TCR/CD3 determines T-cell expression of CD91 and TSP-1. (A) Anti-CD3 Abs (0.125 μg/mL) coated on fibronectin or poly-L-lysine substrata augment CD91 expression on the surface of AF24 cells as demonstrated by quantitative immunocytochemistry and immunoprecipitation of biotinylated cells. (B) Anti-CD3 antibodies (0.125μg/mL) postcoated on a fibronectin-coated surface trigger cell-surface TSP-1 in freshly purified blood lymphocytes. (C) Anti-CD3 antibodies (0.125 μg/mL) postcoated on a fibronectin-coated surface inhibit TSP turnover as shown by abrogation of the inhibitory effect of brefeldin A (10 μg/mL) on TSP expression. For immunocytochemistry, the cells were fixed in PFA after 15 minutes and the expression of TSP-1 and CD91 was determined by quantitative immunocytochemistry. For immunoprecipitation, the adherent cells were biotinylated and subsequently immunoprecipitated and separated on a 4% to 20% SDS-PAGE gel. One representative experiment of 3 to 7 independent experiments is shown. Error bars: SEM. Statistics: Mann-Whitney U test (B-C). Bars: 20 μm.