HUMAN-IMMUNOGLOBULIN BIOSYNTHESIS IN A SERUM-FREE MEDIUM

  • 1 January 1984
    • journal article
    • research article
    • Vol. 103  (5) , 739-748
Abstract
It was previously reported that human peripheral blood lymphocytes are able to survive and undergo blastogenesis when cultured under serum-free conditions. Now it is reported that peripheral blood lymphocytes are capable of producing Ig in response to polyclonal activation during culture for 10 days in medium consisting of 1 part Ham''s F-12 and 1 part Iscove''s modified Dulbecco''s mediun supplemented with sodium bicarbonate, bovine crystalline insulin, human transferrin, 2-mercaptoethanol and bovine serum albumin. Ig levels in tissue culture supernatants obtained on day 10 were determined by enzyme-linked immunosorbent assay [ELISA] and were analyzed using a new computer program. Pokeweed mitogen [PWM] was used as the polyclonal activator, and it stimulated Ig biosynthesis as well in serum-free medium as in medium that contained fetal calf serum. Ig production in serum-free medium began on day 4 and continued through day 9 with peak production on days 5 to 8. Optimal conditions in serum-free medium were: PWM dose, 1:400 to 1:800; day of harvest, 10; and cell concentration, 3.5 .times. 105 cells in 200 .mu.l medium/well. Use of this serum-free medium will allow examination of the effects of various additives to tissue culture without concern for unknown factors, components, influences or potential interaction with serum.