The molecular nature of platelet receptors for quinine- and quinidine- dependent antiplatelet antibodies (Q.Ab and Qd.Ab) was studied by immunoblotting. One Q.Ab caused quinine-dependent IgG binding to platelet proteins with molecular weights (mol wts) of 174 Kd and 93 Kd and another to only a 93-Kd protein. A third Q.Ab caused binding to 174- , 140-, 93-, and 57-Kd proteins, while a fourth Q.Ab and a Qd.Ab caused IgG binding to 174- and 18-Kd proteins. Using platelets from patients with Glanzmann's thrombasthenia or Bernard Soulier syndrome and purified GPIIIa, these proteins were shown to be GPIb, GPIIb, GPIIIa, GPIX, and an unidentified 57-Kd protein missing in Bernard Soulier syndrome. Binding to the 93-Kd protein was independent of the PIA1 antigen. Absorption of one Q.Ab with Glanzmann's thrombasthenia platelets revealed different populations of antibodies with different specificities within the one patient. Thus Q.Ab and Qd.Ab are heterogeneous and may be directed toward different epitopes on major platelet glycoproteins.
SUMMARYImmunoblotting of platelets that have been subjected to SDS-PAGE has revealed thjit sera from normal individuals contain IgG which binds to many platelet components. This binding was seen with aulologous and heterologous platelets using serum of males and of nulliparous females who had not received blood transfusions. Although binding patterns of difTcrcnl sera were not identical, almost all sera caused IgG binding to platelet components of 87-90 kD, 140 kD (identified as vinculin) and 220-240 kD (tentatively identiiied as talin and actin-binding protein). Purified TgG showed the same binding pattern as whole seruni and F(ab')2 fragments retained iheir ability Lo bind lo many eomponents. The litre of IgG binding in serum was 1:50-1600 while thai of alloantibodies to the Pl'^' antigen was 1:3200. IgG bijiding components were not secreted when platelets were stimulated and were rarely associated wiih isolated membranes, bul were located either in platelet cyloplasm or cyloskcletons. IgG binding was decreased by absorbing sera with lysed platelets or isolated cytoskeletons. but only slightly with intact ptaleiels. Microaffinity purification oflgG which formed a major band on immunoblots showed that it was antibody with specificity for vinculin or its degradation products. These findings suggest that normiil sera contain naturally occurring IgG antibodies with specificity for intracellular platelet antigens ajid that in some cases their titre approaches thai of antibodies of pathological significance.
The molecular nature of platelet receptors for quinine- and quinidine- dependent antiplatelet antibodies (Q.Ab and Qd.Ab) was studied by immunoblotting. One Q.Ab caused quinine-dependent IgG binding to platelet proteins with molecular weights (mol wts) of 174 Kd and 93 Kd and another to only a 93-Kd protein. A third Q.Ab caused binding to 174- , 140-, 93-, and 57-Kd proteins, while a fourth Q.Ab and a Qd.Ab caused IgG binding to 174- and 18-Kd proteins. Using platelets from patients with Glanzmann's thrombasthenia or Bernard Soulier syndrome and purified GPIIIa, these proteins were shown to be GPIb, GPIIb, GPIIIa, GPIX, and an unidentified 57-Kd protein missing in Bernard Soulier syndrome. Binding to the 93-Kd protein was independent of the PIA1 antigen. Absorption of one Q.Ab with Glanzmann's thrombasthenia platelets revealed different populations of antibodies with different specificities within the one patient. Thus Q.Ab and Qd.Ab are heterogeneous and may be directed toward different epitopes on major platelet glycoproteins.
SummaryReactivity of quinine- and quinidine-dependent antiplatelet antibodies has been compared in platelet-rich-plasma (PRP) from normal donors and from patients with von Willebrand’s disease (vWd). One quinine-dependent antibody (Q. Ab) caused platelet aggregation and [14C] serotonin release with only 7 of 12 normal donors, while another Q. Ab and a quinidine-dependent antibody (Qd. Ab) caused aggregation and release with all 12. Drug- dependent IgG binding and PF 3 availability induced by the antibodies were, however, comparable in all donors. Differences in responsiveness were associated with platelets and not plasma. vWd platelets showed normal drug-dependent IgG binding, but decreased aggregation and serotonin release to most drug- dependent antibodies. Responsiveness was not restored by purified vWf:Ag, but, in one case, was corrected by normal plasma or cryoprecipitate. Drug-dependent binding of the Q. Ab which caused variable responsiveness in normals was to the same platelet antigens (GPIb and GPIIIa) in both normal and vWd platelets and did not require plasma components. Reduced PF 3 availability was seen with some antibodies in some vWd patients. Plasma from two of these patients inhibited aggregation of normal platelets to Q. Ab and one of these inhibited aggregation to ADP. Antiplatelet antibodies were detected in these two plasmas by ELISA. Thus some Q. Ab produce different responses with platelets from different donors. In vWd, reduced responsiveness to Q.Ab and Qd. Ab may result from production of inhibitory antiplatelet antibodies.
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