Need help with IEF
gab at gnv.ifas.ufl.edu
gab at gnv.ifas.ufl.edu
Tue Apr 20 14:47:53 EST 1993
Hello all!
I have just begun to do some isoelectric focusing and have run into
a snag. I am using BioRad's mini-protean cell, which uses capillary
tubes for the IEF gels. The directions that came with the kit called
for using 1.6% of ampholytes in the 5-7 pH range, and 0.4% of a broad-
range ampholyte, pH 3-10. Since the protein I am working with has a
pI of about 10, I decided to only use the broad range ampholytes, and
bring their concentration up to 2%.
The problem I have encountered is that when I checked the pH of the
gel (by cutting it into 10 pieces, putting each in a small volume
of .05 M KCL under vacuum for 2 hours, then checking the pH), I
still only get a range of 4.3 - 8.
I would appreciate any suggestions on how to get a gel with a pH
gradient of 3-10. Thank you in advance- Beverly
More information about the Methods
mailing list