Demonstration of the Requirements for Cholesterol or Low Density Lipoprotein in the Antibody Response of Murine Lymphocytes in Serum-free Culture

Abstract

We report the essential role of lipid in the development of a serum-free culture medium that supports the primary antibody response by murine lymphocytes in vitro. In our preceding paper, we have reported that β-cyclodextrin (β-CD) is effective as a serum-substitute in murine lymphocyte cultures [H. Ohmori and I. Yamamoto, Eur. J. Immunol., 17, 79 (1987)]. At first, we employed RPMI-1640 that was supplemented with fatty acid-free albumin, insulin, transferrin, and β-CD as the basal serum-free medium (BSF medium). BSF medium supported the antibody response less than 30 % as efficiently as 10% fetal calf serum (FCS)-containing medium. When ether-extracts of FCS (FCS ex.) was added to BSF medium, the antibody response was enhanced by approximately 2-fold. FCS ex. was purified by silica gel thin-layer chromatography. The analyses of the purified active component by high performance liquid chromatography and mass spectrometry revealed it to be cholesterol. Authentic cholesterol or low density lipoprotein enhanced the antibody response as efficiently as FCS ex. in BSF medium. The same level of the antibody response as that in 10 % FCS-containing medium was attained when low density lipoprotein was added to BSF medium in combination with i -a la nine, putrescine, and pyruvate.