Adhesion and spreading of human platelets on immobilized TGFBIp. (A) Washed platelets were allowed to attach to immobilized TGFBIp for 1 hour with or without thrombin. Adhered platelets were detected by the acid phosphatase assay as described in “Platelet adhesion assay.” (B) Washed platelets were attached to BSA, fibronectin (FN), and TGFBIp (each 10 μg/mL) for 1 hour, and attached platelets were stained for anti-CD41 antibody and observed under a fluorescence microscope. The white bar represents 10 μm. The amounts of (C) platelet adhesion and (D) platelet spreading (pixels/cells) in the presence of TGFBIp, BSA, or fibrinogen were determined. Platelet adhesion was detected by the acid phosphatase assay, and spreading was detected by use of the MetaMorph program. (E) Integrin-dependent platelet adhesion on TGFBIp. After washed platelets were incubated with integrins α₅β₁ and α_IIbβ₃, or immunoglobulin G as a control (each 1 μg/mL) for 30 minutes, they were allowed to mix with immobilized TGFBIp (10 μg/mL) or fibronectin (FN; 10 μg/mL) for 1 hour. Platelet adhesion was detected by the acid phosphatase assay. (F) Integrin-dependent platelet spreading on TGFBIp. Platelet spreading (pixels/cell) on TGFBIp or fibronection (FN) was detected by use of the MetaMorph program. All results are shown as the means ± SD of 3 different experiments and ANOVA. *P < .05 compared with BSA or immunoglobulin G.