SEPARATION OF HUMAN THYMOCYTES AT DIFFERENT STAGES OF MATURATION BY CENTRIFUGATION ON A DISCONTINUOUS GRADIENT OF COLLOIDAL SILICA-GEL

  • 1 January 1981
    • journal article
    • research article
    • Vol. 3  (1) , 25-33
Abstract
Separation of human intrathymic cells on a discontinuous gradient of colloidal silica gel (Percoll) yielded 4 layers. The 1st (density 1.054 .+-. 0.002 g/ml) contained stromal cells and a few thymocytes positive for terminal deoxynucleotidyl transferase (Tdt), most of which were bound to large Tdt-negative non-T cells. The 2nd layer (1.069 .+-. 0.003 g/ml) contained large Tdt-negative thymocytes. The 3rd and 4th layers (1.075 .+-. 0.004 and 1.085 .+-. 0.003 g/ml, respectively) contained smaller T cells, more than 95% of which were Tdt-positive. Functional studies revealed that cells from the 1st layer had a high level of spontaneous [3H]thymidine uptake but did not respond to lectins; the 2nd layer responded to PHA [phytohemagglutinin], Con[concanavalin] A and allogeneic stimuli; the 3rd and 4th layers did not respond to lectin stimulation. Addition of cells from the 1st layer to the other layers at a 1 : 10 ratio significantly increased the mitogenic responses of the cells from the 2nd layer, but not of those from the 3rd or 4th layer. As in mice and rats, low-density intrathymic thymocytes in humans probably represent more mature T cells, the percentage of which increases with age.