T-lymphocyte colony formation of murine spleen cells in a one-stage micro agar culture.
A one-stage agar culture technique for stimulation of T-lymphocyte colony-forming units (TL-CFU) derived from murine spleen cells is described. The cultures are in glass capillaries in a volume of 30 microliter. Both, phytohemagglutinin (PHA) and concanavalin A (Con A) stimulate growth of TL-CFU. The technique does not require prestimulation of lymphocytes in liquid culture or addition of growth factor(s). The most important requirements for optimal colony growth are a low concentration of agar (less than 0.1%) and the appropriate number of seeded cells (60 X 10(3) per culture). This one-stage micro agar culture is economical and easily handled. Many cultures can be prepared and rapidly evaluated. The plating efficiency is about 10 colonies per 10(4) nucleated spleen cells. The T-cells character of the TL-CFU was demonstrated by treatment of the spleen cells with a monoclonal anti-Thy-1.2 antibody (anti-Thy-1.2) and complement before assaying for colony growth and also by staining the cells in the cultures with anti-Thy-1.2-FITC. Growth characteristics of TL-CFU indicate that cell interactions are operative during activation and proliferation. The nature of the cells cooperating in this system is discussed.
D000362:Agar; D000818:Animals; D002455:Cell Division / Q000187:drug effects / Q000528:radiation effects; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D004247:DNA / Q000096:biosynthesis; D005260:Female; D007700:Kinetics; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010600:Pharmacology / Q000379:methods; D012313:RNA / Q000096:biosynthesis; D013154:Spleen / Q000166:cytology*; D013601:T-Lymphocytes / Q000276:immunology*; D013997:Time Factors; D014965:X-Rays
D012313:RNA; D000362:Agar; D004247:DNA