Bio-Rad Mini Trans-Blot® Cell - Manual (Page 21)

Mini-Trans-Blot Electrophoretic Transfer Cell 15
Section 4
Strategies for Optimizing Electrophoretic
Transfer
4.1 Optimizing Protein Transfer
Generally, quantitative elution of denatured high molecular
weight proteins is difficult. The following tactics, alone or in
combination, will increase transfer efficiency.
Vary gel composition
Gradient gels are often more effective than single gel
concentrations for elution of a wide range of molecular
weight proteins.
Lower the total monomer to create a more porous gel.
Increase or decrease the percentage of crosslinker. A
5.26% C gel will contain the smallest pore size of all gels
no matter what the concentration of acrylamide. Decrease
in %C will make gels more porous with little loss in
resolution.
grams bis
grams bis + grams acrylamide
Increase transfer time
An initial control should be performed to determine the
time required for complete transfer.
18, 25 Times may vary
from as little as 30 minutes to as long as overnight.
Remember all overnight applications should be performed
at 30 volts to minimize heating problems.
Increase the power
Initial controls should be performed to evaluate the
efficiency of increasing the V/cm as well as its effects on
the temperature of transfer. The temperature increase may
change buffer resistance and subsequent power delivered,
as well as the state of protein denaturation, thus affecting
transfer efficiency.
%C = x 100