Immunoelectron microscopic study of fibrinogen-binding sites on stimulated platelets.

Recently, several investigators reported that fibrinogen-binding sites on the platelet membrane can be exposed following stimulation with ADP or other aggregating agents. We observed fibrinogen-binding sites by immunoelectron microscopy using monoclonal anti-GPIIb-IIIa antibody (NNKY1-32), monoclonal anti-GPIb antibody (NNKY5-5), and anti-fibrinogen antibody. Binding of two monoclonal antibodies was visualized by ferritin particles as a tracer, and binding of anti-fibrinogen antibody by gold particles. Both of ferritin particles which recognize GPIIb-IIIa (F1) and GPIb (F2) were distributed diffusely on the surface of unstimulated platelets. In ADP-stimulated platelets and shape-changed platelets by stimulation with collagen, F1 significantly increased in number, but F2 were distributed similarly in unstimulated platelets. On the other hand, in aggregated platelets by stimulation with collagen, F1 significantly decreased and gold particles which recognize fibrinogen were distributed as clusters between adherent platelets. Our morphological analysis has confirmed the hypothesis that fibrinogen-binding sites on the platelet surface are exposed by stimulation with ADP and collagen. It is seemed that such effect following stimulation with collagen was caused independently of released ADP.