SDS-PAGE and guanidine....
roney.graf at uni-konstanz.de
Thu Jun 17 05:33:11 EST 1999
In article <376861F8.641D8E4B at home.net>, jqpublic at home.net wrote:
> Anyone know a good way to concentrate a protein sample so that it can be
> used is SDS-PAGE if the sample is in 7M guanidine? I have sephacryl
> column fractions in said buffer but need to run 50+ uL in the gel to see
> the bands.
> Using no precipitation causes a 'funky, gunky' ppt to form on top of the
> wells and the electrophoresis does not work. TCA preciptation causes the
> guan. to pp't as well, as does any method requiring a 'cold' step(ice,
> I'm currently using a MeOH/chloroform/water method, but it is very
> tedious for 40+ samples and it cannot accomodate volumes of more than
> 150 uL sample to still be done in eppendorf tubes.
TCA-ppt works if you dilute to about 1-1.5M GuHCl first, then add 1Vol
of 10% TCA. With your sample volumes, this still won't fit into 1.5ml
Eppies, however. HTH.
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