Transfer in SDS of biotinylated proteins from acrylamide gels to an avidin-coated membrane filter.
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 alpha subunit, and fused alpha and beta subunits of receptor. Biotinylated proteins were captured with high specificity compared to their nonbiotinylated counterparts and sensitively detected on the avidin-coated membranes.
Karlin
Arthur
A
Center for Molecular Recognition, College of Physicians and Surgeons, Columbia University, 630 West 168th Street, New York, NY 10032, USA. ak12@columbia.edu
Wang
Chaojian
C
Li
Jing
J
Xu
Qiang
Q
D000178:Acrylamides; D001360:Avidin; D019921:Biotinylation / Q000379:methods*; D015153:Blotting, Western / Q000379:methods*; D020099:Coated Materials, Biocompatible; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D005782:Gels; D008567:Membranes, Artificial; D011506:Proteins / Q000032:analysis* / Q000276:immunology
D000178:Acrylamides; D020099:Coated Materials, Biocompatible; D005782:Gels; D008567:Membranes, Artificial; D011506:Proteins; D001360:Avidin