A METHOD FOR RAPID SCREENING OF RECOMBINANT PROTEINS FOR RECOGNITION BY LYMPHOCYTES-T

A simple, cost-effective method is described that allows rapid screening of recombinant protein sequences for their ability to stimulate T cells. Individual microcultures of E. coli each expressing a gene product or peptide sequence fused to protein A are grown in 96-well plates. Following lysis of the bacteria, the fusion peptide is readily captured with immobilized immunoglobulin in tissue culture wells. No further purification is required. T lymphocytes plus appropriate antigen-presenting cells are added directly to the wells and assayed for proliferation. The DNA in bacteria from wells stimulating T cell proliferation is then sequenced. The technique allows rapid mapping of T cell epitopes by facilitating screening of truncation mutants without extensive purification. Described here is a further application of the technique to study monosubstituted analogues of a known T cell epitope.