Quantitative estimation of HC FN production on differently coated surfaces.

HCs were cultured in serum-free medium for 24 hours in 96-well plates coated as indicated. FN production was quantified by a modified ELISA involving an anti-FN MoAb as the first layer, a biotin-conjugated goat anti-mouse antibody as second layer, and streptavidin-peroxidase as third layer. Peroxidase substrate was then added to each well for 15 minutes, and the product of the reaction was measured in a plate reader. The results expressed in optical density units show relative differences in FN on different surfaces (3 experiments involving cells from 3 patients; each measurement performed 5 times). The increased production rates, as compared with the controls, of FN on HA or anti-CD44 were both significant (P < .01).