Figure 1.
Figure 1. Adhesion and aggregation of platelets on fibrin and fibronectin-fibrin incorporation of soluble fibronectin or 70K fragment into platelet thrombi and effect of perfused fibronectin on the formation of platelet thrombi under shear conditions. (A) A suspension of platelets and red blood cells was perfused through a flow chamber opposed to a coverslip coated with fibrin or fibronectin-fibrin (FN-fibrin) at shear rates of 300 or 1250 s–1 for 5 minutes. Coverslips were taken out of the chamber and washed. Platelets were fixed, permeabilized, stained with rhodamine-phalloidin, and observed by epifluorescence microscopy. The small and large pictures were taken with × 10 (bar = 100 μm) and × 100 objectives (bar = 10 μm), respectively. (B-C) A suspension of platelets and red blood cells without soluble proteins (B) or premixed with 100 nM FITC-fibronectin or FITC-70K fragment (B-C) was perfused through a flow chamber opposed to a coverslip coated with human fibrin or fibronectin-fibrin, or mouse fibrin at a shear rate of 1250 s–1 for 5 minutes. (B) Platelets adhered to mouse fibrin-coated surfaces in the absence (top panel) or presence of FITC-fibronectin (bottom panel). Adherent platelets were fixed and incubated with a rabbit polyclonal antibody against human fibronectin and a mouse mAb against human fibrinogen, followed by incubation with FITC-conjugated anti–rabbit IgG and rhodamine-conjugated anti–mouse IgG antibody (top panel) or incubated only with mouse antifibrinogen antibody and rhodamine-conjugated antimouse IgG (bottom panel). The images in the lower panel were obtained by confocal microscopy and are from a 1-μm slice taken 4 μm above the plane of the coverslip. Bar = 10 μm. (C) Coverslips were taken out of the chamber after perfusion with FITC-fibronectin or FITC-70K fragment, and platelets were stained with rhodamine-phalloidin. Microscopy was performed as described. The small and large pictures were taken with × 10 (bar = 100 μm) and × 100 × (bar = 10 μm), respectively. The images of phalloidin-stained platelets should be compared with the bottom sections of panel A.

Adhesion and aggregation of platelets on fibrin and fibronectin-fibrin incorporation of soluble fibronectin or 70K fragment into platelet thrombi and effect of perfused fibronectin on the formation of platelet thrombi under shear conditions. (A) A suspension of platelets and red blood cells was perfused through a flow chamber opposed to a coverslip coated with fibrin or fibronectin-fibrin (FN-fibrin) at shear rates of 300 or 1250 s–1 for 5 minutes. Coverslips were taken out of the chamber and washed. Platelets were fixed, permeabilized, stained with rhodamine-phalloidin, and observed by epifluorescence microscopy. The small and large pictures were taken with × 10 (bar = 100 μm) and × 100 objectives (bar = 10 μm), respectively. (B-C) A suspension of platelets and red blood cells without soluble proteins (B) or premixed with 100 nM FITC-fibronectin or FITC-70K fragment (B-C) was perfused through a flow chamber opposed to a coverslip coated with human fibrin or fibronectin-fibrin, or mouse fibrin at a shear rate of 1250 s–1 for 5 minutes. (B) Platelets adhered to mouse fibrin-coated surfaces in the absence (top panel) or presence of FITC-fibronectin (bottom panel). Adherent platelets were fixed and incubated with a rabbit polyclonal antibody against human fibronectin and a mouse mAb against human fibrinogen, followed by incubation with FITC-conjugated anti–rabbit IgG and rhodamine-conjugated anti–mouse IgG antibody (top panel) or incubated only with mouse antifibrinogen antibody and rhodamine-conjugated antimouse IgG (bottom panel). The images in the lower panel were obtained by confocal microscopy and are from a 1-μm slice taken 4 μm above the plane of the coverslip. Bar = 10 μm. (C) Coverslips were taken out of the chamber after perfusion with FITC-fibronectin or FITC-70K fragment, and platelets were stained with rhodamine-phalloidin. Microscopy was performed as described. The small and large pictures were taken with × 10 (bar = 100 μm) and × 100 × (bar = 10 μm), respectively. The images of phalloidin-stained platelets should be compared with the bottom sections of panel A.

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