GCANTOR at VETMED.WSU.EDU
Mon Oct 9 12:31:17 EST 1995
Adam Boyt wrote that his Westerns with a biotinylated second
antibody, which is then detected with strepavidin-HRP, was less
sensitive than when he used a second antibody which was HRP-
conjugated, but not biotinylated.
One reason could be that he is using 10% milk as a blocker. Milk
contains some biotin, which would compete for the strepavidin-HRP,
making detection less sensitive. We had a similar problem with
sensitivity, in detection of biotinylated immunoprecipitated
proteins. We switched blockers, and now use a casein preparation,
sold by Pierce. Our sensitivity increased 5-10 fold.
I'd be interested to know if this works for you, Adam.
Dept Vet Microbiology & Pathology
Washington State University
Pullman, WA 99164-7040
gcantor at vetmed.wsu.edu
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