Cai Hui-Xia, Shen Yu-Juan, Han Xiu-Min, Yuan Zhong-Ying, Wang Hu, Xu Yu-Xin, Hu Yuan, Lu Wei-Yuan, Guan Ya-Yi, Cao Jian-Ping
National Institute of Parasitice Diseases, Chinese Center for Disease Control and Prevention, Shanghai 200025, China.
Zhongguo Ji Sheng Chong Xue Yu Ji Sheng Chong Bing Za Zhi. 2011 Jun;29(3):167-71.
To clone and express EPCl gene of Echinococcus granulosus, and investigate its immunogenicity and diagnostic value.
Total RNA was extracted from hydatid cyst protoscoleces and EPC1 gene of Echinococcus granulosus was amplified by RT-PCR. The PCR product was cloned into pGEM-T vector, and then subcloned into the prokaryotic expression vector PET28a(+). The positive recombinants were transformed into Escherichia coli BL21 (DE3), and followed by expression of the protein induced by IPTG. The recombinant protein was identified by SDS-PAGE and Western blotting, and used to establish ELISA. Serum samples from patients with cystic echinococcosis (60 cases), alve-olar echinococcosis (37 cases), cysticercosis (16 cases), clonorchiasis sinensis (7 cases), schistosomiasis japonica (4 cases) and healthy persons (33 cases) were examined.
The recombinant plasmid PET28a-EgEPC1 was identified by restriction enzyme digestion and sequencing. SDS-PAGE result showed that the recombinant containing recombinant plasmid PET28a-EgEPC1 expressed a soluble fission protein of EgEPC1 (about M, 11 000). The protein was recognized by pool sera of cystic echinococcosis patients. The overall sensitivity and specificity of diagnosis by ELISA for cystic echinococcosis were 78.3% (47/60), and 98.3% (59/60), respectively. The cross reaction with sera of alveolar echinococcosis was 40.5% (15/37).
The recombinant EgEPC1 antigen has diagnostic value in cystic echinococcosis.
克隆并表达细粒棘球绦虫EPC1基因，探讨其免疫原性及诊断价值。
从细粒棘球绦虫包虫囊肿原头节中提取总RNA，采用RT-PCR扩增细粒棘球绦虫EPC1基因。将PCR产物克隆至pGEM-T载体，再亚克隆至原核表达载体PET28a(+)。将阳性重组体转化至大肠杆菌BL21(DE3)，经IPTG诱导表达蛋白。通过SDS-PAGE和Western印迹鉴定重组蛋白，并用于建立ELISA。检测囊型棘球蚴病患者（60例）、泡型棘球蚴病患者（37例）、囊尾蚴病患者（16例）、华支睾吸虫病患者（7例）、日本血吸虫病患者（4例）及健康人（33例）的血清样本。
经酶切鉴定和测序确认重组质粒PET28a-EgEPC1。SDS-PAGE结果显示，含重组质粒PET28a-EgEPC1的重组菌表达出可溶性的EgEPC1裂解蛋白（约Mr 11 000）。该蛋白能被囊型棘球蚴病患者混合血清识别。ELISA诊断囊型棘球蚴病的总体敏感性和特异性分别为78.3%（47/60）和98.3%（59/60）。与泡型棘球蚴病患者血清的交叉反应率为40.5%（15/37）。
重组EgEPC1抗原对囊型棘球蚴病具有诊断价值。
