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[利用色谱法、聚丙烯酰胺凝胶电泳和免疫沉淀法对刚地弓形虫进行抗原分析]

[Antigen analysis of Toxoplasma gondii using chromatography, polyacrylamide gel electrophoresis and immunoprecipitation].

作者信息

Hafid J, Sung R T, Raberin H

出版信息

Ann Soc Belg Med Trop. 1989 Mar;69(1):41-8.

PMID:2499276
Abstract

Whole (EAT), cytoplasmic (EAC) and membranous (EAM) extracts were prepared from Toxoplasma gondii RH Strain trophozoites by means of osmotic lysis and/or ultra sonication. Sephacryl S 300 gel chromatography shows respectively 5, 4 and 2 protein fractions from EAT, EAC and EAM. Polyacrylamide gel electrophoresis (PAGE) allowed to detect 22 proteins (WM from 6 to 130 Kd) from cytoplasmic extract and 3 proteins (MW: 8, 30 and 65 Kd) from membranous extract. Immunoprecipitation techniques were performed with polyclonal antibodies against Toxoplasma prepared from serum and rabbits immunized with EAT. Counter electro-immunodiffusion (CEID) on cellulose acetate membrane was found to be more sensitive than double immunodiffusion and immunoelectrophoresis as it detected 8 lines with both cytoplasmic and membranous extracts. The latter seem to be composed of large numbers of proteins which are present in very low quantities difficult to detect in PAGE, but very antigenic and easily shown by CEID.

摘要

通过渗透裂解和/或超声处理,从刚地弓形虫RH株滋养体中制备了全提取物(EAT)、细胞质提取物(EAC)和膜提取物(EAM)。Sephacryl S 300凝胶色谱分别显示EAT、EAC和EAM中有5、4和2个蛋白质组分。聚丙烯酰胺凝胶电泳(PAGE)可从细胞质提取物中检测到22种蛋白质(分子量从6至130千道尔顿),从膜提取物中检测到3种蛋白质(分子量分别为8、30和65千道尔顿)。使用从用EAT免疫的兔血清制备的抗弓形虫多克隆抗体进行免疫沉淀技术。发现醋酸纤维素膜上的对流免疫电泳(CEID)比双向免疫扩散和免疫电泳更灵敏,因为它在细胞质和膜提取物中均检测到8条线。后者似乎由大量蛋白质组成,这些蛋白质在PAGE中含量极低难以检测,但具有很强的抗原性,在CEID中很容易显示出来。

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