School of Biotechnology, East China University of Science and Technology, Shanghai 200237, P,R, China.
Parasit Vectors. 2012 Jun 11;5:115. doi: 10.1186/1756-3305-5-115.
Schistosomiasis is a chronic disease caused by trematode flatworms of the genus Schistosoma. The disease remains a serious public health problem in endemic countries and affects at least 207 million people worldwide. A definite diagnosis of the disease plays a key role in the control of schistosomiasis. The detection of schistosome circulating antigens (CAs) is an effective approach to discriminate between previous exposure and current infection. Different methods have been investigated for detecting the CAs. However, the components of the schistosome CAs remain unclear. In this study, we analyzed the CAs in sera of patients infected with Schistosoma japonicum.
The parasites were collected from the infected rabbits for preparing the adult worm antigen (AWA). The hyline hens were immunized subcutaneously with AWA to produce anti-AWA IgY. The IgY was purified by water-dilution and ammonium sulfate precipitation method and identified by ELISA and Western blotting. After purification and characterization, IgY was immobilized onto the resin as a capture antibody. The circulating antigens were immune-precipitated from patients' serum samples by direct immunoprecipitation. The precipitated proteins were separated by one-dimensional electrophoresis and analyzed by LC-MS/MS.
Firstly, the IgY against AWA was produced from the eggs of immunized hens by AWA, which gave a titer of 1:12800. The purified IgY was used as the capture antibody to enrich the CAs in sera of S. japonicum infected patients through immunoprecipitation. The CAs were determined by LC-MS/MS. There were four proteins, including protein BUD31 homolog, ribonuclease, SJCHGC06971 protein and SJCHGC04754 protein, which were identified among the CAs.
We developed a novel method based on IgY for identification and profiling CAs in sera of S. japonicum infected patients. Four new CAs were identified and have potential value for further development of an antigen assay.
血吸虫病是一种由血吸虫属扁形动物引起的慢性疾病。该疾病在流行国家仍然是一个严重的公共卫生问题，影响全球至少 2.07 亿人。对该疾病的明确诊断对血吸虫病的控制起着关键作用。检测血吸虫循环抗原 (CA) 是区分既往暴露和当前感染的有效方法。已经研究了不同的方法来检测 CA。然而，血吸虫 CA 的成分仍不清楚。在本研究中，我们分析了感染日本血吸虫的患者血清中的 CA。
从感染的兔子中收集寄生虫，用于制备成虫抗原 (AWA)。用 AWA 皮下免疫产蛋母鸡，产生抗-AWA IgY。用稀释法和硫酸铵沉淀法纯化 IgY，并通过 ELISA 和 Western blot 鉴定。纯化和表征后，IgY 被固定在树脂上作为捕获抗体。通过直接免疫沉淀从患者血清样本中免疫沉淀循环抗原。沉淀的蛋白质通过一维电泳分离，并通过 LC-MS/MS 进行分析。
首先，用 AWA 从免疫母鸡的卵中产生了针对 AWA 的 IgY，其效价为 1:12800。纯化的 IgY 用作捕获抗体，通过免疫沉淀从感染日本血吸虫的患者血清中富集 CA。通过 LC-MS/MS 确定 CA。在 CA 中鉴定出四种蛋白质，包括 BUD31 同源蛋白、核糖核酸酶、SJCHGC06971 蛋白和 SJCHGC04754 蛋白。
我们开发了一种基于 IgY 的新方法，用于鉴定和分析感染日本血吸虫的患者血清中的 CA。鉴定出四种新的 CA，它们具有进一步开发抗原检测的潜在价值。
