Antigen‐induced proliferation assay for mouse T lymphocytes. Response to a monovalent antigen

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Abstract
This report describes an antigen‐specific proliferative assay which makes use of lymph node cells obtained from mice injected subcutaneously at the base of the tail one week previously. When lymphocytes from only the draining lymph nodes of mice immunized with a variety of different antigens, were stimulated in flat‐bottomed microculture plates with the immunizing antigen, a substantial degree of proliferation was observed, measured as incorporation of [3H]thymidine. Monovalent immunogen p‐azobenzenearsonate‐L‐tyrosine, (ABA‐Tyr, 500 daltons) which induces T cell responses in guinea pigs without antibody production, was studied in detail. In a typical experiment, ABA‐Tyr gave ∼ 17000 cpm while control cultures without antigen showed ∼ 1600 cpm. The response is antigen‐specific and seems to be mediated by T cells, since (a) it can be abolished by pretreatment of these cells with anti‐Thy‐1.2 serum and complement, (b) cells from nu/nu mice primed with ABA‐Tyr do not show any antigen‐induced proliferation, in contrast to the response observed in nu/+ and BALB/c controls, and (c) purified T cells require macrophages to proliferate, but neither macrophages nor B cells appear to divide in culture. Thus, this rapid and easy technique should prove useful for the in vitro study of murine T lymphocytes.