Format

Send to

Choose Destination
Proteomics. 2004 Mar;4(3):551-61.

About thiol derivatization and resolution of basic proteins in two-dimensional electrophoresis.

Author information

1
CEA-Laboratoire de Bioénergétique Cellulaire et Pathologique, DRDC/BECP Grenoble, France.

Abstract

The influence of thiol blocking on the resolution of basic proteins by two-dimensional electrophoresis was investigated. Cysteine blocking greatly increased resolution and decreased streaking, especially in the basic region of the gels. Two strategies for cysteine blocking were found to be efficient: classical alkylation with maleimide derivatives and mixed disulfide exchange with an excess of a low molecular weight disulfide. The effect on resolution was significant enough to allow correct resolution of basic proteins with in-gel rehydration on wide gradients (e.g. 3-10 and 4-12), but anodic cup-loading was still required for basic gradients (e.g. 6-12 or 8-12). These results demonstrate that thiol-related problems are not solely responsible for streaking of basic proteins on two-dimensional gels.

PMID:
14997479
PMCID:
PMC2781085
DOI:
10.1002/pmic.200300589
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Wiley Icon for PubMed Central
Loading ...
Support Center