SENESCENT FIBROBLASTS AS FEEDER CELLS FOR LYMPHOID-CELL CLONING

Senescent primary human skin fibroblasts were used as feeder layers fo r cloning lymphoid cells by limiting dilution, B-cells including mouse hybridoma cells, human multiple myeloma cells C1R and DIG, as well as an immortalized T-cell line (Jurkat cells) were cloned using this app roach, From heterogenous populations, homogeneous (clonal) populations were obtained and further analyzed. The major advantages of senescent fibroblasts as feeder cells are (i) the need to establish primary cul tures from experimental animals for preparing feeder cells is obviated ; (ii) the risk of contamination with infectious agents is diminished; (iii) primary skin fibroblasts are easily maintained in culture, can be kept at confluence for extended periods of time, and do not undergo spontaneous transformation; and (iv) lymphoid cells are readily separ ated from fibroblast monolayers, The use of senescent fibroblasts over comes limitations inherent with primary mouse cell cultures and irradi ated cells commonly used as feeder cells in cloning techniques. (C) 19 96 Academic Press, Inc.