Proteomic analysis and 2-D-Gel electrophoresis of bile-duct bile: A new diagnostic approach

Aims: Extensive research has been performed on the lipid composition of bile and its mucin content, but not on differences in protein composition. Though this could identify new molecular players and markers of disease.

Methods: Bile samples were acquired by ERC from patients suffering from gallstones, primary sclerosing cholangitis (PSC), liver transplantation (LTx), and/or malignancy of the pancreatico-biliary tract. Purified protein samples were analyzed by 1- and 2-D-Gel electrophoresis, DIGE and mass spectrometry.

Results: We collected >800 bile samples from different cholestatic diseases. A new purification protocol was established removing albumin, IgG and lipids minimizing interference with further analysis and protein loss. 1-D- and 2-D-PAGE verified samples of highest quality and reproducibility. Protein concentrations from patients suffering from PSC or malignancy were markedly elevated. Protein patterns of bile revealed by 1-D and 2-D gel electrophoresis were quite similar within the gallstone group, but different to the majority of patients in the PSC and LTx group. Mass spectrometry of >200 protein spots yielded serum proteins as the major components in all groups, but a number of proteins could be detected with a higher frequency only within subgroups of cholestatic diseases. Protein composition of bile-duct bile is markedly different compared to reported gallbladder bile.

Conclusion: This newly established protocol of bile-duct bile acquisition, storage, purification and analysis enables for a new diagnostic approach in cholestatic diseases. By applying state-of-the-art techniques we could already identify that: 1) Bile-duct bile has a protein composition different than gallbladder bile, 2) Protein composition of bile of patients suffering from different diseases is stable regarding major serum proteins, but variable regarding other secreted proteins. 3) Protein patterns in 2-D-gels are different among various cholestatic diseases.