The selective growth of human T lymphocyte colonies from whole blood in a semisolid culture system.
Human T lymphocyte colonies may be selectively grown from whole blood in a single phase semisolid culture system following stimulation with PHA-P, Con-A, or PPD. This technique eliminates the requirement for gradient-enriched lymphocyte fractions, and provides a sensitive system for the study of T lymphocyte progenitors that more closely approximates the in vivo milieu. Whole blood colonies were composed of lymphoblasts and mature lymphocytes. Individual colony cells, identified as T lymphocytes, lacked lipase and specific esterase activity, formed E rosettes, did not phagocytize latex beads, and were largely ANAE positive. Whole blood was plated at a final concentration of 3%. Optimal mitogen/antigen concentrations were 125 microgram Con-A, 80 microgram PHA-P and 50 microgram PPD/ml culture media. Peak colony growth occurred between days 7 and 8. Colony formation increased as a power function over a wide range of cell concentrations (5 x 10(3)-5 x 10(4) lymphocytes plated). Maximal whole blood colony formation occurred when 5 x 10(4)-10(5) lymphocytes were plated. There was a significant increase in the cloning efficiency using whole blood as compared to gradient-separated cells. This method has wide application for the study of radiation effects, lymphocyte alterations in various disease states, antigen recognition, and the induction and amplification of T cell function.
D000293:Adolescent; D000328:Adult; D002478:Cells, Cultured; D003208:Concanavalin A / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008875:Middle Aged; D010835:Phytohemagglutinins / Q000494:pharmacology; D011043:Pokeweed Mitogens / Q000494:pharmacology; D013601:T-Lymphocytes / Q000166:cytology*; D014373:Tuberculin
D010835:Phytohemagglutinins; D011043:Pokeweed Mitogens; D014373:Tuberculin; D003208:Concanavalin A