGEL User Manual 7
Contents
1 Product overview 13
Intended Use 14
Fractionation Principles of the Agilent 3100 OFFGEL Fractionator 14
Features 16
Instrument and Accessories 17
2 Site requirements and specifications 21
Site Requirements for the Agilent 3100 OFFGEL Fractionator 22
Power Considerations 22
Bench Space 22
Environment 22
The Waste Electrical and Electronic Equipment (WEEE) Directive
(2002/96/EC) 23
Do not dispose of in domestic household waste 23
Sound Emission 23
Instruction for Use 24
Physical Specifications of the Agilent 3100 OFFGEL Fractionator 25
3 PC, Browser and LAN Requirements 27
PC, Browser and LAN Requirements 28
Browser Interface 28
LAN 29
Tested Configurations 29
4 Installation 31
Unpacking the Agilent 3100 OFFGEL Fractionator 32
Setting up the Agilent 3100 OFFGEL Fractionator 34
Setting up the Control PC 36

8OFFGEL User Manual
Contents
Assignment of an IP Address to the Control Computer 36
Local Network Connection (optional) 37
5Operation39
The User Interface 40
General Functions 42
Status tab 43
Working with Methods 46
Displaying Data Graphically 49
Zoom in Plots 50
Pan / Drag in Plots 50
Switching the Y-Scale in Plots 51
Printing of Plots 51
Instrument Logbook 52
Exporting Results 54
Reviewing Result Files 57
How to find the exported files on the USB stick 57
How to Display Exported Results 58
Instrument Configuration 62
Additional Information 64
Performing a Fractionation 65
Preparing a Tray for the OFFGEL Mode 66
Preparing a Tray for the Standard IPG IEF Mode 66
Starting a Run 68
6 Maintenance 69
Cleaning the Agilent 3100 OFFGEL Fractionator 70
Cleaning the Electrodes 70

Contents
OFFGEL User Manual 9
Regular Cleaning Procedure 70
Cleaning Procedure for Heavily Contaminated Electrodes 70
Cleaning the Trays 71
Cleaning the Cooling Platform 72
Cleaning the Anode Connector 72
Cleaning the Top Cover and the Housing of the Instrument 75
Firmware Update 76
Installation of the Firmware Update Tool 77
How to un-install the Firmware Update Tool 77
Installation of the Firmware on your PC 77
Connect your PC to the Agilent 3100 OFFGEL Fractionator 78
Setup of the Firmware Update Tool 79
Configure the Firmware Update Tool 80
Doing the Firmware Update 84
The Log-file 87
Firmware Change via USB Stick 88
Steps for Firmware Change via USB 89
7 Troubleshooting 91
Troubleshooting the Instrument 92
Status Indicators 92
Status LED 92
Status Bars 93
Error Messages 93
Hardware Diagnosis 96
Checking the LED and the cover sensor 96
Checking Temperature Control of the Cooling Platform 96
Checking High Voltage, Electrodes and Electrode Connectors 97
Troubleshooting the Electrodes 100

10 OFFGEL User Manual
Contents
Checking Fixed and Movable Electrodes 100
Troubleshooting the LAN Connection 101
Troubleshooting the Application 102
Looking at Run Parameters 105
Example 1. Typical Results of the Protein Test Sample 105
Example 2 . Current Drop During Fractionation 107
Example 3. Reduced Electrical Charge in One Lane 108
8Safety113
Safety Information 114
Power Cords 114
Operation 115
Safety Symbols 116
Chemical and Biological Safety 116
8安全性 119
安全性について 120
電源コード 121
操作 121
安全記号 122
化学および生物的な安全性 122
8安全 125
安全说明 126
电源线 126
操作 127
安全标志 127
化学和生物安全 128
8Sicherheit129
Sicherheitshinweise 130

Contents
OFFGEL User Manual 11
Netzkabel 131
Betrieb 131
Sicherheitssymbole 132
Chemische und biologische Sicherheit 132
8Sécurité135
Consignes de sécurité 136
Cordons secteur 137
Utilisation 137
Symboles de sécurité 138
Sécurité chimique et biologique 138
8Sicurezza141
Informazioni per la sicurezza 142
Cavo di alimentazione 143
Funzionamento 143
Simboli di sicurezza 144
Sicurezza chimica e biologica 144
8Seguridad147
Información de seguridad 148
Cables de alimentación 148
Operación 149
Símbolos de seguridad 150
Seguridad química y biológica 150
9 Appendix 153
Non-condensing Conditions 154
Ordering Information 155
List of Consumables 155
List of Spare Parts 156

12 OFFGEL User Manual
Contents

13
Agilent 3100 OFFGEL Fractionator
User Manual
Agilent Technologies
1
Product overview
Intended Use 14
Features 16
Instrument and Accessories 17
This chapter gives an overview of the Agilent 3100 OFFGEL Fractionator. It
describes the intended use and the fractionation principles of the instrument,
highlights its features and outlines the different instrument components.

14 OFFGEL User Manual
1 Product overview
Intended Use
Intended Use
The Agilent 3100 OFFGEL Fractionator system fractionates proteins or
peptides based on their isoelectric points (pI) in liquid phase (OFFGEL mode).
In addition, the system performs standard in-gel isoelectric focusing (IEF)
using immobilized pH gradient gel (IPG) strips (in-gel mode). The system
consists of an instrument that can be controlled via LAN by a User Interface
based on a typical web browser. Earlier instruments were controlled by a
handheld control module which can still be used in parallel. The web interface
provides complete control and monitoring of the Agilent 3100 OFFGEL
Fractionator. The system is capable of running a total of 16 samples in parallel
(2×8).
OFFGEL pre-fractionation of complex protein and peptide sample provides
additional sensitivity of downstream analysis tools such as LC-MS. In
addition, the pI is a useful parameter that can be used for validation of peptide
MS hits.
Fractionation Principles of the Agilent 3100 OFFGEL Fractionator
The Agilent 3100 OFFGEL Fractionator performs isoelectric focusing of
proteins or peptides in immobilized pH gradient (IPG) gel strips. The 3100's
OFFGEL electrophoresis differs from conventional gel electrophoresis in that
the sample components do not remain in the gel. Instead, they are recovered
from a buffer solution, making the recovery much easier than with
conventional gels.
The three-part diagram in Figure 1 shows starting and ending states, as well
as the OFFGEL fractionation process itself. After rehydration of gel strips, the
immobilized pH-gradient (IPG) gel seals tightly against the compartment
frame. Then, the diluted sample is distributed across all wells in the strip
(Figure 1A). When a voltage is applied to the ends of the gel strip, the proteins
or peptides move through the gel until the molecules reach a well covering a
portion of the gel where the pH equals the pI of the molecule (Figure 1B).
After fractionation, the liquid fractions containing pI-based separated
proteins or peptides can easily be removed with a (multichannel-) pipette and
processed for downstream experiments (Figure 1C).

OFFGEL User Manual 15
Product overview 1
Intended Use
Figure 1 Fractionation principle of the Agilent 3100 OFFGEL Fractionator.
A
B
C

16 OFFGEL User Manual
1 Product overview
Features
Features
•pI-based OFFGEL fractionation with in-solution recovery for easy transfer
to downstream methods such as LC/MS.
•Reproducible pI-based OFFGEL fractionation down to 0.1 pH resolution
depending on the consumable kit used.
•Preconfigured methods for OFFGEL and traditional in-gel IEF protein and
peptide fractionation.
•Compatible with other upstream or downstream methods
(immunodepletion, LC-MS, gel-based analysis)
•pI obtained acts as an additional validation parameter.
•No MS-incompatible additives used in fractionation buffer
•The instrument can also be operated in the traditional in-gel mode (IEF).
•Fractionation of up to 16 samples in parallel on 2 trays, accommodating 8
samples each.
•Broad loading capacity range from 50 µg sample load up to 5 mg sample
load.
•Two independent power supplies allow two different methods to be run at
the same time.
•Easy method setup and monitoring of important fractionation parameters
and settings such as voltages and currents during a run.
•Online current control for each individual sample or gel-strip during
fractionation.
•Viewing of up to three run parameters in an online plot.
•Easy export of important run parameters of a fractionation via LAN to the
control PC or to a USB stick for further analysis
•Automated storage of up to 1200 data files, including important run
parameters, on the compact flash card
•Monitoring of all operations and error events using the self-updating
logbooks.

OFFGEL User Manual 17
Product overview 1
Instrument and Accessories
Instrument and Accessories
Figure 2 gives an overview of the complete Agilent 3100 OFFGEL Fractionator
system:
Figure 2 Instrument overview
Cooling Platform
Status LED
On/Off Serial number
Top Cover
Trays

18 OFFGEL User Manual
1 Product overview
Instrument and Accessories
The instrument has two individual high voltage power supplies which provide
voltages to the two trays. On the left side of the cooling platform is the anode
connector; across the cooling platform are two cathode connectors (see
Figure 3).
Figure 3 View inside the instrument.
Attached to each tray are a fixed electrode and a movable electrode that
establish the connection to the anode and the cathode connectors (see
Figure 4). Each tray accommodate up to eight samples. Each OFFGEL
fractionation requires a frame to build up the well compartments as depicted
schematically in Figure 1. There are two different frame lengths available,
depending on the consumable kit and application chosen. The 12-well frame is
typically used for fractionations requiring lower resolution, the 24-well frame
is used for fractionations requiring higher resolution. Trays and electrodes can
be ordered as spare parts if needed (for ordering details refer to Table 15 on
page 156).
Cooling Platform
Anode Connector
Trays
Cathode Connectors

OFFGEL User Manual 19
Product overview 1
Instrument and Accessories
Figure 4 Electrodes, trays and frames
The frames are included in the consumable kits. The available consumable kits
are listed in Table 1.
Fixed Electrode [Anode]
Movable Electrode [Cathode]
Tray
Fixed Electrode [Anode]
Movable Electrode [Cathode]
Tray
Cover Seals
Frames
Table 1 Consumable kits
Product Order Number
3100 OFFGEL, High Res Kit, pH 3-10 5188-6424
3100 OFFGEL, Low Res Kit, pH 3-10 5188-6425
3100 OFFGEL, High Res Kit, pH 4-7 5188-6426
3100 OFFGEL, Low Res Kit, pH 4-7 5188-6427
3100 OFFGEL, Starter Kit 5188-6444
OFFGEL, Protein Test Sample 5188-6428

20 OFFGEL User Manual
1 Product overview
Instrument and Accessories