猴免疫缺陷病毒抗体免疫梳检测方法的建立及应用
作者:
作者单位:

(1. 海南出入境检验检疫局检验检疫技术中心,海口 570311; 2. 福建出入境检验检疫局检验检疫技术中心,福州 350001; 3. 深圳出入境检验检疫局动植物检验检疫技术中心,广东深圳 518001; 4. 黑龙江出入境检验检疫局检验检疫技术中心,哈尔滨 150001; 5. 辽宁医学院畜牧兽医学院,辽宁锦州 121001)

作者简介:

通讯作者:

中图分类号:

基金项目:

国家质检总局科技项目(No. 2013IK031,No. 2013IK051,No. 2015IK089);福建省科技计划重点项目(No. 2014N0001);海南省应用技术研究与开发专项项目(No. ZDXM20130025);广东检验检疫局科技计划项目(No. 2011GDK44,No. 2013GDK04)


Establishment and application of a new immune-comb assay for detection of serum antibody against simian immunodeficiency virus
Author:
Affiliation:

(1. Technical Center of Hainan Entry-Exit Inspection and Quarantine Bureau,Haikou 570311,China. 2. Technical Center of Fujian Entry-Exit Inspection and Quarantine Bureau,Fuzhou 35000. 3. Technical Center of Animal and Plant Inspection and Quarantine, Shenzhen Entry-Exit Inspection and Quarantine Bureau, Shengzhen 518001. 4. Technical Center of Heilongjiang Entry-Exit Inspection and Quarantine Bureau,Harbin 150001. 5. Department of Animal Husbandry and Veterinary Medicine, Liaoning Medical University, Jinzhou 121001)

Fund Project:

Science and Technology Project of AQSIQ(No. 2013IK031,No. 2013IK051,No. 2015IK089), Key Projects of Science and Technology Plan of Fujian Province ( No. 2014N0001 ), Special Project on Applied Technology Research and Development in Hainan Province ( No.ZDXM20130025), Science and Technology Project of Guangdong Inspection and Quarantine Bureau(No. 2011GDK44,No. 2013GDK04)

  • 摘要
  • |
  • 图/表
  • |
  • 访问统计
  • |
  • 参考文献
  • |
  • 相似文献
  • |
  • 引证文献
  • |
  • 资源附件
  • |
  • 文章评论
    摘要:

    目的 以基因工程技术制备猴免疫缺陷病毒(SIV)的SIV30 蛋白作为抗原诊断试剂,建立免疫梳方法(IC)用于特异性检测实验猴血清中抗SIV 的抗体IgG?方法 应用原核表达载体pGEX -4T -1 构建SIV SIV30基因的重组表达质粒,并在感受态细胞BL21 中表达,将该重组蛋白纯化后作为诊断抗原,建立免疫梳标准化检测程序,并应用于临床检测?文字结果 最佳抗原包被量为0.02 mg/ mL;制备好的IC 均能够特异性检测到相应的SIV 阳性血清而不与其他病毒血清间发生交叉反应,表明该检测方法特异性强;敏感性分析结果表明,SIV30 蛋白能够敏感地检测到1∶ 400 倍稀释的SIV 阳性血清;稳定性和重复性试验结果表明,同一样品重复检测3 次,变异系数(CV)均小于10%;利用该检测方法在对10 份可疑猴血清样品进行检测,IC 与ELISA 检测结果一致率为100.0%,Kappa=1.000?文字结论 原核表达了SIV30 蛋白,制备了IC,并应用于临床检测?该检测方法灵敏度高?特异性强?重复性好,具有良好的实用性?

    Abstract:

    Objective SIV30 protein of simian immunodeficiency virus ( SIV) was prepared by genetic engineering technique as an antigen diagnostic reagent, to establish an immune comb method for the specific detection of anti SIV IgG in monkey serum. Methods Recombinant expression plasmid of SIV SIV30 gene was constructed by prokaryotic expression vector pGEX-4T-1, and expressed in the competent BL21 cells. The recombinant protein was purified as a diagnostic antigen, and a standardized procedure for the detection of immune comb was established and applied for clinical detection. Results The optimum coating amount of antigen was 0.02 mg/ mL. The prepared IC was able to specifically detect the positive serum of SIV. There was no cross reaction between the sera of other viruses. It showed a high specificity of the detection method. Sensitivity analysis showed that the SIV30 protein was able to detect 1∶400 times diluted SIV positive sera. The result of stability and repeatability test (the same sample was repeated 3 times) showed that the coefficient of variation (CV) was less than 10%. The serum samples of 10 suspicious monkeys were detected by this method , showing a consistent rate of comb method and ELISA test result of 100%, Kappa = 1.000. Conclusions SIV30 protein is expressed in prokaryotic cells. The immune comb is prepared, and is successfullyl applied in clinical examination. It shows that the method has a high sensitivity, strong specificity, good reproducibility and practicability.

    参考文献
    相似文献
    引证文献
引用本文

李丹丹,王绥家,陈平亚,张体银,杨俊兴,刘忠梅,高慎阳.猴免疫缺陷病毒抗体免疫梳检测方法的建立及应用[J].中国实验动物学报,2018,26(2):217~223.

复制
分享
文章指标
  • 点击次数:
  • 下载次数:
  • HTML阅读次数:
  • 引用次数:
历史
  • 收稿日期:2017-10-24
  • 最后修改日期:
  • 录用日期:
  • 在线发布日期: 2018-05-14
  • 出版日期:
征稿啦 |“科技伦理前沿谈”征文通知
关闭