I am creating a sample test of a LFA on a nitrocellulose membrane. I have seen some contradicting protocols regarding blocking the membrane so far. Some suggest doing it before I spot the membrane with my antibodies/conjugates whereas some suggest doing it afterwards or even both. In what order should I be doing this in? Also, would 1% or 0.1% BSA be best for blocking?
Thank you in advance!
L
Typically you do specific protein binding to NC first then block with bulk protein. Your antibody will not be immobilized on the NC if its already completely blocked. A higher BSA concentration would be expected to block more quickly. There are instances where other proteins (casein) or even surfactants (Tween 20) are better choices for blocking NC or PS. All of these variables will need testing in order to be optimized for your specific test, good luck!
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