摘要
目的探讨日本血吸虫信号蛋白14-3-3(Sj14-3-3)在虫卵内的定位和重组Sj14-3-3(rSj14-3-3)的免疫诊断价值。方法从日本血吸虫尾蚴感染42 d的兔肝脏中分离虫卵,用逆转录聚合酶链反应(RT-PCR)检测Sj14-3-3基因在虫卵期的转录水平;兔肝组织石蜡切片免疫组化染色,观察内源性Sj14-3-3在虫卵内的分布和表达丰度。利用纯化的rSj14-3-3和可溶性虫卵抗原(SEA),采用间接酶联免疫吸附试验(ELISA)检测急、慢性血吸虫病患者和正常人血清。结果RT-PCR从日本血吸虫成熟虫卵中检出760 bp左右的基因片断;免疫组化显示Sj14-3-3主要分布在虫卵内毛蚴的体壁和两边的侧腺。检测急性、慢性患者和正常人血清抗rSj14-3-3抗原的抗体阳性率分别为91.0%、78.9%、0.0%;上述标本中抗SEA抗体的阳性率分别为97.4%、88.9%和2.5%。结论用ELISA检测抗SEA抗体和rSj14-3-3抗体诊断血吸虫病具有高度的特异性和敏感性,而Sj14-3-3在成熟虫卵阶段高表达,可能是SEA的主要组分,具有实用价值。
Objective To investigate the localization of 14-3-3 protein （Sj14-3-3） in egg stage of Schistosomajaponicum and its potential use in diagnosis of schistosomiasis. Methods The eggs were purified from liver tissues of the rabbits infected with Schistosomajaponicum on day 42 after infection. The transcription of Sj14-3-3 gene in eggs was confirmed by reverse transcription-polymerase chain reaction（RT-PCR） using total RNA. Immuno-localization of Sj14-3-3 within egg was performed with monoclonal antibody to Sj14-3-3. Sera from patients with schistosomiasis were detected with purified recombinant Sj14-3-3 （rSj14-3-3） and soluble egg antigen （SEA） by indirect enzyme-linked immunosorbent assay （ELISA）. Results Consistent with the expected size of 760 bp, the amplified product of RT-PCR was obtained. The Sj14-3-3 was predominantly localized in integument and lateral gland of miracidia in egg. The results of ELISA showed that the positive rates of anti-Sj14-3-3 antibodies in sera from acute and chronic cases of schistosomiasis were 91.0% and 78.9% ; While the positive rates of anti-SEA antibodies were 97.4% and 88.9%, respectively. No false positive reaction was noted in normal sera detected by rSj14-3-3 but slight nonspecific reaction by SEA. Conclusions The study shows that the SEA and rSj14-3-3 are useful in immunodiagnosis of schistosomiasis. Sj14-3-3 is rich in the egg stage and may be a predominant fraction in SEA.
出处
《检验医学》
CAS
北大核心
2007年第2期128-131,共4页
Laboratory Medicine
基金
国家高技术研究发展计划(863计划)资助项目(2004AA2Z3570)
国家自然科学基金资助项目(30170841)
关键词
日本血吸虫
虫卵
14-3-3
定位
免疫诊断
Schistosomajaponicum
Egg
14-3-3 protein
Immunolocalization
Immunodiagnosis
