Conditions that allow for effective transfer of membrane proteins onto nitrocellulose membrane in Western blots |
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Authors: | Abeyrathne Priyanka D Lam Joseph S |
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Institution: | Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON, Canada. |
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Abstract: | A major hurdle in characterizing bacterial membrane proteins by Western blotting is the ineffectiveness of transferring these proteins from sodium dodecyl sulfate -- polyacrylamide gel electrophoresis (SDS-PAGE) gel onto nitrocellulose membrane, using standard Western blot buffers and electrophoretic conditions. In this study, we compared a number of modified Western blotting buffers and arrived at a composition designated as the SDS-PAGE-Urea Lysis buffer. The use of this buffer and specific conditions allowed the reproducible transfer of highly hydrophobic bacterial membrane proteins with 2-12 transmembrane-spanning segments as well as soluble proteins onto nitrocellulose membranes. This method should be broadly applicable for immunochemical studies of other membrane proteins. |
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