INCREASED FREQUENCY OF INTERLEUKIN-4-PRODUCING T-CELLS AS A RESULT OF POLYCLONAL PRIMING - USE OF A SINGLE-CELL ASSAY TO DETECT INTERLEUKIN-4-PRODUCING CELLS

A limiting dilution assay capable of detecting interleukin 4 (IL4) production by a single cell has been developed. This assay is based on the stimulation of T cells, in the presence of IL 2 (5 U/ml), with anti-CD3 antibody bound to the surface of Terasaki wells. Cells of the IL 4-selective indicator line, CT.4S, are added 24-36 h later and IL 4 production is determined based on their survival 24-48 h thereafter. A frequency of 0.98 was obtained for IL 4 production by T cells of the D10.G4 cell line. T cells from naive donors capable of producing IL 4 in response to anti-CD3 plus IL 2 were quite rare, with a frequency in four experiments ranging between 0.0003 and 0.0018. Treatment of mice with polyclonal activators known to increase the IL 4-producing capacity of T cells when assayed in bulk culture caused striking increases in the frequency of IL 4-producing cells. Similarly, culturing cells in vitro with anti-CD3, IL 2 and IL 4 for 5 days caused a marked increase in the frequency of cells capable of producing IL 4, to 0.031.IL 4 production by individual T cells is dependent upon IL 2. Thus, in naive T cell populations, the frequency of IL 4-producing cells in response to stimulation with anti-CD3 in the absence of IL 2 was below the limit of detection. T cells from primed donors showed a striking inhibition in the frequency of IL 4-producing cells in response to anti-CD3 when IL 2 was not present. The availability of a simple assay to measure the frequency of cells capable of producing IL 4 should have substantial utility in allowing the evaluation of conditions that regulate IL 4 production in vivo and in vitro.