The use of nitrocellulose immunoblots for the analysis of antigen recognition by T lymphocytes.
Antigen insolubilised on nitrocellulose is able to activate T lymphocytes following presentation by appropriate accessory cells. Furthermore, complex mixtures of antigens can be fractionated by polyacrylamide gel electrophoresis under denaturing conditions (SDS-PAGE), transferred to nitrocellulose membranes and then added to T cell proliferation assays. This allows a direct screening for recognition of individual polypeptides by monoclonal and polyclonal T cell populations, without a requirement for extensive biochemical purification and in the absence of initial serological preselection of antigen. The potential use of this technique for screening of recombinant DNA libraries for expression of antigens recognised by T cells is also illustrated. The technical aspects of T cell recognition of solid-phase antigen and its application to the identification of immunodominant epitopes with the potential to modulate T cell-mediated immunity is reviewed.
Lamb
J R
JR
MRC Tuberculosis and Related Infections Unit, Royal Postgraduate Medical School, Hammersmith Hospital, London, U.K.
O'Hehir
R E
RE
Young
D B
DB
D000941:Antigens / Q000276:immunology / Q000302:isolation & purification*; D001402:B-Lymphocytes / Q000276:immunology; D002999:Clone Cells / Q000276:immunology; D003001:Cloning, Molecular / Q000379:methods; D003101:Collodion; D004591:Electrophoresis, Polyacrylamide Gel / Q000295:instrumentation; D006801:Humans; D007158:Immunologic Techniques / Q000295:instrumentation; D008213:Lymphocyte Activation; D008567:Membranes, Artificial; D011993:Recombinant Fusion Proteins / Q000032:analysis; D013601:T-Lymphocytes / Q000276:immunology*
D000941:Antigens; D008567:Membranes, Artificial; D011993:Recombinant Fusion Proteins; D003101:Collodion