猫细小病毒抗体ELISA检测方法建立与初步应用  

Establishment and preliminary application of an ELISA method for detecting antibody to feline panleukopenia virus in cats

作　　者：闫文卓 周洁 胡建华 刘铁龙 张圆圆[1] 赵丽丽[1] 陈洪岩[1] 陆涛峰[1] YAN Wenzhuo;ZHOU Jie;HU Jianhua;LIU Tielong;ZHANG Yuanyuan;ZHAO Lili;CHEN Hongyan;LU Taofeng(Heilongjiang Provincial Key Laboratory of Laboratory Animal and Comparative Medicine,Harbin Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Harbin 100193,China;Shanghai Lab.Animal Research Center,Shanghai 201203;Jiyuan Animal Health Inspection,Jiyuan 454650)

机构地区：[1]黑龙江省实验动物与比较医学重点实验室中国农业科学院哈尔滨兽医研究所,哈尔滨100193 [2]上海实验动物研究中心,上海201203 [3]河南省济源市动物卫生监督所,河南济源454650

出　　处：《中国比较医学杂志》2019年第4期98-101,共4页Chinese Journal of Comparative Medicine

基　　金：上海市科委科研计划项目(16140900500)

摘　　要：目的建立猫细小病毒(feline panleukopenia virus,FPV)抗体ELISA检测方法,应用于临床样品中FPV抗体的检测。方法利用猫肾传代细胞(CRFK)增殖FPV病毒,回收的培养物经纯化后制备FPV抗原,通过对纯化抗原和山羊抗猫Ig G-HRP的最佳工作浓度进行确定,建立FPV抗体ELISA检测方法,并进行特异性、敏感性、稳定性测定。结果抗原和二抗的最佳工作浓度分别为5μg/mL和1∶6000,批次内和批次间的变异系数分别为8.68%和7.14%,检测临床血清的灵敏度大于1∶5000,且与猫疱疹病毒1型(feline herpesvirus 1,FHV-1)和猫杯状病毒(feline calicivurus,FCV)均无交叉反应,与国外试剂盒符合率达到90.7%。结论建立的抗体ELISA方法重复性、稳定性良好,特异性和敏感性良好,可用于猫FPV抗体检测。Objective To establish an enzyme-linked immunosorbent assay (ELISA) method for detecting antibody to feline panleukopenia virus (FPV) and apply this method for FPV antibody detection in cats.Methods The FPV antigen was raised in CRFK cells and purified by ultracentrifugation.The purified FPV antigens were coated in microtiter plates and goat anti-cat IgG-HRP was used as the detection antibody.After a series of experiments on the optimization of reaction conditions,the ELISA method was established.Experiments were carried out to evaluate the sensitivity,specificity,repeatability and stability.Results The optimal working densities of the purified FPV antigens and the enzyme-labeled antibody were 5 μg/mL and 1∶6000,respectively.The inter-assay and intra-assay average coefficients of variation were 8.68% and 7.14%,respectively.The detection sensitivity was more than 1 ∶5000.There was no crossreaction with feline herpesvirus type 1 (FHV-1) and feline calicivirus (FCV).Compared with the results of the ELISA kit from the European Veterinary Laboratory (EVL),the concordance between the two methods was 90.7%.Conclusions The established ELISA method exhibites satisfactory duplication,stability,specificity,and sensitivity.This method can be used for the antibody detection of FPV in cats.

关 键 词：猫细小病毒 猫泛白细胞减少症 ELISA 

分 类 号：R-33[医药卫生]