Human immunoglobulin biosynthesis in a serum-free medium.

We have previously reported that human peripheral blood lymphocytes are able to survive and undergo blastogenesis when cultured under serum-free conditions. Now we report that peripheral blood lymphocytes are capable of producing immunoglobulins in response to polyclonal activation during culture for 10 days in medium consisting of one part Ham's F-12 and one part Iscove's modified Dulbecco's medium supplemented with sodium bicarbonate, bovine crystalline insulin, human transferrin, 2-mercaptoethanol, and bovine serum albumin. Immunoglobulin levels in tissue culture supernatants obtained on day 10 were determined by enzyme-linked immunosorbent assay and were analyzed using a new computer program. Pokeweed mitogen was used as the polyclonal activator, and it stimulated immunoglobulin biosynthesis as well in serum-free medium as in medium that contained fetal calf serum. Immunoglobulin 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 X 10(5) cells in 200 microliters of medium per 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.