Hi All,
I am ordering an overalapping peptide library to study the binding epitope of my antibody. I wonder if there is a formula to calculate the probability of number of epitope hits (e.g. single, double) with different epitope length, peptide length and offset (or peptide overlap). Knowing the probability of double hit would be helpful to determine how many peptides to order (as they are quite expensive!). Thank you for your help.
I did a *lot* of this kind of work as a post-doc, when multiple peptide synthesis was first becoming feasible. There is no easy answer to your question, as asked.
What do you know about your Ab, and what do you need to know about the epitope?
If you have a monoclonal there is about a 12% chance it will bind to a short peptide. Most MAb are specific for conformational epitopes, and it's infamously difficult to draw conclusions about their epitopes using short peptides as antigens. OTOH, if you're looking at antiserum, you're likely to find multiple epitopes, with one or two immunodominant.
What level of resolution do you need? If you plan to follow up with more synthesis and mapping you can get away with an initial experiment using fewer peptides (e.g., 12mers overlapping by 4).
Anyway, yes, making the peptides is a lot of work and expensive. So once you have them, test as many Ab as you can find.