病毒学报

为了建立一种能够检测猫泛白细胞减少症病毒(Feline panleukopenia virus, FPV)的双抗体夹心酶联检测(ELISA)方法，本研究首先构建了FPV VP2蛋白的原核表达载体并进行了VP2蛋白的原核表达。经SDS-PAGE实验验证及反应原性检测后，以VP2重组蛋白作为免疫原，通过间接ELISA方法筛选出2株能特异性识别VP2蛋白的高亲和力单克隆抗体，分别命名为2F10和3D2。通过对捕获抗体包被浓度、酶标检测抗体使用浓度等最适反应条件进行摸索，建立了FPV的双抗体夹心ELISA检测方法，并对方法的敏感性、特异性及与PCR方法的符合率进行验证。结果显示该方法的最低检测限为10^3.2 TCID_50/mL，高于市售商品化试纸条，且与猫杯状病毒(Feline calici virus,FCV)和猫疱疹病毒1型(Feline herpesvirus type 1, FHV-Ι)均无交叉反应，特异性较好，与FPV标准检测方法 PCR的符合率为98.9%，且操作简便、检测快速，为FPV的检测提供了一种有效的工具，为进一步开发FPV检测试剂盒奠定了基础。

To establish a double-antibody sandwich enzyme-linked immunosorbent assay(ELISA) for the detection of feline panleukopenia virus(FPV), a prokaryotic expression vector of FPV VP2 protein was constructed and the VP2 protein was expressed in prokaryotic cells. Following SDS-PAGE verification and antigenicity testing, two high-affinity monoclonal antibodies that specifically recognize the VP2 protein were selected through indirect ELISA using recombinant VP2 protein as the immunogen. These antibodies were designated 2F10 and 3D2. The optimal reaction conditions, including the coating concentration of the capture antibody and the detection antibody concentration, were optimized to establish a double-antibody sandwich ELISA for FPV. The sensitivity, specificity, and concordance with the PCR method were evaluated. The results indicated that the method has a detection limit of 10^3.2 TCID_50/mL, which is superior to that of commercial test strips. No cross-reaction was observed with feline calicivirus(FCV) or feline herpesvirus type 1(FHV-1), demonstrating good specificity. The method showed a concordance rate of 98.9% with the standard PCR detection method for FPV. This ELISA method is simple, rapid, highly sensitive, and specific, providing an effective tool for FPV detection and laying the foundation for the development of diagnostic kits for feline panleukopenia