[Protein-analysis] What is interfering with SDS-PAGE?

Rafael Garcia rgarcia at errc.ars.usda.gov
Wed May 24 11:08:24 EST 2006


I am solubilizing difficult proteins using a pretty harsh extraction solution:
7M Urea
2M Thiourea
0.05 M DTT
0.01M Tris
2.5% SDS
1% N-lauroylsarcosine (detergent)
protease inbitor cocktail
pH7.4

Something in this mixture makes SDS-PAGE gels look streaky and dark.  If I clean up and concentrate the extracted protein with ultrafiltration, the gels work much better.  (dialysis followed by lyophilization doesn't seem to work).  Which component in this extraction solution is causing the problem?

Rafael Garcia
Chemical Engineer
USDA-ARS
Eastern Regional Research Center
600 East Mermaid Lane
Wyndmoor PA 19038
voice: 215-836-3743
fax: 215-233-6795
rgarcia at errc.ars.usda.gov





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