Verbisck N V, Da-Silva S, Mortara R A
Departamento de Microbiologia, Imunologia e Parasitologia, Escola Paulista de Medicina, Universidade Federal de São Paulo, Brasil.
Braz J Med Biol Res. 1998 Dec;31(12):1583-91. doi: 10.1590/s0100-879x1998001200011.
We have raised monoclonal antibodies (mAbs) directed towards amastigote forms of Trypanosoma cruzi, and shown that mAbs ID9 and 4B9 are carbohydrate while mAb 4B5 activity is resistant to periodate oxidation of the antigen. Here we used an ELISA to quantitate and compare the expression of surface epitopes on fixed parasites among different parasite isolates. The expression of markers varied among T. cruzi amastigotes isolated from infected cells or after extracellular differentiation of trypomastigotes. Moreover, we also observed an extensive polymorphic expression of these epitopes among amastigotes derived from different strains and clones. For instance, mAb 2C2 strongly and evenly reacted with 9 strains and clones (G, Y, CL, Tulahuen, MD, and F, and clones Sylvio X-10/4, D11, and CL.B), with absorbance at 492 nm (A492 nm) from 0.6 to 0.8. By contrast, mAb 4B5 had a higher expression in Tulahuen amastigotes (around 0.9 at 492 nm) whereas its reactivity with amastigotes from clones CL.B, Sylvio X-10/4 and D11 was much lower (around 0.4). mAb 1D9 displayed an interesting pattern of reactivity with amastigotes of the different strains and clones (A492 nm of G > D11 > or = Sylvio X-10/4 = MD > Tulahuen = F = Y > CL > CL.B). Finally, we observed that mAb 4B9 had the lowest reaction with the parasites studied, with higher values of A492 nm with Y strain (around 0.6) and lower values with Tulahuen, F and CL.B strains (around 0.2). Immunoblotting analysis also showed extensive variations among amastigotes of the various parasite isolates and mAbs 4B9, 1D9 and 4B5 revealed significant differences in expression between clones and parental strains. These data describe a previously uncharacterized polymorphism of T. cruzi amastigote surface components.
我们制备了针对克氏锥虫无鞭毛体形式的单克隆抗体（mAb），并表明单克隆抗体ID9和4B9识别碳水化合物，而单克隆抗体4B5的活性对抗原的高碘酸盐氧化具有抗性。在此，我们使用酶联免疫吸附测定（ELISA）来定量和比较不同寄生虫分离株的固定寄生虫表面表位的表达。在从感染细胞分离的克氏锥虫无鞭毛体或锥鞭毛体胞外分化后，标志物的表达有所不同。此外，我们还观察到这些表位在源自不同菌株和克隆的无鞭毛体中存在广泛的多态性表达。例如，单克隆抗体2C2与9个菌株和克隆（G、Y、CL、图拉洪、MD和F，以及克隆西尔维奥X-10/4、D11和CL.B）强烈且均匀地反应，492nm处的吸光度（A492nm）为0.6至0.8。相比之下，单克隆抗体4B5在图拉洪无鞭毛体中的表达较高（492nm处约为0.9），而其与克隆CL.B、西尔维奥X-10/4和D11的无鞭毛体的反应性则低得多（约为0.4）。单克隆抗体1D9与不同菌株和克隆的无鞭毛体呈现出有趣的反应模式（G的A492nm > D11 >或=西尔维奥X-10/4 = MD >图拉洪 = F = Y > CL > CL.B）。最后，我们观察到单克隆抗体4B9与所研究的寄生虫反应最低，Y菌株的A492nm值较高（约为0.6），而图拉洪、F和CL.B菌株的值较低（约为0.2）。免疫印迹分析还显示，各种寄生虫分离株的无鞭毛体之间存在广泛差异，单克隆抗体4B9、1D9和4B5揭示了克隆和亲本菌株之间在表达上的显著差异。这些数据描述了克氏锥虫无鞭毛体表面成分以前未被表征的多态性。
