Gawryl M S, Simon M T, Eatman J L, Lint T F
J Immunol Methods. 1986 Dec 24;95(2):217-25. doi: 10.1016/0022-1759(86)90409-6.
A sensitive and simple enzyme-linked immunoabsorbent assay (ELISA) has been developed to measure the terminal complement complex (TCC) in solution. Commercially available antibodies to the native complement (C) components C5 and C9 were used in a double antibody sandwich technique sensitive enough to detect 0.3 microgram/ml of purified TCC. The TCC was not detected in normal human serum (NHS) nor was it generated when sera from patients with a genetic deficiency of functional C5, C7, C8 beta or C9 were activated with cobra venom factor (CVF). If the C8 beta deficient serum was reconstituted with the C8 beta chain and incubated with CVF, TCC were formed and detected by the assay. In in vitro experiments, the TCC was detected in NHS activated by either the classical or alternative pathway even when there was no measurable consumption of C5, C8 or C9. In addition, adaptation of a detergent extraction procedure permitted the quantitation by the assay, of TCC which were generated on sensitized sheep erythrocyte membranes. Experiments to test sample handling conditions showed no generation of TCC in NHS after four freeze/thaw cycles and spontaneous formation only if NHS had been incubated at 37 degrees C for 48 h. The TCC in zymosan-activated NHS were stable at 37 degrees C for 1 week. Patients with C activation associated diseases such as SLE and rheumatoid arthritis had increased levels of TCC that correlated with positive clinical tests for inflammation, even though C levels were normal when measured by routine techniques. These results suggest that this ELISA will provide a valuable tool for studying the role of C in the pathogenesis of C-mediated diseases and in examining the mechanism of tissue injury in in vitro experimental systems.
已开发出一种灵敏且简单的酶联免疫吸附测定法（ELISA）来测量溶液中的末端补体复合物（TCC）。在一种双抗体夹心技术中使用了针对天然补体（C）成分C5和C9的市售抗体，该技术灵敏到足以检测0.3微克/毫升的纯化TCC。在正常人血清（NHS）中未检测到TCC，并且当用眼镜蛇毒因子（CVF）激活功能性C5、C7、C8β或C9基因缺陷患者的血清时也未产生TCC。如果用C8β链重建C8β缺陷血清并与CVF一起孵育，则会形成TCC并通过该测定法检测到。在体外实验中，即使没有可测量的C5、C8或C9消耗，在通过经典途径或替代途径激活的NHS中也检测到了TCC。此外，一种去污剂提取程序的改进使得可以通过该测定法定量在致敏绵羊红细胞膜上产生的TCC。测试样品处理条件的实验表明，经过四个冻融循环后，NHS中未产生TCC，只有当NHS在37℃孵育48小时时才会自发形成TCC。酵母聚糖激活的NHS中的TCC在37℃下稳定1周。患有C激活相关疾病（如系统性红斑狼疮和类风湿性关节炎）的患者TCC水平升高，这与炎症的阳性临床检测结果相关，尽管通过常规技术测量时C水平正常。这些结果表明，这种ELISA将为研究补体在补体介导疾病的发病机制中的作用以及在体外实验系统中研究组织损伤机制提供有价值的工具。
