Fetal calf serum in growth medium obscures the detection of early anticardiolipin antibody secreting clones.
We have examined the effect of fetal calf serum (FCS) on the detection of early anticardiolipin antibody (ACL) secreting clones, using two well-established human IgG clones, LJ1 and AH2. By plating cells at 50/well and growing both clones simultaneously in standard growth (SG) medium containing 10% FCS, and in serum-free (SF) medium, we were able to measure by ELISA the total IgG and ACL levels in the supernatants. The mean OD values (x 1000) against cardiolipin for both LJ1 and AH2 were significantly higher for clones grown in SF than in SG medium: 331 OD units vs. 172, and 275 OD units vs. 166 respectively (p < 0.001). Importantly, the number of wells in which the OD value was > 0.25 units above background was: for LJ1 in SG only 3/36 vs. 30/36 in SF; similarly, for AH2 1/36 in SG vs. 22/36 in SF. In comparison, the total IgG assay using an OD value > 0.7 units above background, detected immunoglobulin secretion in all but one of the wells. We conclude that in ELISA procedures FCS in SG medium competes with solid phase cardiolipin for antibody binding. We suggest that these antibodies are binding to phospholipid from microvesicles found in FCS. We recommend that minimally 'positive' clones on testing should be re-tested and, if necessary, switched to SF medium in order to prevent such clones from being discarded prematurely.
Menon
S
S
Department of Medicine, University College London Medical School, UK.
Isenberg
D A
DA
D000818:Animals; D017153:Antibodies, Anticardiolipin / Q000032:analysis* / Q000096:biosynthesis / Q000276:immunology; D001769:Blood; D002478:Cells, Cultured; D002999:Clone Cells; D003470:Culture Media / Q000737:chemistry*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D008180:Lupus Erythematosus, Systemic / Q000276:immunology; D051379:Mice
D017153:Antibodies, Anticardiolipin; D003470:Culture Media