禽坦布苏病毒NS1-ELISA的初步建立

万春和; 潘异哲; 傅秋玲; 陈珍; 陈翠腾; 傅光华; 陈红梅; 程龙飞; 施少华; 黄瑜

doi:10.19303/j.issn.1008-0384.2015.11.001

禽坦布苏病毒NS1-ELISA的初步建立

doi: 10.19303/j.issn.1008-0384.2015.11.001

1. 福建省农业科学院畜牧兽医研究所/福建省畜禽疫病防治工程技术研究中心,福建福州,350013;
2. 浙江省湖州市动物疫病与预防控制中心,浙江湖州,313000

基金项目:

福建省农业科学院畜牧兽医研究所基金项目(MYQJ2014-3)

国家现代农业产业体系建设专项(CARS-43)

福建省农业科学院青年人才创新基金项目(2014QA-7)

福建省农业科学院“青年科技英才百人计划”项目(YC2015-12)

详细信息

作者简介:
万春和(1982-),男,博士,助理研究员,主要从事水禽病原分子生物学研究;黄瑜(1965-),男,博士,研究员,主要从事畜禽传染病学研究(E-mail:huangyu_815@163.com)

中图分类号: S852.65
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出版历程
- 收稿日期: 2015-08-31
- 刊出日期: 2015-11-18

NS1-ELISA for Antibody Detection on Avian Tembusu Virus

1. Institute of Animal Husbandry and Veterinary Medicine, Fujian Academy of Agricultural Sciences/Fujian Animal Disease Control Technology Development Center, Fuzhou, Fujian 350013, China;
2. Zhejiang Huzhou Center for Animal Disease Control and Prevention, Huzhou, Zhejiang 313000, China

摘要

摘要: 对禽坦布苏病毒非结构蛋白NS1基因进行原核表达,并进行生物学活性鉴定。将表达成功具有生物学活性的重组蛋白经纯化后,作为抗原包被ELISA平板,通过条件优化,建立检测禽坦布苏病毒血清学检测方法。结果表明,表达的NS1重组蛋白大小约为60kd,该蛋白经Western blot检测具有良好的生物学活性。将该重组蛋白纯化后包被ELISA板建立检测禽坦布苏病毒血清学间接ELISA方法,该方法优化后的参数为2.5μg·mL^-1包被ELISA板;一抗(鸭血清)血清稀释度为1∶100;酶标二抗为1∶3 000。本研究建立的方法特异性强,批内和批间重复性较好。用建立的间接ELISA方法对坦布苏病毒灭活苗免疫鸭血清和自然感染鸭血清进行检测,结果表明,本研究建立的NS1-ELISA不能用于区分禽坦布苏自然感染和疫苗免疫血清的鉴别诊断。
- 禽坦布苏病毒 /
- 非结构蛋白NS1 /
- 表达 /
- 间接ELISA
Abstract: Nonstructural protein 1(NS1)from the avian Tembusu virus were amplified and expressed in E.colito establish ELISA procedures for the detection of antibodies in the virus.Biological activity of the expressed protein was analyzed by using SDS-PAGE and western blot.The results indicated that the recombinant NS1 with a molecular weight of 60 kD was expressed in high level with good biological activity.It was subsequently used to establish an indirect ELISA for the detection of antibodies in the virus.The optimized procedures included the uses of 2.5μg·mL^-1 recombination protein,1∶100serum dilution,and the goat anti-duck IgG conjugate(1∶3000).Although the NS1-ELISA showed desirable specificity and repeatability,it failed to effectively differentiate the inactivated Tembusu vaccine serumfrom the naturally infected serum.Thus,this method could not be applied as a diagnosis tool for Tembusu virus.
- avian tembusu virus /
- nonstructural protein 1 /
- expression /
- indirect ELISA

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参考文献(19)

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