Impaired P-selectin expression on Fg^−/− platelets reflects decreased platelet P-selectin content. (A) P-selectin levels in Fg^+/+, Fg^+/−, and Fg^−/− mouse platelets were examined via Western blot. Actin was used as a loading control. A decrease in P-selectin was observed in Fg^−/− compared with wild-type and Fg^+/− platelets (left panel). Total platelet P-selectin levels were significantly decreased in Fg^−/− platelets compared with wild-type platelets, as measured with ELISA (right panel; n = 3; **P < .001). (B) Total platelet PF4 and TSP-1 levels in Fg^+/+ and Fg^−/− mouse platelets were examined via Western blot. Actin was used as a loading control. No significant difference was observed. (C) Total platelet P-selectin levels in platelets of a healthy donor, the homozygous patient, and his mother (Het) were examined via Western blot. Vinculin was used as a loading control. A decrease in P-selectin was observed in the patient's platelets. A slight decrease was also observed in platelets of the heterozygous parent. No significant difference in the levels of TSP-1 was observed. (D) Total platelet P-selectin levels were examined via Western blot (left panel) and ELISA (right panel). Plasma Fg transfusion restored total P-selectin levels in Fg^−/− platelets 1, 2, and 4 days after transfusion. A significant decrease in Fg^−/− compared with wild-type platelets was observed via ELISA (n = 3; ##P < .001). Transfusion of plasma Fg significantly increased total platelet P-selectin levels 1, 2, and 4 days after transfusion compared with untreated Fg^−/− platelets (shaded) (n = 3; **P < .001). The immunoblots in each panel were developed from the same PVDF (A, C-D) or nitrocellulose (B) membrane and the experiments were repeated at least 3 times.