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Western Blotting Using the Invitrogen NuPage Novex Bis Tris MiniGels
Published on: August 22, 2007
Elimination of non-specific binding in western blots from non-reducing gels
1Department of Hormone Research, Weizmann Institute of Science, Rehovot, Israel.
Abstract:
The reaction of some antibodies with Western blots of protein shows strong non-specific binding especially at a region that corresponds to about 70-90 kDa. This binding is independent of protein concentration. Further analysis indicated that the factor responsible for the non-specific binding is 2-mercaptoethanol in the gel sample buffer. Gel electrophoresis of total tissue homogenates in the absence of this reducing agent resulted in dramatic elimination of the non-specific background binding without affecting the mobility of the two proteins we studied.
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