Sophie Richert1, Sylvie Luche2, Mireille Chevallet2, Alain Van Dorsselaer1, Emmanuelle Leize‐Wagner1, Thierry Rabilloud2,3
1IPHC - Institut Pluridisciplinaire Hubert Curien (France)
2BECP - Bioénergétique Cellulaire et Pathologique (CEA-Grenoble 17 rue des martyrs F-38054 Grenoble cedex9 - France)
3LCBM - UMR 5249 - Laboratoire de Chimie et Biologie des Métaux (17, rue des Martyrs 38054 Grenoble cedex 09 - France)
Tóm tắt
AbstractThe mechanism by which silver staining of proteins in polyacrylamide gels interferes with mass spectrometry of peptides produced by proteolysis has been investigated. It was demonstrated that this interference increases with time between silver staining and gel processing, although the silver image is constant. This suggested an important role of the formaldehyde used in silver staining development in this interference process. Consequently, a formaldehyde‐free staining protocol has been devised, using carbohydrazide as the developing agent. This protocol showed much increased peptide coverage and retained the sensitivity of silver staining. These results were however obtained at the expense of an increased background in the stained gels and of a reduced staining homogeneity.
