Platelet surface antigens: Analysis by monoclonal antibodies (original) (raw)
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European journal of biochemistry / FEBS, 1983
The specificity of five monoclonal antibodies (P1-P6) against platelet surface components was determined by immunoprecipitation of surface-labelled platelets from normal donors and patients with known platelet glycoprotein defects, followed by analysis by gel electrophoresis. Three (P2, P4 and P6) precipitated glycoproteins IIb and IIIa and, in addition, P2 precipitated glycoprotein Ia. P1 precipitated normally only glycoprotein Ib also Ia when the platelets were pretreated with neuraminidase. P3 precipitated principally glycoprotein Ia but glycoprotein Ib was also weakly precipitated. The effects of the monoclonals on platelet function were tested. P1 and P2 completely inhibited and P3 slightly inhibited thrombin-induced platelet aggregation. P2 also inhibited collagen-induced aggregation and partially inhibited ADP-induced platelet aggregation. P3, P4 and P6 partially inhibited ADP-induced platelet aggregation. None had any effect on ristocetin-induced aggregation despite P1 and P...
Biochemical characterization of antigens detected with anti-platelet monoclonal antibodies
Veterinary Immunology and Immunopathology, 1996
A panel of 18 monoclonal antibodies (mAbs) defined by the third workshop as specific for platelets, clustered in three preliminary groups: PC7, PC13 and PC27. These mAbs were further analysed by immunoprecipitation using extracts of iodinated and biotinylated peripheral blood mononucleated cells (PBMC) and platelets. We could confirm the existence of mAbs with specificities to WC9 (in PC7) and CD41/61 (in PC13). Two mAbs formed a new cluster, WC13, which may be homologous to human CD31 (in PC27). The influence of EDTA and thrombin on the expression of the different antigens on the platelet membrane was assessed by flow cytometry (FCM) analysis, as well as cross-reactivity with platelets from different species.
Journal of Clinical Investigation, 1987
Neonatal alloimmune thrombocytopenic purpura associated with a new platelet-specific alloantigen Pen' has been reported. We now provide direct evidence that the Pen' determinant is associated with glycoprotein (GP) IIIa, but that it is distinct from epitopes that define the pIA system. By ELISA wherein monoclonal antibodies specific for GPIIb (Tab) and specific for GPIIIa (AP3) were used to capture and hold antigens from a platelet lysate prepared under conditions that generate free GPIIb and GPIIIa, anti-Pen' reacted with GPIIIa held by AP3 but not with GPIIb held by Tab. In an alternative ELISA where purified GPIIIa from both PlAl-positive and PlAl-negative platelets were used individually as antigen, anti-Pen' reacted with both allelic forms of GPIIIa. By radioimmunoprecipitation, anti-Pen' precipitated a single surface-labeled membrane protein with electrophoretic characteristics in sodium dodecyl sulfate-polyacrylamide gels, under nonreduced or reduced conditions, identical to those of GPIIIa. By fluorocytometry, platelets from several donors, regardless of peA phenotype, bound an amount of anti-Pen' roughly equivalent to one-half that amount of anti-PlI' bound by PeAl homozygous (Al/Al) platelets and roughly equal to that amount of anti-P1Ae bound by p1Al heterozygous (Al/A2) platelets. Using platelets from donors typed homozygous for p1Al and Pen' in a quantitative indirect binding assay, 14-24,000 molecules of anti-Pen' and 41-51,000 molecules of anti-PlAl were bound per platelet at saturation. Anti-Pen' completely inhibited ADP-induced aggregation of Pen3-positive platelets, regardless of peA phenotype. These results indicate that the Pen' determinant is associated with GPIIIa but distinct from PA.
Evidence that a 210,000-molecular-weight glycoprotein (GP 210) serves as a platelet Fc receptor
Journal of Clinical Investigation, 1987
We previously identified a 210,000-mol-wt platelet glycoprotein (GP 210) that is missing from Bernard-Soulier platelets, and found that an antibody against GP 210 inhibits ristocetin-induced platelet agglutination. We now show by immunoblotting that GP 210 binds heat-aggregated rabbit and human IgG, as well as keyhole limpet hemocyanin (KLH)anti-KLH and ovalbumin (OA)-anti-OA immune complexes. Immune complex binding to GP 210 was preserved on chymotrypsin-treated platelets that lacked glycoprotein lb (GP Ib). In contrast, ristocetin-induced platelet agglutination resulted in disappearance of immunologically detectable GP 210 and loss of immune complex binding, even though GP lb remained intact. Purified Fc fragments inhibited binding of anti-GP 210 antibody to intact platelets and to GP 210 on immunoblots. The Fc fragments also blocked immune complex binding to GP 210. Conversely, anti-GP 210 antiserum and F(ab)2 fragments inhibited binding of fluoresceinlabeled Fc fragments to intact platelets. We conclude that GP 210 functions as a platelet Fc receptor.
Comparison of rat and human major platelet glycoproteins
1991
Using electrophoretic techniques combined with various detection methods we ascribed rat platelet glycoproteins (GPs) related to human GPIb, GPIIb and GPIIIa. 2. Rat GPIIb and GPIIIa crossreacted with rabbit polyclonal antibodies against human GPIIb and GPIIIa. 3. Species differences in glycosylation of GPs were shown using various lectins. 4. Molecular mass of rat major GPs was determined by SDS-PAGE (unreduced, reduced, kDa): GPIb (200, 166/26), GPIIb (140, 120/32) and GPIIIa (96, 106). 5. Isoelectric points of rat GPIIb and GPIIIa are shifted to the alkaline region as compared to human related GPs.