Interaction of AP-2, a monoclonal antibody specific for the human platelet glycoprotein IIb-IIIa complex, with intact platelets.

D Pidard, RR Montgomery, JS Bennett… - Journal of Biological …, 1983 - Elsevier
D Pidard, RR Montgomery, JS Bennett, TJ Kunicki
Journal of Biological Chemistry, 1983Elsevier
A murine monoclonal antibody, designated AP-2, reacts specifically with the complex formed
by human platelet membrane glycoproteins IIb and IIIa, but does not react at all with the
individual glycoproteins. Purified AP-2 covalently coupled to Sepharose CL4B was used as
an immunoadsorbent column to purify the IIb-IIIa complex from a preparation of Triton X-100-
solubilized human platelet proteins. Radioiodinated AP-2 was shown to bind to a single
class of sites, with 57,400+/-9,700 molecules bound per cell (mean+/-SD) at saturation and a …
A murine monoclonal antibody, designated AP-2, reacts specifically with the complex formed by human platelet membrane glycoproteins IIb and IIIa, but does not react at all with the individual glycoproteins. Purified AP-2 covalently coupled to Sepharose CL4B was used as an immunoadsorbent column to purify the IIb-IIIa complex from a preparation of Triton X-100-solubilized human platelet proteins. Radioiodinated AP-2 was shown to bind to a single class of sites, with 57,400 +/- 9,700 molecules bound per cell (mean +/- S.D.) at saturation and a dissociation constant (Kd) of 0.64 +/- 0.15 nM (mean +/- S.D.). Binding could not be readily reversed even after a 1-h incubation with a 100-fold excess of cold antibody. AP-2 inhibits ADP-induced binding of radiolabeled fibrinogen to gel-filtered platelets in a noncompetitive fashion, consistent with the previous observation that AP-2 also inhibits the aggregation of platelets in plasma induced by a number of physiologic agonists, including adenosine diphosphate, epinephrine, collagen, thrombin, and arachidonic acid. Using AP-2, we have obtained evidence that the IIb-IIIa complex exists in the membrane of intact nonstimulated platelets and that complex integrity is not affected by external calcium ion concentration.
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