isotype controls
Alice L. Givan (Alice.L.Givan@Dartmouth.EDU)
27 Mar 96 15:07:20 EST
Scott Simon's suggestion about using the antibody itself (unconjugated and in
excess concentration) to pre-block cells before staining with the conjugated
antibody may have two practical problems: first, it is expensive, as the
pre-blocking antibody has to be used in high concentration and is, by
definition, different for every stain in your panel. Secondly, it won't work
for an indirect staining system. Another, more theoretical problem may be
that the unconjugated blocking antibody would block both the specific
(antigenic) and the non-specific (eg FC) binding of your staining antibody.
So this "pre-blocked" control may not allow you to determine what binding of
your antibody is a result of its antigen-binding site and what is a result of
its Fc-end. The suggested method could under-estimate the non-specific
binding that occurs. Any thoughts on this?
Alice Givan
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