Development of an ELISA for quantification of human protein S in cell culture fluids using commercial polyclonal antisera.
An enzyme-linked immunosorbent assay (ELISA) was developed to measure protein S antigen released into cell culture fluids. We used readily available commercial polyclonal antisera to develop the assay. This assay was sensitive with a detection limit of about 0.086 ng/ml. Between-assay precision (coefficient of variation) at levels of 0.2, 1.1, and 13.9 ng/ml was 14%, 15%, and 11% respectively. Specificity and accuracy were demonstrated from the use of: 1) culture fluids from 3-primary endothelial cell cultures and 7-cell lines known to constitutively produce protein S; 2) 2-cell lines not synthesizing protein S; and 3) from selected samples of normal and protein S deficient plasma. The ELISA described here was about 12-fold more sensitive and 40-fold more cost effective when compared to a commercial ELISA kit. Thus the assay provided a sensitive, specific, precise and economical method useful for the measurement of the nanogram amounts of protein S commonly encountered in cell culture fluids.
Phillips
D J
DJ
Centers for Disease Control and Prevention, U.S. Department of Health and Human Services, Atlanta, Georgia 30333.
Evatt
B L
BL
Hooper
W C
WC
D002460:Cell Line; D002478:Cells, Cultured; D017077:Culture Media, Conditioned / Q000737:chemistry*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007106:Immune Sera / Q000276:immunology*; D017293:Protein S / Q000032:analysis* / Q000276:immunology; D012680:Sensitivity and Specificity
D017077:Culture Media, Conditioned; D007106:Immune Sera; D017293:Protein S