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Korean J Parasitol > Volume 35(4):1997 > Article

Original Article
Korean J Parasitol. 1997 Dec;35(4):251-258. English.
Published online Dec 20, 1997.  http://dx.doi.org/10.3347/kjp.1997.35.4.251
Copyright © 1997 by The Korean Society for Parasitology
Partially purified Toxoplasma gondii antigens by immunoaffinity chromatography
M H Ahn,*1K H Hyun,1J O Kang,2 and D Y Min1
1Department of Parasitology, College of Medicine, Hanyang University, Seoul 133-791, Korea.
Received July 26, 1997; Accepted September 11, 1997.

Abstract

Tachyzoite antigens of Toxoplasma gondii (RH) were partially purified by immunoaffinity chromatography. The cultivated Toxoplasma in vivo (mouse) and in vitro (Hep-2 cell) and peritoneal fluid of T. gondii infected mice were collected for antigen analysis. Tachyzoite antigens collected from infected mouse showed positive bands of 76 kDa, 70 kDa, 64 kDa, 53 kDa, 46 kDa, 44 kDa, 41 kDa, 35 kDa, 25 kDa, 18 kDa, and 13 kDa on immunoblot with anti-Toxoplasma rabbit sera, and those from infected Hep-2 cells revealed reactive bands of 70 kDa, 64 kDa, 53 kDa, 35 kDa 28 kDa, and 13-10 kDa. After applying to an IgG-Sepharose column, two elution peaks, E-1 and E-2 fractions, were obtained from both soluble antigen of T. gondii and the peritoneal fluid of infected mice, respectively. Immunoblots of soluble antigen with immunized rabbit sera revealed positive bands of 97 kDa, 63 kDa, 53 kDa, and 35 kDa from E-1 fraction and 53 kDa and 35 kDa from E-2. In the case of the eluted peaks from mice peritoneal fluid, E-1 showed protein bands of 84 kDa, 76 kDa, 53 kDa, and 29 kDa bands and 53 kDa and 45 kDa from E-2 on immunoblots. Serum IgG antibody titer of mice immunized with T. gondii tachyzoites was increased on 1 week after booster immunization when analysed by ELISA using crude antigen, while it was elevated on 3 weeks after booster immunization by ELISA using purified antigen.

Figures


Fig. 1
SDS-PAGE (A) and immunoblot (B) analyses of T. gondii tachyzoite. M, marker; lane 1, T. gondii tachyzoite from infected mouse peritoneum; lane 2, T. gondii tachyzoite cultured in vitro with Hep-2 cell.


Fig. 2
SDS-PAGE (A) and immunoblot (B) analyses of crude or purified T. gondii antigen. M, marker; lane 1, T. gondii tachyzoite lysate; lane 2, the first elution peak of purified Toxoplasma antigen; lane 3, the second elution peak of purified Toxoplasma antigen.


Fig. 3
SDS-PAGE (A) and immunoblot (B) analyses of Toxoplasma infected mouse peritoneal fluid and purified antigen. M, marker; lane 1, ascitic fluid from infected mouse; lane 2, the first elution peak of purified antigen; lane 3, the second elution peak of purified antigen.

Tables


Table 1
Serum IgG antibody levels of T. gondii immunized mice by ELISA using crude antigen


Table 2
Serum IgG antibody levels of T. gondii immunized mice by ELISA using purified antigen

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