GenScript eStain User Manual

Version 2.0
June 4, 2010

Prot e in Staining System

User Manual

For electric staining of proteins in mini polyacrylamide gels

The Biology CRO

Table of Contents

Product Contents

Product Specifications

eStain

TM

Protein Staining Device

Accessory Products

Introduction

Quick Reference Guide

Protocols

Troubleshooting

Examples of Results

Technical Support

Warranty

1

3

5

7

8

11

12

21

22

23

24

eStainTM Protein Staining System www.genscript.com

Product Contents Product Contents,

Type of products

eStain

TM

Protein Staining

Device Contents

Upon Receiving
the Instrument

eStain

TM

Protein Staining

Pads

Product

eStainTM Protein Staining Device

Cat. No.

L02010

Component

eStainTM Protein Staining Device
Specific Power Cord based on the type
of unit ordered (for U.S./Canada/China/
Taiwan/Japan, Europe, or UK)
Forceps
Shovel
eStainTM Graphite Electrode (11mm)
(installed inside the device)
Ice Pack

Quantity

1 each

1 each

1 each
1 each
1 each

1 each

Product

eStainTM Protein Staining Pads (R-250, 20-pak)
eStainTM Protein Staining Pads (G-250, 20-pak)

Cat. No.

L02011
L02012

This manual is supplied with the following products:

The contents of the eStainTM Protein Staining Device are listed below

Examine the unit carefully for any damage incurred during transit. File any
damage claims with the carrier. The warranty does not cover in-transit
damage.

The following eStain

TM

Protein Staining Pads are available from GenScript:

If you ordered the eStainTM Protein Staining Pads, you will receive the
components listed in the table below. Store the eStainTM Protein Staining
Pads at room temperature. For best results, use the eStainTM Staining Pads
before the expiration date printed on the package.

eStainTM Protein Staining System www.genscript.com eStain

continued

eStain

TM

Protein Staining

Pads, continued

eStainTM Graphite
Electrode

Product

eStainTM Graphite Electrode (11mm, 1-pak)

Cat. No.

L02014

The following eStainTM Graphite Electrode is available from GenScript:

For best results, when a Graphite Electrode was used for 200 times of electric
staining, replace it with a new one.

Component

The eStain

TM

Protein Staining Pads
(R-250, 20-pak) contains:
eStainTM Protein Staining Pad (R-250)
Gel Window (66mm×56mm)
Gel Window (76mm×64mm)
The eStainTM Protein Staining Pads
(G-250, 20-pak) contains:
eStainTM Protein Staining Pad (G-250)
Gel Window (66mm×56mm)
Gel Window (76mm×64mm)
each pack eStain

TM

Protein Staining Pad
(R-250) contains:
1×eStainTM Cathode Pad (R-250)
1×eStainTM Anode Pad
each pack eStainTM Protein Staining Pad
(G-250) contains:
1×eStainTM Cathode Pad (G-250)
1×eStainTM Anode Pad

TM

Protein Staining System www.genscript.com

Quantity

20

1
1

20

1
1

1
1

1
1

- 2 --1 -

Product Specifications Product Specifications,

eStainTM Protein Staining
Device Specifications

- 3 -

Weight:
Dimensions:
Electrical Parameters:
Built-in Features:
Compatibility:

Materials:

Operating Temperature:
Forceps:
Shovel:

2.3 kg
200 mm (l) ×180 mm (w) ×90 mm (h)
100-120 V, 220-240 V, 50/60 Hz, 3.2 A
Digital display, alarm, light LED
Suitable for electric staining of proteins in mini
polyacrylamide gels
Acrylonitrile Butadiene Styrene, Polycarbonate,
Aluminum, Titanium, Plasticized silicone.
15 - 40°C
Stainless steel
Polycarbonate

eStainTM Protein Staining
Pads Specifications

size:
Materials:

8cm (l) x 7cm (w) x2.5mm (thick)
Blotting filter paper presoaked with proprietary
cathode buffer containing CBB R-250 or G-250

size:
Materials:

8cm (l) x 7cm (w) x2.5mm (thick)
Blotting filter paper presoaked with proprietary
anode buffer

The eStainTM Protein Staining Pads are used with the eStainTM Protein Staining
Device.

The specifications of the eStainTM Protein Staining Pads are listed below:

eStainTM Cathode Pad (R-250 or G-250)

Avoid acetone, dimethyl sulfoxide, and acetic acid. These reagents can erode or
damage the device.

eStainTM Anode Pad

Inter frame size:

Materials:

66mm×56mm
76mm×64mm
Polycarbonate

Gel Window

eStainTM Protein Staining System www.genscript.com eStain

- 4 -

continued

eStainTM Graphite
Electrode Specifications

Dimensions:
Weight:
Materials:

100 mm (l) ×90 mm (w) ×11mm (h)
180g
Powdered carbon, clay

eStain

TM

Graphite Electrode (11mm)

The eStain

TM

Graphite Electrode is used as the replaceable anode of eStain

TM

Protein Staining Device and available separately from GenScript.
The specifications for eStainTM Graphite Electrode are listed below:

TM

Protein Staining System www.genscript.com

eStainTM Protein Staining Device eStain

Front View of
eStain

TM

Device

Rear View of
eStain

TM

Device

- 5 -

The rear view showing various parts of the eStain

TM

Protein Staining Device

is shown below.

The front top view showing various parts of the eStain

TM

Protein Staining Device

is shown below.

Control Panel

Handle Bar

eStainTM Protein Staining System www.genscript.com eStain

Power Switch AC Inlet

- 6 -

TM

Protein Staining Device,

continued

Control Panel of
eStain

TM

Device

Inside View of
eStain

TM

Device

The control panel of the eStainTM Protein Staining Device is described below.
When upper status light is on, which indicates staining mode, the Digital
Display shows running time; otherwise when both upper and lower status lights
are on, which indicates numbering mode, the Digital Display shows how many
times of electric staining the eStainTM Graphite Electrode has been used for.
The Reset button is used to clear parameters.
The Time button is used to set running time and toggle between staining mode
and numbering mode.
The Start/Stop button is used to activate/stop the staining program.

The inside view showing various parts of the eStain

TM

Protein Staining

Device is shown below:

TM

Protein Staining System www.genscript.com

Titanium Cathode

Graphite Anode

Accessory Products Introduction

eStainTM Protein Staining System www.genscript.com

eStainTM Protein Staining
Pads

eStain

TM

Graphite

Electrode

Additional Products

- 7 -

eStain

TM

Graphite Electrode is available separately from GenScript. Ordering

information is provided below.

The precast Express

TM

PAGE Gels as well as premade buffers or buffer powders
are available from GenScript. For details, contact Technical Support or visit

www.genscript.com.

eStain

TM

Protein Staining Pads are available separately from GenScript. Ordering

information is provided below.

Product

eStainTM Protein Staining Pads
(R-250, 20-pak)
eStainTM Protein Staining Pads
(G-250, 20-pak)

Cat. No.

L02011

L02012

Quantity

1 box

1 box

Product

eStainTM Graphite Electrode (11mm, 1-pak)

Cat. No.

L02014

Quantity

1 unit

- 8 -

eStainTM Protein Staining System www.genscript.com

System Overview

System Components

The eStainTM Protein Staining System is a unique electric staining system that
allows you to quickly, reliably and efficiently stain proteins in various types of
mini polyacrylamide gels with Coomassie blue dye reagents. It consists of the
eStainTM Protein Staining Device and eStainTM Protein Staining Pads.
The proprietary electric staining technology of the eStainTM Protein Staining
Device combined with the eStainTM Protein Staining Pads applies a high
electric force generated between graphite anode and titanium cathode to
allow for quick and directional movement of negatively charged Coomassie
blue dye reagents into the gel matrix to stain the proteins and also the quick
and directional movement of the free staining reagents out of the gel matrix to
destain the gel matrix within only 7 minutes or less. The eStainTM Protein
Staining System integrates three steps of conventional Coomassie blue
staining method into a single step and greatly cuts down the time required for
protein staining analysis. The proteins stained using the eStainTM Protein
Staining System exhibit as high detection sensitivity as proteins stained using
conventional Coomassie blue staining method.

The eStain

TM

Protein Staining System consists of:

eStainTM Protein Staining Device

The eStainTM Protein Staining Device is a user-friendly electric staining unit
that allows for fast, convenient and efficient in-gel protein staining with
Coomassie blue dye reagents.

eStainTM Protein Staining Pads

The eStainTM Protein Staining Pads are disposable pads to perform electric
staining of proteins in gels with Coomassie blue dye. Two types of eStainTM
Protein Staining Pad, R-250 and G-250, are available separately from
GenScript and for different requirements. Each pack eStainTM Protein Staining
Pad contains an eStainTM Cathode Pad presoaked with proprietary cathode
buffer containing CBB dye R-250 or G-250, and an eStainTM Anode Pad
presoaked with proprietary anode buffer, allowing for rapid, convenient and
reliable in-gel protein staining without the need to prepare any solutions and
buffers.

Introduction,

continued

eStainTM Protein Staining System www.genscript.com

System Components,

continued

System Mechanism

- 9 -

The eStain

TM

Protein Staining System is based on the electric staining concept
which utilizes the unique and patent-pending technology developed by GenScript.
To use the eStainTM Protein Staining System for in-gel protein staining,
assemble the eStainTM Protein Staining Pad with your pre-run gel on the eStainTM
Protein Staining Device. Similar to semi-dry blotting, the eStainTM Cathode Pad
and eStainTM Anode Pad act as ion reservoirs that contain the appropriate anode
and cathode buffers incorporated into the gel matrix. Simultaneously the eStainTM
Cathode Pad presoaked with proprietary cathode buffer containing CBB dye
R-250 or G250, also supplies negatively charged staining reagents. The high
electric force generated between graphite anode and titanium cathode allows
for rapid and directional movement of negatively charged Coomassie blue dye
reagents into the gel matrix to stain the proteins and also the homogenous
movement of the free staining reagents out of the gel matrix to destain the gel
matrix within only 7 minutes or less. Therefore the eStainTM Protein Staining
System provides fast, convenient and reliable in-gel protein staining without the
need to prepare any solutions and buffers.

eStain

TM

Graphite Electrode

The eStainTM Graphite Electrode serves as the replaceable anode of the eStainTM
Protein Staining Device. For the common use, one eStainTM Graphite Electrode
(11mm) is pre-installed inside the eStainTM Protein Staining Device to constitute a
complete electric staining unit. Each eStainTM Graphite Electrode is produced with
special process and can be steadily used for 200 times of electric staining.

- 10 -

Introduction,

continued

System Mechanism,

continued

eStainTM Protein Staining System www.genscript.com

Features

Important features of the eStainTM Protein Staining System are listed below:

· Unique electric staining technology created for fast, reliable and efficient
protein gel staining in as few as 7 minutes.

· User-friendly electric staining unit allowing for easy and convenient
procedures.

· Consumable staining pads offering convenience without the need for
additional solutions and buffers.

· High staining efficiency as conventional Coomassie blue staining method.

· Compatible for use with various types of mini polyacrylamide gels.

Schematic mechanism of eStain

TM

Protein Staining System showing the

movement of Coomassie blue dye:

eStainTM Cathode Pad

eStain

TM

Anode Pad

Pre-run gel

E

E

E

E

CBB
dye

CBB
dye

CBB
dye

CBB
dye

CBB
dye

CBB
dye

Titanium Cathode
2e

-

+ 2H2O H2 + 2OH

-

Graphite Anode
4OH- - 4e- O2 + 2H2O

eStainTM Protein Staining System www.genscript.com

- 11 -

Quick Reference Guide

Mode

eStainTM plugged in

eStainTM device and

staining pad assembled

Time selection

Run

End of run

Checking using
frequency
of graphite anode

Replacement of worn
graphite anode

Action

Connect eStain

TM

Device to an electrical
outlet and power
switch is on
Place staining pads
on the device and close
lid
Press Time button to
select desired running
time
Press Start/Stop
button
Automatic

Press and hold Time
button for 2 seconds

Switch off the device
and replace the worn
graphite anode with a
new one

Sound

–

–

–

–

Continuous beeping
for 2 minutes

–

–

Light

Steady upper
light

Steady upper
light

Steady upper
light

flashing upper
light
Steady upper
light

Steady upper
and lower light

–

Display

Default

running

time (000)

Default

running

time (000)

Running

time

defined

Count down

time

Default running

time (000)

Staining times

the graphite

anode has

been used for

–

A quick reference guide for operating the eStain

TM

Protein Staining Device

is provided below.

Introduction

- 12 -

Protocols

eStainTM Protein Staining System www.genscript.com

Recommendations

1. Check the Power Cord supplied with the unit to ensure that the cord is
compatible with the local socket format.

2. Place the eStainTM Protein Staining Device on a level laboratory bench.
Keep the area around the device clear to ensure proper ventilation of the
unit.

3. For your safety: Position the device properly such that the power switch
and the AC inlet located on the rear of the unit are easily accessible.

4. Ensure the AC power switch is in the Off position.

5. Attach the power cord to the AC inlet and then to the electrical outlet.
Use only properly grounded AC outlets and power cords.

6. When the electrophoresis of your samples is almost complete, press the
power switch (located on the rear of the device) to turn ON the eStainTM
Protein Staining Device. The upper status light is on indicating you are
using staining mode. Digital Display shows the default parameters (000).

To obtain the best results follow these recommendations:

· Wear gloves at all times during the entire staining procedures to prevent
contamination of pads and gels.

· Avoid using expired eStain

TM

Protein Staining Pads. Always use the pads

before the specified expiration date printed on the package.

· Always keep the Ice Pack supplied with eStain

TM

Device stored at -20℃ or

below to facilitate cooling electrodes procedure.

Installing the
eStain

TM

Device

You are ready to use the eStainTM Protein Staining Device for staining
applications.

eStainTM Protein Staining System www.genscript.com

Assembling Staining
Stack with eStain

TM

Pads and Pre-run Gel

- 13 -

1. Lift up the handle bar to vertical position, then push it forward to open the
lid of eStain

TM

Device.

2. Remove one package labeled as eStain

TM

Protein Staining Pad from the
eStainTM Protein Staining Pads box and tear the laminated sealing of the
package. Remove the two small packages respectively labeled as 1 x
eStainTM Cathode Pad R-250 (or G-250), and 1 x eStainTM Anode Pad.

Protocols,

continued

- 14 -

Assembling Staining
Stack with eStain

TM

Pads and Pre-run Gel,

continued

eStainTM Protein Staining System www.genscript.com

4. Carefully remove the pre-run gel containing your protein samples from the
gel cassette and rinse the gel with 50 to 100 ml distilled water for about 1
minute.

5. Place the gel on the eStainTM Anode Pad. Gently remove air bubbles
between gel and anode pad using the small shovel supplied with the
device.

3. Tear the sealing of the 1 x eStain

TM

Anode Pad package. Remove the
eStainTM Anode Pad from the package and place it on the anode plate of
the eStain

TM

Protein Staining Device.

Protocols,

continued

Protocols,

continued

eStainTM Protein Staining System www.genscript.com

Assembling Staining
Stack with eStain

TM

Pads

and Pre-run Gel,

continued

- 15 -

8. Pull back the handle bar to close the lid of eStain

TM

Device while keeping

the bar vertical to the lid, and then return the bar to horizontal position.

Note: Gel Window is used as the spacer between anode pad and cathode pad to prevent
short circuit. The edges of the pre-run gel may also be covered by Gel Window and
will probably be stained to deep blue after running an electric staining program.

6. Select appropriate Gel Window according to actual size of the gel (see the
table below). Place Gel Window on the gel. Ensure that the Gel Window fully
cover the margin of the anode pad.

Pre-run gel

Gel with size 66 x 56mm to 80 x 60 mm
Gel with size <66 x 56 mm

Gel Window

Gel Windows 76 x 64mm
Gel Windows 66 x 56mm

7. Tear the sealing of the 1 x eStainTM Cathode Pad package. Remove the
eStainTM Cathode Pad from the package and place it on the gel.

- 16 -

Protocols,

continued

eStainTM Protein Staining System www.genscript.com

Gel Thickness

0.75 mm

1.0 mm

1.5 mm

Running Time

6 minutes
7 minutes
9 minutes

Performing electric
staining

2. Press the Start/Stop button to activate the electric staining program. The
running time begins to count down and upper status light keeps flashing
during the whole staining program.

After assembling the staining stack, perform electric staining as described
below.

1. Press the Time button to set appropriate running time based on the gel
thickness (see table below). If an undesired running time is set by mistake,
press Reset button to clear the wrong time, and then press again the Time
button to choose the desired running time.

Note: During electric staining process, the graphite electrode may be heated to 60-70℃.
For continuous use and best results, it is necessary to cool down both graphite
anode and titanium cathode to room temperature before the next run

．

(see next page for cooling procedures)

eStainTM Protein Staining System www.genscript.com

- 17 -

3. At the end of the staining, current automatically shuts off and the eStain

TM

Protein Staining Device signals the end of staining with repeated beeping
sounds. Upper status light stops flashing and the Digital Display shows
text (000).

4. Press any button on the control panel to stop the beeping.

5. Proceed to disassemble the stack and clean the device.

To obtain good staining results, disassemble the staining stack right away after
ending the staining procedure.

1. Open the lid of eStain

TM

Protein Staining Device.

2. Carefully separate the stained gel from the staining stack and proceed to be
photographed.

3. Discard the used eStainTM Protein Staining Pad.

Note: Do not re-use the eStainTM Protein Staining Pad after staining. Discard after each
use.

4. Clean the titanium cathode plate, graphite anode plate and its surrounding
area with a dry cloth or paper towel.

For continuous use and best results, it is necessary to cool down both graphite
anode and titanium cathode to room temperature before the next run. Place the
pre-frozen ice pack on the graphite anode, close the lid gently to let the titanium
cathode also touches the ice pack. Do not force to close the lid otherwise the ice
pack will damage the device.

At this point, the eStain

TM

Protein Staining Device is ready for another run. If
you are not using the device, turn Off the power switch located on the back of
the device.

Disassembling and
cleaning the
eStain

TM

Device

Cooling Graphite
Anode and Titanium
Cathode

Protocols,

continued

Performing electric
staining, continued

- 18 -

eStainTM Protein Staining System www.genscript.com

2. Switch Off the eStainTM Protein Staining Device and then open the lid of
the device.

3. Use small shovel to pry out the worn Graphite Electrode along the gap at
either side of the anode tank.

1. If the eStainTM Protein Staining Device works at staining mode, press and
hold Time button for 2 seconds to toggle to numbering mode. If the Digital
Display shows “200” or a number greater than 200, perform the replacing
protocol as describe below.

Protocols,

continued

During electric staining process, the eStainTM Graphite Electrode will absorb ions
from anode pad and as well as lose carbon composition, thereby changing the
characteristics of the Graphite Electrode and composition of the anode buffer.
For the best staining results, after having been used for 200 times of electric
staining, the worn Graphite Electrode should be replaced by a new one.

Replacing the eStainTM
Graphite Electrode

eStainTM Protein Staining System www.genscript.com

- 19 -

4. Tear the sealing of a new eStain

TM

Graphite Electrode package and take the
new Graphite Electrode out of the package. Place the new Graphite Electrode
into the anode tank and close the lid of the device.

5. Switch On the eStainTM Device. Press and hold Time button for 2 seconds
to toggle to numbering mode.

Protocols,

continued

Replacing the eStainTM
Graphite Electrode,

continued

- 20 -

eStainTM Protein Staining System www.genscript.com

Protocols,

continued

Replacing the eStain

TM

Graphite Electrode,

continued

After successfully installing Graphite Electrode into the eStainTM Device, you
are ready to use the eStainTM Device for staining applications again.

6. Press Reset button to zero the staining times.

Note: For the best results, after using Graphite Electrode per 40 times of electric
staining, you may also need to perform a washing step by soaking the Graphite
Electrode in distilled water for half an hour and then drying it with a dry cloth or
paper towel.

eStain

TM

Protein Staining System www.genscript.com

- 21 -

Troubleshooting

Problem

The upper status light doesn’t

flash during electric staining

process

The upper and lower status

lights flash simultaneously

The stained gel has very tinged

or nearly invisible protein bands

with high blue background.

The stained gel has very tinged

or nearly invisible protein bands

with clear background.

The stained gel has hazy blue

spots.

Cause

Incomplete electric circuit due

to improper assembly of the

staining pads.

Excessive current is flowing

through the Device.

1. The staining time is not

long enough.

2. Cooling the electrodes

for too long a time.

3. The graphite electrode has

been used for 200 times or

above.

1. The staining time is too long

2. The electrodes have not

been cooled or the cooling time

is not long enough

1. The gel has not been destained

sufficiently.

2. Cooling the electrodes for too

long a time

3. Air bubbles present in staining

stack interfering gel staining.

Solution

Ensure the staining stack is

assembled correctly: use the

eStain

TM

Anode Pad first

followed by the gel, Gel

Window and eStain

TM

Cathode

Pad.

Check the staining stack and

ensure Gel Window covered

correctly on the gel.

1. Cover Gel Window and the

eStain

TM

Cathode Pad again

and perform electric

staining for 2 more minutes.

2. Follow the instruction to

perform cooling procedure.

3. Replace the worn graphite

electrode using a new one.

1. Re-assemble the staining

stack using a new set of

eStain

TM

Protein Staining

Pad and then perform

another 7-min electric

staining.

2. Follow the instruction to

perform cooling procedure.

1. Cover Gel Window and the

eStain

TM

Cathode Pad again

and perform electric

staining for 1 more minute.

2. Follow the instruction to

perform cooling procedure.

3. Be sure to remove all air

bubbles present in the

staining stack by using the

small shovel supplied with

eStain

TM

Device

Review the information below to troubleshoot your experiments using the eStain

TM

Protein Staining Device and eStain

TM

Protein Staining Pad.

Introduction

- 22 -

eStain

TM

Protein Staining System www.genscript.com

Examples of Results

Samples of 5μl New England Biolabs® Protein Ladder (P7703S) in each lane

were separated on a 4-20% Bis-Tris gel and stained using eStain

TM

Protein

Staining System (R-250).

Samples were separated on a 12% Tris-Glycine gel and stained using eStain

TM

Protein Staining System (R-250).

Tris-Glycine gel
results

Bis-Tris gel
results

1 2 3 4 5 6 7 8 Lane 1 500ng rabbit IgG

Lane 2 10μl

E.coli

lysate

Lane 3 200ng reduced BSA

Lane 4 125ng reduced BSA

Lane 5 50ng reduced BSA

Lane 6 15ng reduced BSA

Lane 7 10μl New England Biolabs

®

Protein Ladder

(Cat. No. P7703S)

Lane 8 10μl GenScript Smart Middle

Range Protein Standard

(Cat. No. MM0900)

eStainTM Protein Staining System www.genscript.com

- 23 -

Technical Support

Visit the GenScript Web site at www.genscript.com for:

1. Technical resources, including manuals, vector maps and sequences,
application notes, MSDSs, FAQs, formulations, citations, handbooks, etc.

2. Complete technical support contact information

3. Access to the GenScript Online Catalog

4. Additional product information and special offers

For more information or technical assistance, call, write, fax, or email.

GenScript USA Inc.

120 Centennial Ave.
Piscataway, NJ 08854
Tel: [phone]
Fax: [phone]
Email: [email protected]

Web Resources

Contact Us

- 24 -

eStainTM Protein Staining System www.genscript.com

eStain

TM

Protein Staining

Device

GenScript warrants that eStainTM Protein Staining Device will be free from
defects in material and workmanship for a period of one year from date of
purchase. If any defects occur in the product during this warranty period,
GenScript will, at its option, repair, replace, or refund the purchase price of
this product at no charge to you. The following defects, however, are
specifically excluded:

1. Defects caused by improper operation.

2. Repair or modification done by anyone other than GenScript or an
authorized agent.

3. Use of fittings or other spare parts supplied by anyone other than
GenScript.

4. Damage caused by accident or misuse.

5. Damage caused by disaster.

6. Corrosion due to use of improper solvent or sample.
For any inquiry or request for repair service, contact GenScript after
confirming the model and serial number of your instrument. For your
protection, items being returned must be insured against possible damage or
loss. This warranty shall be limited to the replacement of defective products.
It is expressly agreed that this warranty will be in lieu of all warranties of
fitness and in lieu of the warranty of merchantability.

Warranty

GenScript USA Inc.

120 Centennial Ave.
Piscataway, NJ 08854
Tel: [phone]
Fax: [phone]
Email: [email protected]