Navarrete-Perea José, Orozco-Ramírez Rodrigo, Moguel Bárbara, Sciutto Edda, Bobes Raúl J, Laclette Juan P
Department of Immunology, Institute for Biomedical Research, Universidad Nacional Autónoma de México, Mexico D.F. 04510, Mexico.
Department of Immunology, Institute for Biomedical Research, Universidad Nacional Autónoma de México, Mexico D.F. 04510, Mexico.
Mol Biochem Parasitol. 2015 Jul;202(1):22-8. doi: 10.1016/j.molbiopara.2015.08.005. Epub 2015 Sep 1.
Human and porcine cysticercosis is caused by the larval stage of the flatworm Taenia solium (Cestoda). The protein extracts of T. solium cysts are complex mixtures including cyst's and host proteins. Little is known about the influence of using different detergents in the efficiency of solubilization-extraction of these proteins, including relevant antigens. Here, we describe the use of CHAPS, ASB-14 and Triton X-100, alone or in combination in the extraction buffers, as a strategy to notably increase the recovery of proteins that are usually left aside in insoluble fractions of cysts. Using buffer with CHAPS alone, 315 protein spots were detected through 2D-PAGE. A total of 255 and 258 spots were detected using buffers with Triton X-100 or ASB-14, respectively. More protein spots were detected when detergents were combined, i.e., 2% CHAPS, 1% Triton X-100 and 1% ASB-14 allowed detection of up to 368 spots. Our results indicated that insoluble fractions of T. solium cysts were rich in antigens, including several glycoproteins that were sensitive to metaperiodate treatment. Host proteins, a common component in protein extracts of cysts, were present in larger amounts in soluble than insoluble fractions of cysts proteins. Finally, antigens present in the insoluble fraction were more appropriate as a source of antigens for diagnostic procedures.
人和猪的囊尾蚴病是由扁形虫猪带绦虫(绦虫纲)的幼虫阶段引起的。猪带绦虫囊肿的蛋白质提取物是包括囊肿蛋白和宿主蛋白的复杂混合物。关于使用不同去污剂对这些蛋白质(包括相关抗原)的溶解提取效率的影响知之甚少。在此,我们描述了在提取缓冲液中单独或组合使用CHAPS、ASB - 14和Triton X - 100,作为显著提高通常留在囊肿不溶性部分中的蛋白质回收率的策略。单独使用含CHAPS的缓冲液时,通过二维聚丙烯酰胺凝胶电泳检测到315个蛋白点。分别使用含Triton X - 100或ASB - 14的缓冲液时,共检测到255个和258个蛋白点。当去污剂组合使用时检测到更多蛋白点,即2% CHAPS、1% Triton X - 100和1% ASB - 14可检测到多达368个蛋白点。我们的结果表明,猪带绦虫囊肿的不溶性部分富含抗原,包括几种对高碘酸盐处理敏感的糖蛋白。宿主蛋白是囊肿蛋白质提取物中的常见成分,在囊肿蛋白质的可溶性部分中比不溶性部分含量更高。最后,不溶性部分中存在的抗原更适合作为诊断程序的抗原来源。