An improved method for the growth of human lymphocyte colonies
Alder, S.J.; Morley, A.A.; Seshadri, R.S.
Experimental Hematology 9(10): 972-977
1981
ISSN/ISBN: 0301-472X PMID: 7308326 Document Number: 170534
An improved method was developed for growth of human lymphocyte colonies in agar. The method is a single-step assay involving the addition of irradiated lymphocytes which do not themselves form colonies, but which stimulate a small number of non-irradiated cells to do so. Over the range 0-1 .times. 105 target lymphocytes, there is a linear relationship between number of lymphocytes cultured and number of colonies produced. Technical factors which influence colony growth include osmolality of the medium, fetal calf serum concentration, agar concentration, PHA type and concentration, and 2-mercaptoethanol. Colony formation is not affected by using homologous irradiated cells from an individual of the same age. For normal individuals aged 11-45, colonies of 20-200 cells were grown to a plating efficiency of 2.18%, with a range 1.28-4.00%. The advantages of the method are its simplicity, the linear relationship between cells plated and colonies formed, which enables the method to be used as a quantitative assay, and the ability to determine whether an observed abnormality is due to an abnormality of colony forming cells or of irradiated stimulating cells.