Zhongguo xue xi chong bing fang zhi za zhi = Chinese journal of schistosomiasis control · 2011
[Cloning and expression of echinococcus granulosus Eg18 and preliminary evaluation of immunological assay with recombinant antigen].
Аннотация на английскомСтатьяПоддержка исследования: вне правительства СШАНазвание и аннотация: Перевод пока не подготовлен
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Wang Yong-Shun, Han Xiu-Min, Wang Hu
Аннотация
OBJECTIVE: To clone and express Echinococcus granulosus Eg18 gene and evaluate the immunoreactivity of the recombinant protein.
METHODS: Eg18 coding sequence was amplified by RT-PCR using primers designed according to the sequence of Eg18 in GenBank from total RNA extracted from Echinococcus granulosus protoscoleces isolated from infected sheep in Qinhai Province and cloned into the prokaryotic expression vector pET-28a(+). The recombinant expression vector was transformed to Escherichia coli BL21 (DE) and induced to express by IPTG. The expressed products were analyzed by SDS-PAGE and purified with Ni-IDA agarose affinity chromatography. rEg18 was evaluated for its reactivity with the sera from the patients infected with hydatid and other helminthes by Western blotting and ELISA.
RESULTS: The sequence of cloned Eg18 was completely identical wit the original sequences of Eg18 and Em18 deposited in GenBank. The recombinant protein strongly reacted to the sera from the patients with alveolar echinococcosis, cystic echinococcosis, and cysticercosis, and weakly reacted to the sera from the patients with nematodiasis, schistosomiasis and clonorchiasis. The detection of specific IgG4 was much more specific than that of IgG.
CONCLUSION: Eg18/Em18 is the common antigen of cestode and its specific IgG4 is a serum marker of alveolar echinococcosis.
METHODS: Eg18 coding sequence was amplified by RT-PCR using primers designed according to the sequence of Eg18 in GenBank from total RNA extracted from Echinococcus granulosus protoscoleces isolated from infected sheep in Qinhai Province and cloned into the prokaryotic expression vector pET-28a(+). The recombinant expression vector was transformed to Escherichia coli BL21 (DE) and induced to express by IPTG. The expressed products were analyzed by SDS-PAGE and purified with Ni-IDA agarose affinity chromatography. rEg18 was evaluated for its reactivity with the sera from the patients infected with hydatid and other helminthes by Western blotting and ELISA.
RESULTS: The sequence of cloned Eg18 was completely identical wit the original sequences of Eg18 and Em18 deposited in GenBank. The recombinant protein strongly reacted to the sera from the patients with alveolar echinococcosis, cystic echinococcosis, and cysticercosis, and weakly reacted to the sera from the patients with nematodiasis, schistosomiasis and clonorchiasis. The detection of specific IgG4 was much more specific than that of IgG.
CONCLUSION: Eg18/Em18 is the common antigen of cestode and its specific IgG4 is a serum marker of alveolar echinococcosis.