Hello
I have two antibodies for flow cytometry for the same antigen but they seem to bind differently as they give different percentages (one is low and the other is high). I don't know the sequence of both antibodies but would like to know the epitope that the antibodies bind to. Is there a database for antigens or is there a tool when I can put my sequence so it predicts the epitope?
Many thanks
Abed
Dear the binding of the antibodies will be predicted with the specificity of the antibodies.
If you are working with flow, you need to take FSC, SSC antibody markers separately by reading instructions/SOPs were available.
There are a number of factors, besides epitope specificty, that could influence your FACS results. One of them is affinity, as a higher affinity usually results in stronger signals irrespectively of the epitope recognized.
Anyway, if you want to explore the influence of epitope recognition, you could look for information about the atibodies nyou are using, perhaps their specificities are already known and published.
A computer program could tell you regions on the target antigen that are likely to be recognized by antibodies (in general) taking into account solvent accessibility and other properties, but no method can predict the epitope specificity of a given antibody unless you have more information about this particular antibody-antigen pair.
For instance, docking methods can be used to predict the binding interface, but to use them you need to know the sequence of the target antigen and of the antibody variable regions in order to model their structures (or use their available structures if already known). Co-cristallyzation of the antigen-antibody complex, followed by X-ray diffraction, can provide you a picture of the interaction showing the recognized epitope.
Other experimental approaches also allow to map the epitope recognized by each antibody , by studying recognition of antigen fragments or synthetic peptides resembling the epitope or by introducing mutations in the antigen to identify critical residues contributing to binding. All of them require performing specific epitope mapping studies.
Are you interested in predicting the epitope bound by each mAb, or do you just want to know if they are binding the same/overlapping epitopes? If the former, there is no easy way (as Gertrudis Rojas explained above), but if the latter, a competition ELISA will give you the answer.
Hope it helps!
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