Identification of surface proteins and antigens from larval stages of Ascaris suum by two-dimensional electrophoresis
Identification of surface proteins and antigens from larval stages of Ascaris suum by two-dimensional electrophoresis
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DOI：
10.1017/s0031182000006892
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发表时间：
2000-12-01
期刊：
影响因子：
2.4
通讯作者：
Yoshihara, S
中科院分区：
文献类型：
--
作者：
Kasuga-Aoki, H;Tsuji, N;Yoshihara, S
了解蛔虫幼虫阶段角质层的生物学非常重要，因为角质层分子不仅具有与生存相关的多种功能，而且还具有作为免疫预防靶标的潜在作用。因此，我们使用两种生物素衍生物和二维聚丙烯酰胺凝胶电泳 (2D-PAGE) 对猪蛔虫三期幼虫 (L3) 和肺期幼虫的表面蛋白和抗原进行了初步表征。 L3 期幼虫和肺期幼虫中生物素标记的蛋白质分别包含 37 个和 32 个点，分子量 (M-r) 范围为 15 至 101 kDa，等电点 (pI) 范围为 3.8 至 7.6。分布图显示各个斑点与一种或两种生物素衍生物结合。此外，仅在L3和肺期幼虫中发现阶段常见和阶段特异性斑点。使用反复接种 A. suum L3 的兔子的抗血清进行 2D-PAGE/免疫印迹分析。将免疫印迹与表面蛋白谱进行比对后，来自 L3 和肺期幼虫的 19 个点被识别为表面抗原，M-r 范围为 32 至 66 kDa，pI 范围为 4.9 至 7.6。发现这些斑点包括阶段常见和阶段特异性抗原。通过生物素标记结合2D-PAGE来鉴定表面蛋白可以大大缩短目标蛋白的样品制备时间，并且将成为猪A. suum L3和肺期幼虫表面蛋白和抗原的蛋白质分析的可行方法。
An understanding of the biology of the cuticle in the larval stages of Ascariodea is of importance since the cuticle molecules not only possess a variety of functions related to survival but also have a potential role as a target for immunoprophylaxis. Thus, we made a preliminary characterization of surface proteins and antigens from 3rd-stage larvae (L3) and lung-stage larvae of Ascaris suum using two biotin-derivatives and two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). The proteins labelled with biotin comprised a total of 37 and 32 spots, with molecular weights (M-r) ranging from 15 to 101 kDa and isoelectric points (pI) from 3.8 to 7.6, in L3 and lung-stage larvae, respectively. The profiles revealed that the individual spots bound to one or both biotin derivatives. In addition, stage-common and stage-specific spots mere found in L3 and lung-stage larvae. 2D-PAGE/immunoblotting analysis was performed with antisera from rabbits repeatedly inoculated with A. suum L3. Nineteen spots were recognized as surface antigens, with M-r ranging from 32 to 66 kDa and pI from 4.9 to 7.6, from L3 and lung-stage larvae after alignment of the immunoblots with the profile of the surface proteins. These spots were found to include stage-common and stage-specific antigens. Identification of surface proteins by biotin labelling combined with 2D-PAGE allows a substantial shortening of sample preparation time for the target proteins, and will be a viable method for protein analysis of surface proteins and antigens of A. suum L3 and lung-stage larvae.