Bio-Rad Bio-Gel P Polyacrylamide Gel - Manual (Page 6)

Bio-Gel P gel is compatible with solubilizing and denaturing con-
ditions used in molecular weight determinations such as 6 M guani-
dine-HCl, chaotropic agents, reducing agents such as dithiothreitol
and mercaptoethanol, and detergents such as SDS, CHAPS, and
Triton X-100.
Volatile buffer salts, for example pyridine, acetic acid, ammonium
formate, or ammonium bicarbonate, may be employed if the final
product must be free of buffer salts. These substances are easily
removed from effluent fractions by lyophilization.
Removal of dissolved gases, primarily carbon dioxide, should be
performed to prevent bubble formation within the system. This is
done by aspirating the buffer in a vacuum flask either with a water aspi-
rator or central vacuum source.
The use of eluants with pH above 10 or below 2 should be avoid-
ed to prevent hydrolysis of the gel. Strong oxidizing agents should be
avoided because they will react with the gel and increase the con-
tent of charged groups on the matrix.
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Section 3
Instructions For Use
3.1 Column Selection
The ideal column dimensions will be those that allow baseline
resolution of analytes without significant sample dilution. Typically,
the column length to diameter ratio will be between 5 and 10 and a
bed volume 4 to 20 times the volume of the sample. The minimal
dilution factor that can be obtained for an excluded substance is
approximately 1.25. Difficult fractionation procedures generally
require bed length to diameter ratios of 25 to 100 or greater and bed
volumes 25 to 100 times the sample volume.
3.2 Eluant Selection
The eluant chosen should provide maximum stability for labile
sample solutes. The ionic strength should be at least 20 mM to elim-
inate the effect of small amounts of negatively charged groups on
the gel. Using highly concentrated salt solutions may cause small
changes in gel bed volume and exclusion limits.
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