The protocol presented was specifically optimized for in-depth analysis of the human colon mucosa proteome. After cell lysis in a sodium deoxycholate/urea buffer, a tandem digestion with Lys-C and trypsin was performed. Prior to LC-MS/MS analysis, peptides were TMT-labeled and fractionated by high pH reversed-phase spin columns. This protocol is a powerful, reproducible, sample-saving, and cost-effective option when an in-depth quantitative proteome analysis is desired.
Keywords: Colon mucosa proteome; High pH reversed-phase fractionation; Lys-C; Sodium deoxycholate; TMT labeling.