BIO RAD Trans-Blot SD User manual

Trans-Blot®SD
Semi-Dry
Electrophoretic
Transfer Cell
Instruction
Manual
Catalog Number
170-3940
For Technical Service Call Your Local Bio-Rad Office or in the U.S. Call 1-800-4BIORAD (1-800-424-6723)

Note
To insure the best performance from the Trans-Blot SD semi-dry electrophoretic transfer
cell, become fully acquainted with these operating instructions before using the cell to
transfer samples. Bio-Rad recommends that you first read these instructions carefully. Then
assemble and disassemble the cell completely without transferring sample. After these
preliminary steps, you should be ready to transfer a sample.
Bio-Rad also recommends that all Trans-Blot SD cell components and accessories be
cleaned with a suitable laboratory cleaner (such as Bio-Rad Cleaning Concentrate, catalog
number 161-0722) and rinsed thoroughly with distilled water, before use.
Warranty
Bio-Rad Laboratories warrants the Trans-Blot SD semi-dry electrophoretic transfer cell
against defects in materials and workmanship for 1 year. If any defects occur in the instrument
during this warranty period, Bio-Rad Laboratories will repair or replace the defective parts free.
The following defects, however, are specifically excluded:
1. Defects caused by improper operation.
2. Repair or modification done by anyone other than Bio-Rad Laboratories or an authorized
agent.
3. Use of fittings or other spare parts supplied by anyone other than Bio-Rad Laboratories.
4. Damage caused by accident or misuse.
5. Damage caused by disaster.
6. Corrosion due to use of improper solvent or sample.
This warranty does not apply to parts listed below:
1. Platinum plate electrode.
For any inquiry or request for repair service, contact Bio-Rad Laboratories after con-
firming the model and serial number of your instrument.
Model___________________________________
Catalog Number__________________________
Date of Delivery___________________________
Warranty Period__________________________
Serial Number____________________________
Invoice Number___________________________
Purchase Order Number____________________

Table of Contents
Page
Section 1 Introduction.................................................................................................. 1
1.1 Specifications............................................................................................................. 1
Section 2 Equipment and Reagents............................................................................ 2
2.1 Equipment and Accessories....................................................................................... 2
2.2 Related Instruments.................................................................................................... 4
2.3 Chemical Reagents..................................................................................................... 4
Section 3 Safety Instructions....................................................................................... 5
Section 4 Trans-Blot SD Assembly ............................................................................. 6
4.1 Preparation for Blotting ............................................................................................. 6
4.2 Assembly of the Unit for Standard Transfers............................................................ 7
4.3 Assembly of the Unit for Acidic Transfers ............................................................... 10
Section 5 Buffer Formulation...................................................................................... 10
Section 6 Examples of Specific Protocols................................................................... 11
6.1 SDS-Protein Blotting ................................................................................................. 11
6.2 DNA Blotting (For acrylamide gels with DNA 250 bp to ~1 kb) ............................ 12
6.3 DNA & RNA Blotting (For agarose gels with DNA up to 23 kb,
RNA up to 3.5 kb)...................................................................................................... 12
Section 7 Properties of Protein Blotting Media......................................................... 12
Section 8 Troubleshooting Guide................................................................................ 13
8.1 Poor Transfer.............................................................................................................. 13
8.2 Poor Binding to Nitrocellulose Membrane................................................................ 14
8.3 High Background After Incubation with Antibody Probes; Nonspecific
or Nonquantitative Detection..................................................................................... 14
8.4 Poor Detection Sensitivity or No Reactivity ............................................................. 15
Section 9 References..................................................................................................... 15

Section 1
Introduction
Blotting was first performed by Southern1in 1975 with the transfer of DNA from agarose
gels to nitrocellulose membranes. Blotting has subsequently been applied to RNA2-4 and
protein5,6 from both agarose and polyacrylamide gels. Membrane materials have been expanded
to include PVDF for improved protein binding capacity. To overcome the inefficiency of
capillary transfers, electric current has been adopted for eluting proteins from polyacrylamide
gels, as first described by Towbin et al.7in 1979. Since that time, electrophoretic transfer has
also been used for DNA and RNA blotting.8-14
For blotting PCR fragments, plasmid and vector DNA, and RNA with the SD cell, use the
Trans-Blot SD DNA blotting kit. DNA or RNA can be blotted from agarose gel to
Zeta-Probe®GT membrane in only 10 minutes, without any gel pretreatments. The kit comes
complete with DNA/RNA blotting accessories and a detailed instruction manual.
Semi-dry blotting was first reported by Kyhse-Andersen in 1984.15 Blotting was
performed with plate electrodes in a horizontal configuration. The gel and nitrocellulose
membrane were sandwiched between sheets of buffer-soaked filter paper, which served as
the ion reservoir and replaced the buffer tank. The plate electrodes, separated only by the filter
paper stack, provided high field strength (V/cm) across the gel, and very efficient, rapid
transfers.
The Trans-Blot semi-dry transfer cell incorporates the original concepts of semi-dry
blotting along with innovative features for quick set-up and ease of use. The platinum-coated
titanium and stainless steel electrode pair provides efficient, background-free blotting with
trouble-free service.
1.1 Specifications
Construction
Trans-Blot SD body Molded polycarbonate
Anode Platinum-coated titanium
Cathode Stainless steel
Anode platform Precision machined acrylic
Overall size 37 cm x 24 cm x 11 cm
Maximum gel size 25 cm x 18.5 cm
Cleaning Do not immerse the unit in liquid. Use special care
when cleaning the anode plate to avoid scratching
or marring the platinum. Do not use abrasives or
strong detergents. The cathode plate (stainless
steel) can be cleaned with a mild abrasive to
remove salt that may deposit during normal opera-
tion. The entire unit can also be periodically disas-
sembled and cleaned with water to remove salt
deposits.
Chemical compatibility The semi-dry blotter components are not compati-
ble with chlorinated hydrocarbons (e.g., chloro-
form), aromatic hydrocarbons (e.g., toluene,
benzene), or acetone. Use of organic solvents
voids all warranties.
1

Section 2
Equipment and Reagents
2.1 Equipment and Accessories
Catalog
Number Product Description
170-3940 Trans-Blot SD Electrophoretic Transfer Cell
Replacement Parts
170-3942 Trans-Blot SD Anode, platinum
170-3947 Trans-Blot SD Cathode, stainless steel
DNA Blotting Kit
170-3957 Trans-Blot SD DNA/RNA Blotting Kit
Power Supply
165-4761 Model 200/2.0 Constant Voltage Power Supply, 100/120 V,
50/60 Hz
165-4762 Model 200/2.0 Constant Voltage Power Supply, 220/240 V,
50/60 Hz
Protein Blotting Accesories
Blot Absorbent Filter Paper (extra thick) Recommended uses
170-3966 Sheets, 7 x 8.4 cm pre-cut to fit
mini PROTEAN 3 Gels, 60
170-3967 Sheets, 8.3 x 13.6 cm pre-cut to fit
Criterion Gels, 60
170-3968 Sheets, 14 x 16 cm pre-cut to fit
PROTEAN xi Gels, 30
170-3969 Sheets, 19 x 19.5 cm pre-cut to fit
PROTEAN xL Gels, 30
Immuno Blot PVDF Membrane
162-0174 Sheets, 7 x 8.4 cm, 10 (mini blot size)
162-0175 Sheets, 10 x 15 cm, 10
162-0176 Sheets, 20 x 20 cm, 10
162-0177 Roll, 26 cm x 3.3 m, 1
Sequi-Blot PVDF Membrane
162-0180 Sheets, 10 x 15 cm, 10
162-0181 Sheets, 15 x 15 cm, 10
162-0182 Sheets, 20 x 20 cm, 10
162-0184 Roll, 24 cm x 3.3 m, 1
162-0185 Sheets, 20 x 20 cm, 3
162-0186 Sheets, 7 x 8. 4 cm, 10
Nitrocellulose Membrane (0.45 micron)
162-0115 Roll, 33 cm x 3 m, 1
162-0113 Sheets, 20 x 20 cm, 5
162-0114 Sheets, 15 x 9.2 cm, 10
162-0116 Sheets, 15 x 15 cm, 10
162-0117 Sheets, 9 x 12 cm, 10
162-0145 Sheets, 7 x 8.4 cm, 10 2
Immunoblotting of
protein.
High capacity for all
molecular weight
ranges.
Low background for
immunodensity.
Protein sequencing
and amino acid
analysis.
Protein blotting
Transfer of proteins
(high molecular
weight or abundant,
low molecular weight
tend to penetrate this
membrane)