Bio-Rad PROTEAN IEF Cell Accessories User Manual

Cup Loading Tray for the
PROTEAN

®

IEF Cell

For technical service
call your local Bio-Rad office or
in the U.S. call 1-800-4BIORAD
(1-800-424-6723)
On the Web at discover.bio-rad.com

Table of Contents

SSeeccttiioonn 11 IInnttrroodduuccttiioonn ........................................................................................................................................................................11

1.1 Overview..........................................................................................1

1.2 Introduction ....................................................................................1

1.3 Description ......................................................................................3

1.4 Specifications ..................................................................................4

SSeeccttiioonn 22 IInnssttrruuccttiioonnss............................................................................................................................................................................44

2.1 General Overview ............................................................................4

2.2 IPG Strips Rehydration Solution Preparation and

Strip Rehydration ............................................................................5

2.3 Transfer and Placement of Rehydrated IPG Strips in the

Focusing Tray ..................................................................................5

2.4 Placement of Movable Electrodes....................................................6

2.5 Positioning Sample Loading Cups and Loading Sample..................6

2.6 Focusing Conditions ........................................................................8

2.7 After Focusing Run is Complete ......................................................9

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3.1 Focusing Tray ................................................................................10

3.2 Movable Electrode Assemblies ......................................................11

3.3 Cups ............................................................................................11

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Section 1
Introduction

1.1 Overview

The Cup Loading Tray for the PROTEAN IEF cell is designed for the optimal resolution and
separation of proteins with isoelectric points (pI) at either extreme of the pH range. The Cup
Loading Tray can accommodate up to 12 IPG strips from 7 cm to 24 cm in length.

The Cup Loading Tray includes a tray base, a pair of movable electrode assemblies, and one bag
each (120 count) of 100 and 150 µl sample cups. No modifications to the PROTEAN IEF cell are
needed for compatibility with the Cup Loading Tray.

The Cup Loading Tray offers the following benefits:

• Enhanced resolution and separation of proteins with pIs at extreme pH ranges

• Optimal focusing for users of ReadyStrip™ IPG strips pH 3-6, pH 7-10, and Micro-range
ReadyStrip IPG strips pH 6.3–8.3

• Tray runs up to 12 IPG strips per run for high throughput

• Tray accommodates 7 cm to 24 cm IPG strips with movable electrode assemblies

• Two sample cup sizes, 100 and 150 µl, are available to allow greater flexibility

• The Cup Loading Tray is compatible with all PROTEAN IEF cells, so no modification to
existing PROTEAN IEF cell is necessary.

1.2 Introduction

Isoelectric focusing of proteins in the acidic and basic pH ranges is often associated with
streaking due, in part, to protein precipitation when utilizing standard in-gel rehydration of IPG
strips. Protein resolution can be markedly improved when the proteins are applied to the IPG
strips by means of a sample cup positioned in a discreet area of a rehydrated IPG strip.

The Cup Loading Tray allows for sample application via cup loading after rehydration of the IPG
strips. Certain conditions make cup loading the preferred method of sample loading. The
examples below compare in gel rehydration resolution with cup loading resolution of an E. coli
sample on pH 3-6 and pH 7-10 IPG strips.

1

pH 3 pH 6 pH 3 pH 6

A. pH 3-6 cup loading tray B. pH 3-6 in-gel rehydration

pH 7 pH 10 pH 7 pH 10

C. pH 7-10 cup loading tray D. pH 7-10 in-gel rehydration

Figure 1.
In-gel rehydration: 400 µg E.coli lysate in 8M Urea, 4% Chaps, 50 mM DTT, 0.2% Bio-Lytes.
Cup loading: 400 µg E.coli lysate in 100 µl 8M Urea, 4% Chaps, 50 mM DTT, 0.2% Bio-Lytes.
A. pH 3-6 cup loading at cathode (-), B. pH 3-6 in-gel rehydration
C. pH 7-10 cup loading at anode (+), D. pH 7-10 in-gel rehydration

2

10%

20%

8%

16%

1.3 Description

The Cup Loading Tray for use with the PROTEAN IEF cell consists of the following components:

Figure 2. Diagram of Cup Loading Tray components.

1. A twelve-cup strip that holds up to 100 µl of sample per cup and a twelve-cup strip that holds
up to 150 µl of sample per cup.

Note: For precise mechanical fit, a minimum of two cups is required. A single sample cup
provides an incomplete “stop” on the channel walls and the gel could be punctured.

2. A twelve channel focusing tray that accommodates IPG strips from 7cm to 24 cm.

3. Two movable electrode assemblies, each having 12 individual teeth with cavities which allow
mineral oil to flow through the entire channel of the focusing tray.

3

Spring cup

Large (150 µl) sample cup

Small (100 µl) sample cup

µl

Electrode

Electrode

Spring clip

Right handle

Left handle

Tray base

Electrode
contact area

Cup loading tray

Electrode
contact area