Cytosolic and Transmembrane Protein Extraction Methods of Breast and Ovarian Cancer Cells: A Comparative Study

Journal of Biomolecular Techniques : JBT
Thingreila MuinaoHari Prasanna Deka Boruah

Abstract

Efficient extraction of proteins is a great challenge for numerous downstream proteomic analyses. During the protein extraction procedure, it is critical to maintain the conformational stability, integrity, as well as higher yield of the protein. To do so, 5-different lysis buffers of Tris and HEPES have been used as the primary buffering reagents with variable compositions at different concentrations and pH using human cancer cells. In this study, different protein lysates of human breast cancer cells T47D and MDA-MB-231 and ovarian cancer cell PA-1 were subjected to run SDS-PAGE for separation of proteins based on their molecular size, followed by Coomassie blue, silver staining, and immunoblot assays to compare the extraction yield of total cytoplasmic proteins, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the integral membrane protein, integrin β-1. Our results revealed that Tris-based lysis buffer with 50 mM concentration, pH 7.5, is relatively the efficient and reliable protein extraction method for a wide range of MW subcellular markers, cytoplasmic GAPDH and transmembrane integrin β-1 proteins. We anticipate that this simple and cost-effective protein extraction protocol might be extremely useful across a broad ...Continue Reading

Citations

Jan 29, 2022·Proceedings of the National Academy of Sciences of the United States of America·Uri Nimrod Ramírez-JarquínSrinivasa Subramaniam

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