Bio-Rad Trans-Blot® Cell - Manual (Page 13)

2. Pre-equilibration of gels
All electrophoresis gels should be pre-equilibrated in transfer buffer prior to electrophoretic
transfer. Pre-equilibration will facilitate the removal of contaminating electrophoresis
buffer salts and neutralization salts (salts resulting from the denaturation of nucleic acids
prior to transfer). If the salts are not removed, they will increase the conductivity of the
transfer buffer and the amount of heat generated during the transfer. Also, low percent-
age gels (<12%) will shrink in methanol buffers. Equilibration allows the gel to adjust to
its final size prior to electrophoretic transfer.
3. Current limits
The PowerPac 200 Power Supply is capable of a 200 watt output. Unless a current limit
is set, uncontrolled conductivity changes may result in full power being delivered to the
Trans-Blot cell. The gel holder and electrode cards may warp, and the transfer buffer may
boil and evaporate (further increasing conductivity). This would result in a potential safe-
ty hazard. Refer to the PowerPac 200 Power Supply Instruction Manual for setting cur-
rent limits and run times.
4. Polarity of transfer
Do not reverse polarity with the plate electrodes. This will result in corrosion and rusting
of the stainless steel cathode. If this should occur, the stainless steel should be cleaned with
a mild abrasive cleanser to remove the rust.
5. Heat dissipation with the Super Cooling Coil - Use of the Super Cooling Coil with the
Plate Electrodes
For high power applications, such as high intensity transfers and transfers performed with
the plate electrodes, efficient heat removal is only obtained using the Super Cooling Coil
connected to a refrigerated recirculating bath. Placing the Trans-Blot cell in the cold room
is an inadequate means of controlling transfer buffer temperature. The plastic tank of the
Trans-Blot cell is an effective thermal insulator, thus limiting the efficient dissipation of
heat. Use of the Super Cooling Coil is necessary with all experiments using the plate elec-
trodes and with high intensity transfers using the standard platinum wire electrodes.
6. Use of a stir bar during transfer
For all blotting applications a stir bar must be placed inside the Trans-Blot cell, so that the
transfer buffer is stirred during the course of the experiment. This will help to maintain
uniform conductivity and temperature during electrophoretic transfer. Failure to proper-
ly control transfer buffer temperature results in poor transfer of macromolecules and poses
a potential safety hazard.
7. Transfer buffer pH
Do not adjust the pH of transfer buffers unless specifically indicated. Adjustments of the
pH of transfer buffers, when not indicated, will result in increased buffer conductivity. This
is manifested by a higher than expected initial current output and a decreased resistance.
It is recommended that the buffer conductivity and resistance be checked with the
PowerPac 200 Power Supply before starting each transfer.
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