Induction of CD69 expression by T-cell interaction with immobilized ICAM-1. (A) PBLs were added to wells precoated with BSA (1%), rsICAM-1 (20 μg/mL), rsVCAM-1 (20 μg/mL), FN (20 μg/mL), and PLL (40 μg/mL). After 18 hours of incubation at 37°C, CD69 expression was measured in CD3⁺ cells by two-color flow cytometry. The results of three independent experiments are depicted as arithmetic mean ± 1 SD. *, P < .01 compared with BSA (Mann-Whitney U test). (B) ICAM-1 dose-dependent T-cell activation. PBLs were incubated on wells precoated with different concentrations of rsICAM-1 (▪) or rsVCAM-1 (•). After 18 hours, CD69 expression was analyzed in CD3⁺ cells. The results of four independent experiments are represented as arithmetic mean ± 1 SD. (C) Kinetics analysis of CD69 T-cell expression induced by rsICAM-1 and IL-15. PBLs were incubated on BSA (1%) or rsICAM-1 (20 μg/mL) precoated wells, and then IL-15 (5 ng/mL) was added where indicated. After 4, 16, 24, 40, 62, and 120 hours, cells were collected and CD69 expression was measured in CD3⁺ cells. The results of three independent experiments are shown as arithmetic mean ± 1 SD. (○), BSA; (•), ICAM-1; (□), BSA + IL-15; (▪), ICAM-1 + IL-15.