A statistical approach to determine monoclonality after limiting cell plating of a hybridoma clone

One of the standard methods to isolate a hybridoma clone producing a monoclonal antibody requires successive steps of limiting dilution. The probability of obtaining a monoclonal antibody increases with the number of limiting dilution steps. However, without meticulous visual screening monoclonality is hard to prove. Here we describe a statistical analysis, based on Poisson's approximation, which allows one to calculate the number of hybridoma cells at a given plating efficiency so that when seeded a predictable number of mono-, bi-, etc.-clonal cultures are obtained after the first step of limiting cell plating.