Enumeration of IL 2-secreting helper T cells by limiting dilution analysis, and demonstration of unexpectedly high levels of IL 2 production per responding cell.
We describe here a method for estimating the frequency of alloantigen-responsive precursors of interleukin 2-(IL 2) secreting mouse T helper cells in spleen cell populations, and for measuring the amount of IL 2 generated by the progeny of each responsive cell over a 5-day culture period. Depending on the particular stimulus, about 1 T cell in every 30 to 300 can generate detectable levels of IL 2. Mls-locus incompatibilities activate a higher fraction of cells than do differences at the H-2 complex. The responder cell is sensitive to anti-Thy-1.2 antibody and complement, and is highly enriched in Lyt-2- populations as compared to positively selected Lyt-2+ cells. The dilution curve obtained is consistent with the idea that IL 2 production depends on a single class of T cell. The amount of IL 2 produced in these limiting dilution cultures is unexpectedly high; about 30-fold greater than the levels predicted by extrapolation from conventional mixed lymphocyte cultures. These results suggest that conventional cultures may be very responsive to regulatory events that do not occur at very low responder cell concentrations. Thus, limiting dilution analyses will provide insights into the effects of drugs, age, subset manipulation, etc., on the IL 2-producing cell itself, information that cannot be provided by conventional methods alone.
D000818:Animals; D002452:Cell Count / Q000379:methods; D002454:Cell Differentiation; D004306:Dose-Response Relationship, Immunologic; D007376:Interleukin-2 / Q000096:biosynthesis / Q000378:metabolism*; D007519:Isoantigens / Q000276:immunology; D008213:Lymphocyte Activation; D008222:Lymphokines / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008808:Mice, Inbred CBA; D008811:Mice, Inbred DBA; D010641:Phenotype; D011817:Rabbits; D013154:Spleen / Q000378:metabolism; D013601:T-Lymphocytes / Q000145:classification / Q000166:cytology / Q000378:metabolism*
D007376:Interleukin-2; D007519:Isoantigens; D008222:Lymphokines