I would like to perform a immunocytochemistry (ICC) on differentiated THP-1 to evaluate the expression of a membrane receptor. I will use for that a primary conjugated antibody. On blocking steps do I really need to use a Fc receptor blocking solution or can I use regular blocking agents (like FBS or BSA)?
Hi,
It's not mandatory to use FcR blocking reagents while staining with fluorophore conjugated antibodies. You will find lot's of literature supporting this. However there is a chance of non specific binding of antibodies to FcR on monocyte/macrophage, if the antibody, which you will be using is human origin (very less likely), then I believe you should consider use of FcR blocking reagents. If it is from other species eg mouse/rabbit (more likely), then there will be no point of using this. 3% BEA will be fine.
good luck. Let me know if I can help with this further.
In our experience we have saw binding of conjugated antibodies to macrophages only once. So technically it could be possible. Since serum contains immunoglobulins (in contrast to BSA) thus using of BSA or pooled human heat inactivated AB serum could be used as source of a concurrent ligands for Fc receptor and effectively downgrades false positive signal (if exist) to minimum.
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