Open AccessTransfer in SDS of biotinylated proteins from acrylamide gels to an avidin-coated membrane filter
Author(s) -
Arthur Karlin,
Chaojian Wang,
Jing Li,
Qiang Xu
Publication year - 2004
Publication title -
biotechniques/biotechniques
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.617
H-Index - 131
eISSN - 1940-9818
pISSN - 0736-6205
DOI - 10.2144/04366rr02
Subject(s) - biotinylation , avidin , chemistry , gel electrophoresis , biotin , sodium dodecyl sulfate , polyacrylamide gel electrophoresis , biochemistry , isoelectric focusing , chromatography , microbiology and biotechnology , biology , enzyme
Avidin was covalently linked to aldehyde-derivatized polyethersulfone membrane filters. These filters were used in Western blot analysis of proteins reacted with biotinylation reagents and electrophoresed in sodium dodecyl sulfate (SDS) on polyacrylamide gels. Electrophoretic transfer from the gels to these filters was in 0.1% SDS, in which the covalently bound avidin retained its biotin-binding capacity. We compared Western blots on avidin-coated membrane filters of biotinylated and nonbiotinylated forms of mouse immunoglobulin G (IgG), mouse IgG heavy chain, muscle-type acetylcholine receptor α subunit, and fused α and β subunits of receptor. Biotinylated proteins were captured with high specificity compared to their nonbiotinylated counterparts and sensitively detected on the avidin-coated membranes.