ThermoFisher Scientific Invitrogen iBlot 3 User manual

For Research Use Only. Not for use in diagnostic procedures.
iBlot™3 Western Blot Transfer System
USER GUIDE
For dry electroblotting of proteins from mini and midi gels
Catalog Number IB31001
Publication Number MAN0028464
Revision A.0

Life Technologies Holdings Pte Ltd | Block 33 | Marsiling Industrial Estate Road 3 | #07-06, Singapore 739256
For descriptions of symbols on product labels or product documents, go to thermofisher.com/symbols-definition.
Revision history:MAN0028464 A.0 (English)
Revision Date Description
A.0 5 January 2023 New document for iBlot™3 Western Blot Transfer System.
The information in this guide is subject to change without notice.
DISCLAIMER: TO THE EXTENT ALLOWED BY LAW, THERMO FISHER SCIENTIFIC INC. AND/OR ITS AFFILIATE(S) WILL NOT BE
LIABLE FOR SPECIAL, INCIDENTAL, INDIRECT, PUNITIVE, MULTIPLE, OR CONSEQUENTIAL DAMAGES IN CONNECTION WITH OR
ARISING FROM THIS DOCUMENT, INCLUDING YOUR USE OF IT.
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TRADEMARKS: All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified.
©2023 Thermo Fisher Scientific Inc. All rights reserved.

Contents
■CHAPTER1Productinformation .................................................. 6
Product contents ................................................................ 6
iBlot™3 Western Blot Transfer Device contents ................................. 6
Before starting ............................................................. 6
Installing theinstrument ..................................................... 7
Service operation requirements ............................................... 8
Unpackinginstructions .......................................................... 8
Upon receiving theinstrument ................................................ 8
Unpacking the iBlot™3 Western Blot TransferDevice ............................ 8
iBlot™3 Western Blot TransferDevice .............................................. 8
Deviceviews ............................................................... 8
About the system ............................................................... 9
iBlot™3 Western Blot Transfer System ......................................... 9
Features ................................................................... 9
Systemcomponents ........................................................ 9
System overview ........................................................... 9
Description of parts ............................................................ 11
iBlot™3 Western Blot TransferDevice ........................................ 11
Blotting surface ........................................................... 11
Lids andcooling ........................................................... 12
Controlpanel .............................................................. 12
Blotting roller .............................................................. 12
Power cord and power adapters ............................................. 12
iBlot™3 Transfer Stacks .................................................... 12
Guidelines for iBlot™3 Transfer Stacks ....................................... 13
Topstack ................................................................. 14
Bottomstack .............................................................. 15
Transfer membrane ........................................................ 16
iBlot™Filter Paper .......................................................... 17
iBlot™3 Absorbent Pad ..................................................... 17
iBlot™3 Western Blot Transfer System User Guide 3

Operating the iBlot™3 Western Blot TransferDevice ................................ 18
First time usage of the iBlot™3 Western Blot TransferDevice .................... 18
Control panel of the iBlot™3 Gel TransferDevice .............................. 18
Set the date andtime ...................................................... 19
Write logs to a USB storagedevice .......................................... 19
Description ofmethods ......................................................... 20
Methods .................................................................. 20
Recommended running parameters .......................................... 20
■CHAPTER2Protein transfer protocol ........................................... 22
Experimental overview .......................................................... 22
Experimentaloutline ....................................................... 22
Generalguidelines ............................................................. 22
Recommended geltypes ................................................... 23
Using the iBlot™3 Western Blot Transfer System ................................... 23
Materialsneeded .......................................................... 23
Select amethod ........................................................... 24
Assemble the iBlot™3 Transfer Stack ......................................... 24
Performblotting ........................................................... 26
Disassemble the iBlot™3 Transfer Stack ...................................... 26
■CHAPTER3Custommethods ................................................... 28
Create custommethods ........................................................ 28
Create a newmethod ...................................................... 28
Edit existingmethods ...................................................... 30
Create custom methods from an existing template (up to 10 steps) .............. 31
■APPENDIXATroubleshooting .................................................... 32
Introduction ................................................................... 32
■APPENDIXBPost transferanalysis .............................................. 35
Post transferanalysis ........................................................... 35
■APPENDIXCOptimizingblotting ................................................ 36
Optimizingblotting ............................................................. 36
Contents
4iBlot™3 Western Blot Transfer System User Guide

■APPENDIXDMaintenance ....................................................... 37
Cleaning ...................................................................... 37
General maintenance ........................................................... 37
Replacing thefuse ............................................................. 37
Replacing electricalcontacts .................................................... 39
■APPENDIXEProductspecifications ............................................. 41
iBlot™3 Western Blot Transfer Devicespecifications ................................ 41
iBlot™3 Transfer Stackspecifications ............................................. 42
■APPENDIXFAccessory products ............................................... 43
iBlot™3 Transfer Stacks ......................................................... 43
Additional products ............................................................ 43
Precast gels and premadebuers ................................................ 44
■APPENDIXGSafety ............................................................... 45
Symbols on thisinstrument ..................................................... 45
Standard safetysymbols ................................................... 45
Control and connectionsymbols ............................................. 46
Safety information for instruments not manufactured by Thermo FisherScientific ...... 47
Instrumentsafety .............................................................. 47
Electricalsafety ........................................................... 47
Cleaning anddecontamination .............................................. 48
Instrument component and accessorydisposal ................................ 48
Safety and electromagnetic compatibility (EMC) standards .......................... 48
Symbols oninstrument ..................................................... 48
Safety standards .......................................................... 49
EMC standards ............................................................ 49
Environmental design standards ............................................. 50
Radio compliance standards ................................................ 50
Chemicalsafety ................................................................ 51
■APPENDIXHDocumentation and support ...................................... 52
Customer and technical support ................................................. 52
Limited product warranty ........................................................ 52
Contents
iBlot™3 Western Blot Transfer System User Guide 5

Product information
Product contents
iBlot™3 Western Blot Transfer Device contents
This manual is intended to provide instructions for the operation of the iBlot™3 Western Blot Transfer
Device (Cat. No.IB31001).
The contents of the iBlot™3 Western Blot Transfer Device are listed below. See “iBlot™3 Western Blot
Transfer Device specifications” on page41 for specifications and description of the iBlot™3 Western
Blot Transfer Device.
Component Quantity
iBlot™3 Western Blot Transfer Device 1
Blotting Roller 1
Power Cord 1
Before starting
Before you begin using this product, or any installation or service operation, please read the following
safety information. Attention to these warnings will help prevent personal injuries and damage to the
products.
It is your responsibility to use the product in an appropriate manner. This product is designed for use
solely in laboratory environments, and must not be used in any way that may cause personal injury or
property damage.
You are responsible if the product is used for any intention other than its designated purpose
or in disregard of Thermo Fisher Scientific instructions. Thermo Fisher Scientific shall assume no
responsibility for such use of the product.
The product is used for its designated purpose if it is used in accordance with its product
documentation and within its performance limits.
Using the product requires technical skills and a basic knowledge of English. It is therefore essential
that only skilled and specialized sta or thoroughly trained personnel with the required skills be allowed
to use the product.
Keep the basic safety instructions and the product documentation in a safe place and pass them on to
the subsequent users.
Applicable local or national safety regulations and rules for the prevention of accidents must be
observed in all work performed.
1
6iBlot™3 Western Blot Transfer System User Guide

The iBlot™3 Western Blot Transfer Device complies with the TUV Rhineland North America Inc. safety
requirements, part 15 of the FCC rules, and the European Community Safety requirements. Operation of
the iBlot™3 Western Blot Transfer Device is subject to the conditions described in this manual.
Operation of the iBlot™3 Western Blot Transfer Device is subject to the following conditions:
•Indoor use
•Altitude below 2,000 meters
•Temperature range: 4 to 30°C
•Maximum relative humidity: 80% (maximum relative humidity 80% for temperatures up to 31°C,
decreasing linearly to 50% relative humidity at 40°C)
•Installation categories (over voltage categories) II; Pollution degree 2
•Mains supply voltage fluctuations not to exceed 10% of the nominal voltage (100–240 V, 50/60 Hz,
6.3 A)
•Mains plug is a disconnect device and must be easily accessible.
•Do not attempt to open the iBlot™3 Western Blot Transfer Device. To honor the warranty, the iBlot™
3 Western Blot Transfer Device can only be opened and serviced by Thermo Fisher Scientific.
•The protection provided by the equipment may be impaired if the equipment is used in a manner
not specified by Thermo Fisher Scientific.
•The device must be connected to a mains socket outlet with protective earthing connections.
•Ventilation requirements: Room ventilation
•Do not use if device becomes cracked or broken in any area.
Installing the instrument
The product may be installed only under the conditions and in the positions specified by Thermo Fisher
Scientific.
Following are the required operating position and conditions:
•Do not place the product in an area where it will be subject to vibration.
•Do not place the product on surfaces, vehicles, cabinets or tables that for reasons of weight or
stability are unsuitable for this purpose.
•Do not place the product on heat-generating surface or near heat emitting devices such equipment
racks or heaters. Verify that there is sucient clearance between the product and any other system
that may exhaust warm air.
•The product's ventilation should not be obstructed. If proper ventilation is not provided it can result
in electric shock, fire and/or serious personal injury or death.
•The product is for indoor use only.
•Use only with suitably rated mains supply cord (having 3 conductors, min. 16 AWG or 1.5 mm2,
min. 300V, Harmonized Type for Europe and UL Listed/CSA Certified for North America, with
molded plug rated min. 10A).
•A tolerance of ±10% shall apply to the nominal input voltage and ±3% to the nominal frequency,
over voltage category 2.
•Maximum operating altitude 2,000 m asl. Maximum transport altitude 4,500 m asl.
Chapter1Product information
Product contents 1
iBlot™3 Western Blot Transfer System User Guide 7

Service operation requirements
In the event of an equipment malfunction, it is the responsibility of the customer to report the need for
service to Thermo Fisher Scientific or to one of the authorized agents. For service information, contact
Technical Support.
Servicing of this device is to be performed by trained service personnel only.
Unpacking instructions
Upon receiving the instrument
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.
Unpacking the iBlot™3 Western Blot Transfer Device
•Remove the roller from the Styrofoam packaging.
•Remove the device from the Styrofoam packaging.
•Remove the protective film from the touch screen.
•Remove blue protective tape from the electrode contact positions on the base.
iBlot™3 Western Blot Transfer Device
Device views
Figure1Front, rear, and top views of the iBlot™3 Western Blot Transfer Device
Chapter1Product information
Unpacking instructions
1
8iBlot™3 Western Blot Transfer System User Guide

About the system
iBlot™3 Western Blot Transfer System
The iBlot™3 Western Blot Transfer System consists of the iBlot™3 Western Blot Transfer Device and
associated iBlot™3 Transfer Stacks (sold separately). The iBlot™3 Western Blot Transfer Device has a
unique design, which, in conjunction with the patented gel matrix technology of the iBlot™3 Transfer
Stacks, results in a shortened distance between electrodes, high field strength, and high currents to
reduce transfer times when blotting proteins onto membranes.
Western blotting of proteins from midi- or mini-sized polyacrylamide gels onto nitrocellulose or PVDF
membranes can be performed with iBlot™3 Transfer Stacks in less than 10 minutes.
Features
•Pre-programmed methods for protein transfers from various gel types in 3–8 minutes
•Enhanced user safety with built-in safety features in the device
•User-friendly design with an integrated power supply to avoid inconsistencies associated with the
use of an external power supply
•Fast, reliable protein transfer with ready-to-use iBlot™3 Transfer Stacks with no need to prepare
buers or membranes
•Compatible for use with Novex™and NuPAGE™gel types and other gels
•Not compatible with E-PAGE™gels
System components
iBlot™3 Western Blot Transfer Device
The iBlot™3 Western Blot Transfer Device is a self-contained blotting unit with integrated power supply
used for fast dry blotting of proteins.
iBlot™3 Transfer Stacks
The iBlot™3 Transfer Stacks are disposable stacks that have integrated PVDF or nitrocellulose transfer
membranes to perform dry blotting of proteins. Each iBlot™3 Transfer Stack contains a copper
electrode and appropriate cathode and anode buers in the gel matrix to allow fast, reliable transfer of
proteins. See “iBlot™3 Transfer Stacks” on page43 for details.
System overview
The iBlot™3 Western Blot Transfer System is based on the dry blotting concept, utilizing the unique,
patented gel matrix technology developed for E‑Gel™gels and applied in iBlot™3 Transfer Stacks.
Each iBlot™3 Transfer Stack consists of a Bottom Stack and a Top Stack sandwiching a pre-run gel
and a nitrocellulose or PVDF membrane. The iBlot™3 Transfer Stack is assembled with the blotting
membrane on the anode side, and a pre-run gel on the cathode side.
Chapter1Product information
About the system 1
iBlot™3 Western Blot Transfer System User Guide 9

Schematic of iBlot™3 Transfer Stack showing the flow of current
9
+
_
Current flow
1
3
4
5
6
7
8
2
1Top felt with folded aluminum tab
2Copper cathode
3Cathode gel matrix
4Plastic divider
5Membrane (NC or PVDF)
6Improved anode gel matrix
7Copper anode
8Bottom felt
9Transfer tray
After the stack is assembled on the iBlot™3 Western Blot Transfer Device, and the appropriate
method is selected, the run is initiated. Transfer of proteins from the gel to the blotting membrane
is accomplished in approximately 3–10 minutes. The rapid transfer without the need for external power
supply or premade buers is possible due to the following features of the iBlot™3 Western Blot Transfer
System:
•The gel matrix of the Bottom Stack and Top Stack incorporates the appropriate anode and cathode
buers to act as ion reservoirs. This format eliminates the need for premade buers or soaked
membranes and minimizes handling that can lead to inconsistent performance.
•The copper anode does not generate oxygen gas as a result of water electrolysis, resulting in
increased transfer consistency. Conventional inert electrodes present in other blotting systems
result in oxygen generation, which can result in blotting distortion.
•The transfer stack gel matrix technology and reduced distance between the electrodes combined
with the integrated power supply allows the system to generate high field strength and increase
transfer speed.
Chapter1Product information
About the system
1
10 iBlot™3 Western Blot Transfer System User Guide