GenScript eBlot User manual

GenScript USA Inc.
860 Centennial Ave.
Piscataway, NJ 08854
Tel: [phone]
Fax: [phone]
Email: [email protected]
Version No. 08152014
For fast semi-dry electroblotting of proteins from mini polyacrylamide
gels to nitrocellulose or PVDF membranes

Table of Contents
Product Contents
Product Specifications
eBlotTM Protein Transfer Device
Accessory Products
Introduction
Quick Reference Guide
Protocols
Troubleshooting
Examples of Results
Technical Support
Warranty
Appendix
1
3
5
7
8
11
12
21
22
23
24
25
eBlotTM Protein Transfer System www.genscript.com

Product Contents Product Contents, continued
Type of products
eBlotTM Protein Transfer
Device Contents
Upon Receiving
the Instrument
eBlotTM Protein Transfer
Pads
Component
eBlotTM Protein Transfer Device
eBlotTM Graphite Electrode (Installed inside the device)
Sponge Cushion (Installed inside the device)
Regional specific power cord
Forceps
Shovel
Shallow Tray
Quantity
1 each
1 each
2 each
1 each
1 each
1 each
2 each
Product
eBlotTM Protein Transfer Pads
(Basic, without membrane, 20-pak)
eBlotTM Protein Transfer Pads (Nitrocellulose, 20-pak)
eBlotTM Protein Transfer Pads (PVDF, 20-pak)
Cat. No.
L03011
L03013
L03014
This manual is supplied with the eBlotTM Protein Transfer Device (L03010). Components for eBlotTM Protein Transfer Pad are as follows:
The contents of the eBlotTM Protein Transfer 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 eBlotTM Protein Transfer Pads are available from GenScript:
Component
eBlotTM Protein Transfer Pad
Nitrocellulose membranes (20 sheets)
PVDF membranes (20 sheets)
eBlotTM Equilibration Buffer (125 ml)
Gel Window (66 mm×56 mm)
Gel Window (76 mm×64 mm)
Gel Window (88 mm×78 mm)
Absorbent Filter Paper
Sponge Cushion
Basic
20
-
-
2
1
1
1
1
2
PVDF
20
-
1
2
1
1
1
1
2
Nitrocellulose
20
1
-
2
1
1
1
1
2
The eBlotTM Protein Transfer Pads come with the following components.
eBlotTM Graphite Electrode
eBlotTM Equilibration Buffer
The eBlotTM Graphite Electrode (L03012) is available separately from GenScript:
For best results, when a graphite electrode has been used for 100 times of
protein transfer, replace it with a new one.
The eBlotTM Equilibration Buffer (M01078, 125 ml) is available separately from
GenScript.
Store the eBlotTM Protein Transfer Pads at room temperature. For best results,
use the eBlotTM Protein Transfer Pads before the expiration date printed on the
package.
Component
1× eBlotTM Cathode Pad
1× eBlotTM Anode Pad
eBlotTM Protein Transfer System www.genscript.com eBlotTM Protein Transfer System www.genscript.com
Quantity
1
1
- 2 --1 -
DO NOT start a run without properly assembled Transfer Stacks in place!
! Important
Product
eBlotTM Graphite Electrode
Cat. No.
L03012
Product
eBlotTM Equilibration Buffer
Cat. No.
M01078

- 4 -
Product Specifications Product Specifications, continued
eBlotTM Protein Transfer
Device Specifications
- 3 -
Weight:
Dimensions:
Electrical Parameters:
Built-in Features:
Compatibility:
Materials:
Operating Temperature:
Forceps:
Shovel:
Shallow Tray:
1.6 kg
325 mm (l) ×195 mm (w) ×70 mm (h)
100-120 V, 220-240 V, 50/60 Hz, 3.2 A
Digital Display, Alarm, Light LED
Suitable for fast electroblotting of proteins from mini
polyacrylamide gels to PVDF or nitrocellulose membranes
Acrylonitrile Butadiene Styrene, Polycarbonate,
Aluminum, Titanium, Plasticized silicone.
15 - 40 °C
Stainless steel
Polycarbonate
Polycarbonate eBlotTM Graphite Electrode
Specifications
Dimensions:
Weight:
Materials:
105 mm (l) × 95 mm (w) × 10 mm (h)
140 g
Powdered carbon, Clay, and Stainless steel
eBlotTM Graphite Electrode
The eBlotTM Electrode is used as the replaceable anode electrode
of eBlotTM Protein Transfer Device and available separately from GenScript.
The specifications for eBlotTM Graphite Electrode are listed below:
eBlotTM Protein Transfer
Pads Specifications
eBlotTM Anode Pad:
eBlotTM Cathode Pad:
Materials:
90 mm (l) × 80 mm (w) × 2.5 mm (thickness)
90 mm (l) × 80 mm (w) × 2.5 mm (thickness)
Blotting filter paper presoaked with proprietary
anode or cathode buffer
Nitrocellulose:
PVDF:
90 mm (l) × 80 mm (w)
90 mm (l) × 80 mm (w)
The eBlotTM Protein Transfer Pads are used with the eBlotTM Protein Transfer Device.
The specifications of the eBlotTM Protein Transfer Pads are listed below:
eBlotTM Protein Transfer Pad
Intended Use For research use only. Not intended for human or animal diagnostic or therapeutic uses.
Avoid acetone, dimethyl sulfoxide, and acetic acid. These reagents can erode
or damage the device.
Membrane
Size:
Formulation:
125 ml × 2
Proprietary
eBlotTM Equilibration Buffer
Inter frame size:
Materials:
66 mm × 56 mm
76 mm × 64 mm
88 mm × 78 mm
Polycarbonate
Gel Window
Size:
Materials:
80 mm × 70 mm
Vegetable fiber
Absorbent Filter Paper
eBlotTM Protein Transfer System www.genscript.com eBlotTM Protein Transfer System www.genscript.com

- 6 -
eBlotTM Protein Transfer Device eBlotTM Protein Transfer Device, continued
Front View of
eBlotTM Device
Rear View of
eBlotTM Device
- 5 -
Control Panel of
eBlotTM Device
Top View of
Open eBlotTM Device
The control panel of the eBlotTM Protein Transfer Device is described below.
The Digital Display shows two rows of multi-digits that specify the transfer
conditions as follows:
The upper three digits after text <PN> indicate how many times of transfer the
graphite electrode has been used for.
The lower four digits specify the running time of protein transfer in minute and
second.
The two Status Lights show the working mode of the eBlotTM Protein
Transfer Device. When the right status light is on, the device is switched on
and working at transferring mode; when both left and right status lights are on,
the device is working at numbering mode.
The Reset button is used to clear parameters.
The Min. button is used to shift between
transferring and numbering mode, and to
set running time. Each short press will
increase one minute. Each long press
(2 seconds) will toggle working mode from
transferring to numbering or the opposite.
The Sec. button is also used to set running
time, each press will increase 5 seconds.
The Start/Stop button is used to activate/stop
the transfer program.
The top view of open eBlotTM Protein Transfer Device identifying various parts.
The rear view showing various parts of the eBlotTM Protein Transfer Device
is shown below.
The front-top view showing various parts of the eBlotTM Protein Transfer Device
is shown below.
eBlotTM Protein Transfer System www.genscript.com eBlotTM Protein Transfer System www.genscript.com
Power SwitchAC Inlet
Titanium Cathode
Graphite Anode
Control Panel
Open Button

- 8 -
Accessory Products Introduction
eBlotTM Protein Transfer System www.genscript.com
Precast Gels and Premade
Buffers
ONE-HOUR WesternTM
Detection Kits
- 7 -
eBlotTM Protein Transfer System www.genscript.com
System Overview
System Components
Semi-dry Western blotting is a common technique applied in protein research.
Conventional semi-dry blotting is a cumbersome process, requiring time
-consuming reagent preparation and setup, followed by an electrophoretic
transfer that could take one hour or more. GenScript’s eBlotTM Protein Transfer
System accelerates the semi-dry blotting process without sacrificing performance.
The eBlotTM Protein Transfer System, consisting of the eBlotTM Protein Transfer
Device and eBlotTM Protein Transfer Pads, enables researchers to quickly,
reliably perform electrophoretic transfer of proteins from various types of mini
polyacrylamide gels to membranes in 7 to 10 minutes without the need to
prepare additional buffers.The proteins transferred using the eBlotTM Protein
Transfer System exhibit high detection sensitivity as to proteins transferred
using other existing blotting methods.
The eBlotTM Protein Transfer System consists of:
eBlotTM Protein Transfer Device
The eBlotTM Protein Transfer Device is a self-contained electroblotting unit
with a power supply built-in that allows for fast transfer of proteins.
eBlotTM Protein Transfer Pads
As the consumable part of eBlotTM Protein Transfer System, the eBlotTM
Protein Transfer Pads are the mixed assortment of eBlotTM Protein Transfer
Pad, nitrocellulose or PVDF membranes, Equilibration Buffer and Gel
Window. Each pack of eBlotTM Protein Transfer Pad contains an 1×eBlotTM
Cathode Pad and an 1×eBlotTM Anode Pad presoaked with proprietary
cathode buffer and anode buffer respectively, allowing for rapid, convenient
and reliable protein blotting without the need to prepare additional buffers.
The ONE-HOUR WesternTM Detection Kits used for Western blotting analysis
are available from GenScript. Ordering information is provided below. For
more information, visit www.genscript.com or call Technical Support.
The precast ExpressTM PAGE Gels as well as premade buffers and buffer
powders are available from GenScript. For details, contact Technical
Support or visit www.genscript.com.
Product
ONE-HOUR Western
TM
Basic Kit (Rabbit)
ONE-HOUR Western
TM
Basic Kit (Mouse)
ONE-HOUR Western
TM
Basic Kit (Goat)
ONE-HOUR Western
TM
Standard Kit (Rabbit)
ONE-HOUR Western
TM
Standard Kit (Mouse)
ONE-HOUR Western
TM
Standard Kit (Goat)
ONE-HOUR Western
TM
Standard Kit with TMB (Rabbit)
ONE-HOUR Western
TM
Standard Kit with TMB (Mouse)
ONE-HOUR Western
TM
Standard Kit with TMB (Goat)
ONE-HOUR Western
TM
Advanced Kit (Rabbit)
ONE-HOUR Western
TM
AdvancedKit (Mouse)
ONE-HOUR Western
TM
Advanced Kit (Goat)
ONE-HOUR IP-Western Kit (Rabbit)
ONE-HOUR IP-Western Kit (Mouse)
ONE-HOUR IP-Western Kit (Goat)
ONE-HOUR Western
TM
Fluorescent Kit
ONE-HOUR Western
TM
Multiplex Fluorescent Kit
Cat. No.
L00204
L00205
L00399
L00204C
L00205C
L00228
L00204T
L00205T
L00228T
L00241
L00242
L00243
L00231
L00232
L00233
L00397
L00398
Quantity
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 Kit (5 Assays)
1 kit (5 Assays)
1 Kit (10 Assays)
1 Kit (10 Assays)

- 10 -
Introduction, continued Introduction, continued
eBlotTM Protein Transfer System www.genscript.com
System Features
- 9 -
eBlotTM Protein Transfer System www.genscript.com
Important features of the eBlotTM Protein Transfer System are listed below:
· Unique semi-dry electroblotting technique created for fast, reliable protein
transfer within 7-10 minutes.
· Self-contained electroblotting unit with an integrated power supply for easy
and convenient procedures.
· Consumable transfer pads offering convenience without the need for
additional buffers.
· Compatible for use with various types of mini polyacrylamide gels.
· Proprietary formula without methanol .
· High transfer efficiency as compared to other existing blotting methods.
System Mechanism The eBlotTM Protein Transfer System is based on the proprietary fast semi-dry
blotting technique developed by GenScript.
To use the eBlotTM Protein Transfer System for rapid protein blotting, assemble
the eBlotTM Protein Transfer Pad with your pre-run gel and the membrane on
the eBlotTM Protein Transfer Device. The eBlotTM Cathode Pad and eBlotTM
Anode Pad act as ion reservoirs that contain the appropriate anode and
cathode buffers. The design of the eBlotTM Protein Transfer Device reduces
the distance between electrodes and the integrated power supply enables the
system to generate a certain definite voltage allowing for rapid and directional
movement of negatively charged protein molecules from the gel matrix onto
the membrane within 7-10 minutes. Meanwhile, the membrane presoaked with
eBlotTM Equilibration Buffer shows good combination ability with proteins that
prevents the flow through of low molecular weight proteins from the membrane.
Schematic mechanism of eBlotTM Protein Transfer System showing the flow
of current:
TM
TM
Window

- 12 -
Protocols
eBlotTM Protein Transfer System www.genscript.com
- 11 -
eBlotTM Protein Transfer 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 eBlotTM Protein Transfer Device on a levelled 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 Power Switch is in the Off position.
5. Open the closed lid of the eBlotTM Protein Transfer Device by pressing
the Open button. Place one or two pieces of Sponge Cushion in the anode
tank depending on gel thickness. For 0.75 and 1.0mm gel, use one or two
pieces of Spong Cushion; for 1.5mm gel, use one or zero piece of Sponge
Cushion.
Note: After 20 times of protein transfer, replace the used Sponge Cushions with new
ones. A pair of new Sponge Cushions are included in each box of eBlotTM Protein
Transfer Pads.
6. Insert the eBlotTM Graphite Electrode into the anode tank as described in
Section "Replacing the eBlotTM Graphite Electrode", then close the lid of
the device.
Installing the
eBlotTM Device
Quick Reference Guide
Mode
eBlotTM Device
plugged in
eBlotTM Device and
transfer stack
assembled
Time selection
Run
End of run
Checking the number
of uses of the
graphite anode
Replacement of
worn graphite
anode
Action
Connect eBlotTM
Device to an electrical
outlet and power
switch is on
Assemble transfer
stack on the device
and close lid
Press Min. and Sec.
button to select
desired running time
Press Start/Stop
button
Automatic
Press and hold Min.
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 right
light
Steady right
light
Steady right
light
Flashing right
light
Steady right
light
Steady left
and right lights
–
Display
Default
running
time (00:00)
Default
running
time (00:00)
User specified
running
time (00:00)
Counting down
time
Default running
time (00:00)
Times of the
graphite anode
has been used
for transfer
–
A quick reference guide for operating the eBlotTM Protein Transfer
Device is provided below.
Introduction To obtain the best results, follow these recommendations:
1. Wear gloves at all times during the entire blotting procedures to prevent
contamination of pads, gels and membranes.
2. Do not touch the gel or membrane with bare or gloved hands.
This may contaminate the gel or membrane and interfere with further
analysis. If needed, always use forceps to adjust the membrane or gel.
3. Avoid using expired eBlotTM Protein Transfer Pads and eBlotTM Equilibration
Buffer. Always use the pads and buffer before the specified expiration date
printed on the package.
4. Remove any trapped air bubbles between the gel and membrane during
the assembly of the transfer stack using the small shovel supplied with the
device.

- 14 -
eBlotTM Protein Transfer System www.genscript.com
Assembling the Transfer
Stack, continued
- 13 -
eBlotTM Protein Transfer System www.genscript.com
4. Pour 10 ml eBlotTM Equilibration Buffer into the shallow tray supplied
with the eBlotTM Device.
5. Tear the sealing of the Nitrocellulose or PVDF Membranes package.
Remove one sheet of membrane and soak it in the eBlotTM Equilibration
Buffer for 1 minute.
Note: If a PVDF membrane is used, the membrane must be pre-wetted with methanol
before equilibrating in eBlotTM Equilibration Buffer. For users of eBlotTM Protein
Transfer Pads (Basic, 20-pak) (L03011), the precut membrane should be
prepared by themselves.
3. Tear the sealing of the 1×eBlotTM Anode Pad package. Remove the
eBlotTM Anode Pad from the package and place it on the anode plate of
the eBlotTM Protein Transfer Device.
Note: The pad must be placed in the middle of the graphite electrode position, can not
contact on both sides of the metal frame.
Protocols, continued Protocols, continued
8. Attach the power cord to the AC inlet and then to the electrical outlet.
Use only properly grounded AC outlets and power cords.
9. When the electrophoresis of your samples is almost complete, press
the Power Switch (located on the rear of the device) to turn ON the
eBlotTM Protein Transfer Device. The right Status Light is on indicating
you are using transfer mode. The lower four digits of the Digital Display
show the default running time (00:00).
You are ready to use the eBlotTM Protein Transfer Device for blotting application.
Assembling the Transfer
Stack
1. Open the closed lid by pressing the Open button.
2. Remove one package labeled as eBlotTM Protein Transfer Pad from the
eBlot™ Protein Transfer Pads box and tear the laminated sealing of the
package. Remove the two small packages respectively labeled 1×eBlotTM
Cathode Pad and 1×eBlotTM Anode Pad.
7. Pull out the Waste Tray from the right side of the device. Place a new
Absorbent Filter Paper inside the tray and then push the tray in.
Note: Change absorbent paper with a new one when opening a new box of pads.
Installing the
eBlotTM Device , continued

- 16 -
Protocols, continued Protocols, continued
eBlotTM Protein Transfer System www.genscript.com
Assembling the Transfer
Stack, continued Assembling the Transfer Stack, continued
- 15 -
eBlotTM Protein Transfer System www.genscript.com
8. Place the gel on the membrane. Gently remove air bubbles between the
gel and the membrane.
9. 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 covers the margin of the membrane.
10. Tear the sealing of the 1×eBlotTM Cathode Pad package. Remove the
eBlotTM Cathode Pad from the package and place it on the gel.
Note: Gel Window is used as the spacer between anode pad and cathode pad to
prevent short circuit.
Note: During assembling of the transfer stack, make sure to remove all the air bubbles
trapped between the transfer pads, pre-run gel and the membrane, which may
prevent the transfer of proteins and cause empty spots on the transferred membrane.
6. Place the equilibrated membrane on the eBlotTM Anode Pad. Gently remove
air bubbles between the membrane and the anode pad using the small
shovel supplied with the device.
7. Carefully remove the pre-run gel containing your protein samples from the
gel cassette and briefly rinse the gel with distilled water in another shallow
tray to remove any small gel pieces attached to the gel and facilitate easy
positioning of the gel on the membrane.
Pre-run gel
Gel with size > 80 × 60 mm
Gel with size 66 × 56 mm to 80 × 60 mm
Gel with size < 66 × 56 mm
Gel Window
Gel Window 88 × 78 mm
Gel Window 76 × 64 mm
Gel Window 66 × 56 mm