Gorski J P
J Immunol Methods. 1981;47(1):61-73. doi: 10.1016/0022-1759(81)90257-x.
A method is described to quantitate complement fragment C4ai in human plasma, synovial fluid, and urine. Samples are first precipitated with 50% saturated (NH4)2SO4 to remove cross-reactive macromolecules C4 and pro-C4. Whereas greater than 97% of C4 is removed by this precipitation step, 88% of C4ai remains in solution. Second, the concentration of C4ai in supernatant fractions is determined by double antibody competitive inhibition radioimmunoassay. C4a was recently completely sequenced (Moon et al., 1981) and is readily available as a pure standard. Examination of the specificity of this method confirmed it was indeed specific for C4a antigenicity. Immunochemically depleted C4-deficient plasma and inulin-activated reconstituted C4-deficient plasma exhibited less than 0.1% of the immunoreactivity of untreated plasma. In addition, good agreement was observed in analyses of aggregated IgG activated serum between the experimentally determined concentration of C4ai and that expected from the initial concentration of C4. As a result, recovery and measurement of C4ai in physiological fluids with this method appear both quantitative and specific. Based on results from 17 adult volunteers, the average concentration of C4ai in normal plasma is 488 ng/ml. Interestingly, significant correlation could not be demonstrated between the levels of C4 and C4ai in normal plasma. The mean concentration of C4ai in human urine is 0.5 ng/ml.
本文描述了一种定量检测人血浆、滑液和尿液中补体片段C4ai的方法。首先用50%饱和硫酸铵沉淀样品，以去除交叉反应性大分子C4和前体C4。通过这一步沉淀可去除超过97%的C4，但88%的C4ai仍留在溶液中。其次，通过双抗体竞争性抑制放射免疫测定法测定上清液中C4ai的浓度。C4a最近已完成全序列测定（Moon等人，1981年），并且可容易地获得作为纯标准品。对该方法特异性的检测证实它确实对C4a抗原性具有特异性。免疫化学耗尽C4的缺陷血浆和菊粉激活的重组C4缺陷血浆的免疫反应性不到未处理血浆的0.1%。此外，在分析聚集IgG激活的血清时，实验测定的C4ai浓度与根据C4初始浓度预期的浓度之间观察到良好的一致性。因此，用该方法对生理液中C4ai的回收和测量似乎是定量且特异的。基于17名成年志愿者的结果，正常血浆中C4ai的平均浓度为488 ng/ml。有趣的是，正常血浆中C4和C4ai的水平之间未显示出显著相关性。人尿液中C4ai的平均浓度为0.5 ng/ml。
