Croos reactivity - (Mar/16/2012 )

Hi Everyone
I have a very simple quetion . How can i checked the cross reactivity between my coating antibody and secondary antibody??

Coating antibody is anti human IgG
secondary antibody is Alkaline phospatase labbeled antihuman IgG.

plz share your ideas
Thanks

The cross reactivity depends on which is the host that each antibody (capture or detection) have been produced. One good choise is to use the same host (then there is no cross reactivity) for both of antbodies, Fc fragment for coating, Fab for secondary. In order to check yr system now, make a gradient of your secondary antibody concentration and use it direct in the plate (with coating antibody) without antigen. If there will be a significant standard curve, there is strong cross reactivity.