Corbeil L B, Hodgson J L, Widders P R
Department of Pathology, University of California, San Diego Medical Center 92103.
J Clin Microbiol. 1991 Dec;29(12):2710-4. doi: 10.1128/jcm.29.12.2710-2714.1991.
A better method for diagnosis of bovine trichomoniasis is needed because culture is slow and somewhat lacking in sensitivity. Immunodiagnosis of Tritrichomonas foetus infection usually involves detection of antigen-antibody reactions with an anti-immunoglobulin conjugate. However, nonspecific immunoglobulin (Ig), bound to the surface of T. foetus, would also be detected by an anti-Ig conjugate and thus would interfere with the specificity of the immunoassay. The goals of this study were to define the binding of bovine immunoglobulins to T. foetus. To determine whether nonimmune binding of Ig to T. foetus occurs, we immunized rabbits with organisms that had been grown in medium containing normal bovine serum and vigorously washed three times with phosphate-buffered saline. The rabbit antiserum had similar titers to T. foetus and to normal bovine serum by enzyme-linked immunosorbent assay (ELISA). Furthermore, two bovine serum proteins were immunoprecipitated by the rabbit antiserum in an immunoelectrophoretogram. One of the serum proteins had a distribution characteristic of IgG2. The rabbit antiserum was then shown to react with purified bovine IgG and IgM by ELISA. Reactivity to IgG was greater. To identify the IgG subisotypes bound and to confirm nonimmune binding of Ig, we grew T. foetus in agammaglobulinemic fetal calf serum and reacted it with IgG1, IgG2, and IgM specific for dinitrophenol (DNP) in ELISA. The binding of IgG2 was greatest, that of IgG1 was next, and that of IgM was least. Little competitive inhibition by DNP was detected, indicating that binding of DNP-specific antibodies was predominantly nonimmune rather than antigen-specific Ig binding. We also demonstrated that T. foetus grew well in medium containing agammaglobulinemic fetal calf serum or serum made agammaglobulinemic by ammonium sulfate precipitation of Igs. This may overcome the problem of low specificity in diagnostic assays as a result of antigen with Ig bound by nonimmune mechanisms.
由于培养方法耗时且灵敏度有所欠缺，因此需要一种更好的牛毛滴虫病诊断方法。胎儿三毛滴虫感染的免疫诊断通常涉及用抗免疫球蛋白缀合物检测抗原 - 抗体反应。然而，结合在胎儿三毛滴虫表面的非特异性免疫球蛋白（Ig）也会被抗Ig缀合物检测到，从而干扰免疫测定的特异性。本研究的目的是确定牛免疫球蛋白与胎儿三毛滴虫的结合情况。为了确定Ig与胎儿三毛滴虫是否发生非免疫结合，我们用在含有正常牛血清的培养基中培养并经磷酸盐缓冲盐水剧烈洗涤三次的生物体免疫兔子。通过酶联免疫吸附测定（ELISA），兔抗血清对胎儿三毛滴虫和正常牛血清的滴度相似。此外，在免疫电泳图谱中，兔抗血清使两种牛血清蛋白发生免疫沉淀。其中一种血清蛋白具有IgG2的分布特征。然后通过ELISA证明兔抗血清与纯化的牛IgG和IgM发生反应。对IgG的反应性更强。为了鉴定结合的IgG亚类并确认Ig的非免疫结合，我们在无丙种球蛋白血症的胎牛血清中培养胎儿三毛滴虫，并在ELISA中使其与对二硝基苯酚（DNP）特异的IgG1、IgG2和IgM反应。IgG2的结合量最大，其次是IgG1，IgM的结合量最小。几乎未检测到DNP的竞争性抑制作用，这表明DNP特异性抗体的结合主要是非免疫性的，而非抗原特异性Ig结合。我们还证明，胎儿三毛滴虫在含有无丙种球蛋白血症的胎牛血清或通过硫酸铵沉淀Ig制成的无丙种球蛋白血症血清的培养基中生长良好。这可能克服了由于抗原与Ig通过非免疫机制结合而导致诊断试验特异性低的问题。
