Evaluation of two cell surface modification methods for proteomic analysis of plasma membrane from isolated mouse...

The hepatocyte is a highly polarized cell with a heterogeneous distribution of plasma-membrane (PM) proteins. To reduce the complexity of the proteome of liver tissue and give a comprehensive profile of hepatocyte PM, two PM purification methods based on cell surface modification, named the biotin-avidin (BA) and cationic silica-polyanion (CSP) strategies were evaluated and compared with the traditional cell fractionation method to prepare highly enriched PM from freshly isolated C57 mouse hepatocytes. Employing different principles for PM modification, both methods were effective in the isolation of general and purified PM fraction. The CSP strategy showed better yield for the PM purification from freshly isolated hepatocytes. 189 non-redundant proteins were identified, including 49 from the BA method and 185 from CSP strategy. Many known and novel PM-associated proteins were also found. Our evaluation here should give implications for PM preparation from other freshly isolated tissue-derived cells. The hepatocyte PM proteins identified here should be taken as a references for the PM-related functional and diseases research.