目的筛选合适的内参基因以检测细粒棘球绦虫水通道蛋白(aquaporins of Echinococcus granulosus,EgAQPs)基因在原头蚴发育为棘球蚴过程中的转录表达水平。方法通过geNorm软件分析5个候选内参基因(ND1、ATP6、elp、actin和ActⅡ)在原头...目的筛选合适的内参基因以检测细粒棘球绦虫水通道蛋白(aquaporins of Echinococcus granulosus,EgAQPs)基因在原头蚴发育为棘球蚴过程中的转录表达水平。方法通过geNorm软件分析5个候选内参基因(ND1、ATP6、elp、actin和ActⅡ)在原头蚴成囊过程中的稳定性,并运用RT-q PCR方法检测体外培养原头蚴及原头蚴在小鼠体内囊化形成的继发棘球蚴中EgAQPs基因转录表达量。结果actin和ActⅡ、ATP6和ND1分别为检测体外培养原头蚴和原头蚴囊化形成的继发棘球蚴囊壁生发层细胞EgAQPs基因转录表达状态的理想内参基因。EgAQP3和EgAQP9在体外培养第3天的原头蚴中转录表达量最高,随着培养时间的增加,转录表达量呈持续下降的趋势,且不同培养时间转录表达量的差异有统计学意义(P<0.05);EgAQP4在体外培养第25天的原头蚴中转录表达量最高(P<0.05),在体外培养第3、15、35天的原头蚴中转录表达量较低,这3个时间点的差异无统计学意义(P>0.05)。EgAQP3、EgAQP4和EgAQP9在原头蚴中转录表达量最高(P<0.05),随着感染时间的延长,EgAQP3、EgAQP4和EgAQP9的转录表达水平在接种原头蚴感染小鼠后第3~7个月的继发棘球蚴囊壁生发层细胞中转录表达水平很低,差异无统计学意义(P>0.05)。EgAQP、EgAQP1、EgAQPFA-CHIP和EgAQPAnG在原头蚴和继发棘球蚴囊壁生发层细胞中的转录表达水平均很低,基本上检测不到。结论通过geNorm软件筛选了原头蚴发育为棘球蚴过程中稳定的内参基因;原头蚴囊化的继发棘球蚴囊壁生发层细胞中EgAQPs基因的转录表达水平很低,可能与棘球蚴液形成或积累有关。展开更多
Objective:To explore the serodiagnosis of hydatid cyst in human using different antigens of sheep(hydatid fluid,somatic and Lxcretory/secretory antigens of protoscolexi by ELISA and compares this result with commercia...Objective:To explore the serodiagnosis of hydatid cyst in human using different antigens of sheep(hydatid fluid,somatic and Lxcretory/secretory antigens of protoscolexi by ELISA and compares this result with commercial human ELISA kit.Methods:one hundred blood samples from patients with history of severe abdominal pain and cosinophilia were obtained.Ten serum samples were obtained from surgically and pathologically confirmed cystic echinococcosis patients from Mashhad university hospital as positive control and 5 serum samples from infant under one year old as negative control.Blood samples were centrifuged at 3000xg at 20℃for 15 min and sera were stored at-20℃.First,these samples were tested for the presence of antibody by commercial human ELISA.Then.ELISA was developed on microplates coated with hydatid fluid,Somatic and Excretory/secretory antigens of protoscolex of sheep.Results:The results of this study as analyzed by Kappa test showed that,hydatid fluid antigen could be used as a precise source of detection in indirect ELISA test.Conclusions:Hydatid fluid in comparison with Excretory-secretory and somatic antigens showed more compatibility agreement in kappa test which can be used for further studies in development of any ELISA test for diagnosis of human hydatidosis.展开更多
文摘目的筛选合适的内参基因以检测细粒棘球绦虫水通道蛋白(aquaporins of Echinococcus granulosus,EgAQPs)基因在原头蚴发育为棘球蚴过程中的转录表达水平。方法通过geNorm软件分析5个候选内参基因(ND1、ATP6、elp、actin和ActⅡ)在原头蚴成囊过程中的稳定性,并运用RT-q PCR方法检测体外培养原头蚴及原头蚴在小鼠体内囊化形成的继发棘球蚴中EgAQPs基因转录表达量。结果actin和ActⅡ、ATP6和ND1分别为检测体外培养原头蚴和原头蚴囊化形成的继发棘球蚴囊壁生发层细胞EgAQPs基因转录表达状态的理想内参基因。EgAQP3和EgAQP9在体外培养第3天的原头蚴中转录表达量最高,随着培养时间的增加,转录表达量呈持续下降的趋势,且不同培养时间转录表达量的差异有统计学意义(P<0.05);EgAQP4在体外培养第25天的原头蚴中转录表达量最高(P<0.05),在体外培养第3、15、35天的原头蚴中转录表达量较低,这3个时间点的差异无统计学意义(P>0.05)。EgAQP3、EgAQP4和EgAQP9在原头蚴中转录表达量最高(P<0.05),随着感染时间的延长,EgAQP3、EgAQP4和EgAQP9的转录表达水平在接种原头蚴感染小鼠后第3~7个月的继发棘球蚴囊壁生发层细胞中转录表达水平很低,差异无统计学意义(P>0.05)。EgAQP、EgAQP1、EgAQPFA-CHIP和EgAQPAnG在原头蚴和继发棘球蚴囊壁生发层细胞中的转录表达水平均很低,基本上检测不到。结论通过geNorm软件筛选了原头蚴发育为棘球蚴过程中稳定的内参基因;原头蚴囊化的继发棘球蚴囊壁生发层细胞中EgAQPs基因的转录表达水平很低,可能与棘球蚴液形成或积累有关。
基金funded by a grant from Ferdowsi University of Mashhad
文摘Objective:To explore the serodiagnosis of hydatid cyst in human using different antigens of sheep(hydatid fluid,somatic and Lxcretory/secretory antigens of protoscolexi by ELISA and compares this result with commercial human ELISA kit.Methods:one hundred blood samples from patients with history of severe abdominal pain and cosinophilia were obtained.Ten serum samples were obtained from surgically and pathologically confirmed cystic echinococcosis patients from Mashhad university hospital as positive control and 5 serum samples from infant under one year old as negative control.Blood samples were centrifuged at 3000xg at 20℃for 15 min and sera were stored at-20℃.First,these samples were tested for the presence of antibody by commercial human ELISA.Then.ELISA was developed on microplates coated with hydatid fluid,Somatic and Excretory/secretory antigens of protoscolex of sheep.Results:The results of this study as analyzed by Kappa test showed that,hydatid fluid antigen could be used as a precise source of detection in indirect ELISA test.Conclusions:Hydatid fluid in comparison with Excretory-secretory and somatic antigens showed more compatibility agreement in kappa test which can be used for further studies in development of any ELISA test for diagnosis of human hydatidosis.
