Bessos H, Murphy W G
Edinburgh and South-East Scotland Regional Blood Transfusion Service, Royal Infirmary, UK.
Thromb Res. 1990 Aug 1;59(3):497-507. doi: 10.1016/0049-3848(90)90410-e.
A new competitive binding enzyme-linked immunosorbent assay (CB ELISA) for glycocalicin (GC) was developed using GC coated wells and a monoclonal antibody (MAb) to glycoprotein Ib (AN51). The principal stages of the CB ELISA consisted of coating the plate with GC extract overnight, blocking with 3% BSA, incubating the wells with test or standard sample dilution and AN51, and a final incubation with horseradish peroxidase-conjugated goat anti-mouse IgG. Serial dilutions of purified GC, starting in 2% BSA, yielded standard curves which were linear between 10 and 0.4 micrograms/ml. Parallel curves were obtained for platelet concentrate supernatants and for citrated plasma. We used the ELISA to measure GC levels in platelet concentrates during storage. The results indicated that soluble GC increased progressively during storage from 3.3 to 6.7 micrograms/ml, while GC levels in platelet-poor plasma remained at 1.9-2.2 micrograms/ml. These results show that the new CB ELISA is a simple and short assay for the direct measurement of GC in plasma solutions, and may be of use in clinical studies.
利用包被有糖钙蛋白（GC）的微孔板和针对糖蛋白Ib的单克隆抗体（MAb，AN51），开发了一种用于检测糖钙蛋白（GC）的新型竞争性结合酶联免疫吸附测定法（CB ELISA）。CB ELISA的主要步骤包括：用GC提取物包被微孔板过夜，用3%牛血清白蛋白封闭，将微孔板与测试样品或标准样品稀释液以及AN51一起孵育，最后与辣根过氧化物酶偶联的山羊抗小鼠IgG一起孵育。以2%牛血清白蛋白为起始浓度的纯化GC系列稀释液产生了标准曲线，该曲线在10至0.4微克/毫升之间呈线性。血小板浓缩物上清液和枸橼酸盐血浆获得了平行曲线。我们使用该ELISA法测量储存期间血小板浓缩物中的GC水平。结果表明，可溶性GC在储存期间从3.3微克/毫升逐渐增加至6.7微克/毫升，而血小板贫乏血浆中的GC水平保持在1.9 - 2.2微克/毫升。这些结果表明，新型CB ELISA是一种用于直接测量血浆溶液中GC的简单且耗时短的检测方法，可能在临床研究中有用。
