Ruan C G, Du X P, Xi X D, Castaldi P A, Berndt M C
Blood. 1987 Feb;69(2):570-7.
A new monoclonal antibody (MoAb), SZ 2, reactive with the human platelet glycoprotein Ib complex has been produced by the hybridoma technique. SZ 2 immunoprecipitated the components of the glycoprotein Ib complex, glycoprotein Ib and glycoprotein IX, from Triton-X-100-solubilized, periodate-labeled platelets. Western blot analysis indicated that the epitope for SZ 2 was on the alpha-subunit of glycoprotein Ib. Scatchard analysis of SZ 2 binding to formaldehyde-fixed, washed platelets revealed a single class of binding sites with Kd = 6.6 +/- 3.3 X 10(-10) mol/L and 15,200 +/- 4,100 binding sites per platelet (mean +/- SD, n = 10). Intact antibody and its purified (Fab')2 fragments not only inhibited the ristocetin-dependent binding of von Willebrand factor to platelets and ristocetin-induced platelet agglutination but also inhibited platelet aggregation induced by Type I collagen and platelet-activating factor (PAF). SZ 2 inhibited platelet serotonin and beta-thromboglobulin release in response to these stimuli and also platelet thromboxane A2 formation in response to ristocetin and collagen. SZ 2 was without effect on platelet aggregation or release in response to other platelet stimuli such as ADP, thrombin, or arachidonic acid. The inhibition by SZ 2 of collagen- and PAF-induced platelet aggregation is surprising in that Bernard-Soulier syndrome platelets, which lack the glycoprotein Ib complex, respond normally to both these stimuli. SZ 2 was unreactive toward Bernard-Soulier syndrome platelets, as evaluated by fluorescence-associated cell sorting, and had no effect on the collagen- and PAF-induced aggregation of Bernard-Soulier syndrome platelets. The combined results suggest that the inhibition by SZ 2 of collagen- and PAF-induced aggregation of normal platelets is steric and are consistent with the glycoprotein Ib complex and the platelet collagen and PAF receptor(s) being adjacent in the human platelet plasma membrane.
通过杂交瘤技术制备了一种新的与人血小板糖蛋白 Ib 复合物反应的单克隆抗体（MoAb）SZ 2。SZ 2 从经 Triton-X-100 溶解、高碘酸盐标记的血小板中免疫沉淀糖蛋白 Ib 复合物的成分，即糖蛋白 Ib 和糖蛋白 IX。蛋白质印迹分析表明，SZ 2 的表位位于糖蛋白 Ib 的α亚基上。对 SZ 2 与甲醛固定、洗涤后的血小板结合进行 Scatchard 分析显示，存在一类结合位点，解离常数（Kd）为 6.6±3.3×10⁻¹⁰ mol/L，每个血小板有 15200±4100 个结合位点（平均值±标准差，n = 10）。完整抗体及其纯化的（Fab'）₂片段不仅抑制血管性血友病因子与血小板的瑞斯托霉素依赖性结合以及瑞斯托霉素诱导的血小板凝集，还抑制 I 型胶原和血小板活化因子（PAF）诱导的血小板聚集。SZ 2 抑制这些刺激引起的血小板 5-羟色胺和β-血小板球蛋白释放，以及对瑞斯托霉素和胶原的反应中血小板血栓素 A₂ 的形成。SZ 2 对其他血小板刺激物（如 ADP、凝血酶或花生四烯酸）诱导的血小板聚集或释放没有影响。SZ 2 对胶原和 PAF 诱导的血小板聚集的抑制作用令人惊讶，因为缺乏糖蛋白 Ib 复合物的伯纳德-索利尔综合征血小板对这两种刺激的反应正常。通过荧光相关细胞分选评估，SZ 2 与伯纳德-索利尔综合征血小板无反应，且对伯纳德-索利尔综合征血小板的胶原和 PAF 诱导的聚集没有影响。综合结果表明，SZ 2 对正常血小板胶原和 PAF 诱导的聚集的抑制是空间位阻性的，并且与糖蛋白 Ib 复合物以及人血小板质膜中相邻的血小板胶原和 PAF 受体一致。
