Dancis A, Ehmann C, Ferziger R, Grima K, Karpatkin S
Department of Medicine, New York University Medical School 10016.
Blood. 1988 Apr;71(4):1056-61.
The mol wt of the glycoprotein(s) carrying the PLA1 antigen was examined on platelets, megakaryocytes and endothelial cells by immunoblotting with a human polyclonal anti-PLA1 antibody (BE), as well as on four different monoclonal antibodies (MoAbs; DEK-1, DEK-2C, DEK-10, and DEK-16) raised against GPIIIa, the 100,000-mol wt platelet glycoprotein known to carry the PLA1 antigen. BE reacted with PLA1 positive but not with PLA1 negative platelets. DEK-1 reacted strongly with PLA1 positive platelets but weakly with PLA1 negative platelets. The remaining three MoAbs reacted equally with PLA1 positive as well as negative platelets. BE, DEK-1, DEK-10, and DEK-16 reacted with a 120,000- as well as 100,000-mol wt band on immunoblot of PLA1 positive platelets. The 120,000-mol wt band copurified with affinity purified 100,000-mol wt GPIIIa. Megakaryocytes had a prominent 120,000- as well as 105,000-mol wt band that reacted with BE on immunoblot (the 100,000-mol wt band was not detectable). Umbilical cord endothelial cells from presumed PLA-positive infants had a prominent 100,000-mol wt band that reacted with BE, DEK-16, and DEK-1 (the 120,000-mol wt band was not visualized). The 120,000- and 100,000-mol wt PLA1-positive bands could be digested with proteolytic enzymes to 55,000- to 65,000-mol wt-resistant fragments that retain PLA1 epitopes. Further digestion with endoglycosidase-H lowered the apparent mol wt by approximately 2,000 to 6,000 daltons without affecting PLA1 reactivity. We conclude that the PLA1 antigen is present on a 120,000- as well as 100,000-mol wt glycoprotein of platelets and megakaryocytes, a 105,000-mol wt band of megakaryocytes, and a 100,000-mol wt glycoprotein of endothelial cells. We postulate that the 120,000-mol wt glycoprotein, which shares three or more epitopes with the 100,000-mol wt GPIIIa, may be a post-translational precursor of this species.
通过用一种人源多克隆抗PLA1抗体（BE）进行免疫印迹，以及用针对已知携带PLA1抗原的100,000道尔顿分子量血小板糖蛋白GPIIIa产生的四种不同单克隆抗体（MoAbs；DEK-1、DEK-2C、DEK-10和DEK-16），检测了携带PLA1抗原的糖蛋白在血小板、巨核细胞和内皮细胞上的分子量。BE与PLA1阳性血小板反应，但不与PLA1阴性血小板反应。DEK-1与PLA1阳性血小板强烈反应，但与PLA1阴性血小板反应较弱。其余三种单克隆抗体与PLA1阳性和阴性血小板反应程度相同。BE、DEK-1、DEK-10和DEK-16在PLA1阳性血小板的免疫印迹上与120,000道尔顿以及100,000道尔顿分子量的条带反应。120,000道尔顿分子量的条带与亲和纯化的100,000道尔顿分子量的GPIIIa共纯化。巨核细胞有一条突出的120,000道尔顿以及105,000道尔顿分子量的条带，在免疫印迹上与BE反应（100,000道尔顿分子量的条带未检测到）。推测为PLA阳性婴儿的脐带内皮细胞有一条突出的100,000道尔顿分子量的条带，与BE、DEK-16和DEK-1反应（120,000道尔顿分子量的条带未显现）。120,000道尔顿和100,000道尔顿分子量的PLA1阳性条带可以被蛋白水解酶消化成保留PLA1表位的55,000至65,000道尔顿分子量的抗性片段。用内切糖苷酶-H进一步消化使表观分子量降低约2,000至6,000道尔顿，而不影响PLA1反应性。我们得出结论，PLA1抗原存在于血小板和巨核细胞的120,000道尔顿以及100,000道尔顿分子量的糖蛋白上、巨核细胞的105,000道尔顿分子量的条带上以及内皮细胞的100,000道尔顿分子量的糖蛋白上。我们推测，与100,000道尔顿分子量的GPIIIa共有三个或更多表位的120,000道尔顿分子量的糖蛋白可能是该物种的翻译后前体。
