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目的鉴定杜氏利什曼原虫无鞭毛体特异表达抗原。方法培养杜氏利什曼原虫前鞭毛体并体外转化无鞭毛体,其总蛋白经2-DE电泳后以小鼠抗杜氏利什曼原虫无鞭毛体血清进行Western blot,对前鞭毛体与无鞭毛体特异表达抗原蛋白进行MALDI-TOF/TOF串联质谱鉴定。重组表达无鞭毛体特异表达抗原编码基因,以Western blot法对重组蛋白进行鉴定。结果等量的杜氏利什曼原虫前鞭毛体与无鞭毛体蛋白经2-DE电泳均可呈现680~742个蛋白点,Western blot及MALDI-TOF/TOF-MS分析甘油醛3-磷酸脱氢酶与延伸因子2为杜氏利什曼原虫前鞭毛体特异表达抗原,核苷二磷酸激酶为无鞭毛体特异表达抗原。重组核苷二磷酸激酶编码基因表达产物经Western blot证实为杜氏利什曼原虫无鞭毛体特异表达强抗原。结论杜氏利什曼原虫前鞭毛体与无鞭毛体抗原表达存在差异,核苷二磷酸激酶为杜氏利什曼原虫无鞭毛体特异表达强抗原。
Objective To identify Leishmania donovani specific expressed antigen. Methods The Leishmania donovani promastigotes were cultured and transformed into amastigotes in vitro. The total protein was detected by 2-DE electrophoresis and Western blot was performed on mouse anti-Leishmania donovani serum. The body-specific antigenic protein was identified by MALDI-TOF / TOF tandem mass spectrometry. Recombinant expression of amastigotes specific expression of antigen-encoding genes, Western blot method for the identification of recombinant proteins. Results Equivalent amount of Leishmania donovani promastigotes and amastigotes protein showed 680 ~ 742 protein spots by 2-DE electrophoresis, Western blot and MALDI-TOF / TOF-MS analysis of glyceraldehyde 3-phosphate dehydrogenation Enzymes and elongation factor 2 are Leishmania donovani promastigotes specific expression of antigens, nucleoside diphosphate kinase is amastigote-specific expression of antigens. Recombinant nucleoside diphosphate kinase gene expression products confirmed by Leishmania donovani amastigote-specific expression of strong antigen. Conclusion The expression of Leishmania donovani promastigotes and amastigote antigen are different. Nucleoside diphosphate kinase is a strong antigen expressed specifically by Leishmania donovani.