Comprehensive proteomic analysis of human bile

Bile serves diverse functions from metabolism to transport. In addition to acids and salts, bile is composed of proteins secreted or shed by the hepatobiliary system. Although there have been previous efforts to catalog biliary proteins, an in‐depth analysis of the bile proteome has not yet been reported. We carried out fractionation of non‐cancerous bile samples using a multipronged approach (SDS‐PAGE, SCX and OFFGEL) followed by MS analysis on an LTQ‐Orbitrap Velos mass spectrometer using high resolution at both MS and MS/MS levels. We identified 2552 proteins – the largest number of proteins reported in human bile till date. To our knowledge, there are no previous studies employing high‐resolution MS reporting a more detailed catalog of any body fluid proteome in a single study. We propose that extensive fractionation coupled to high‐resolution MS can be used as a standard methodology for in‐depth characterization of any body fluid. This catalog should serve as a baseline for the future studies aimed at discovering biomarkers from bile in gallbladder, hepatic, and biliary cancers.