动物造模 z-bio 2022-05-09 378

　　Objective To establish a mouse hepatitis virus serum antibody ELISA method for daily monitoring of MHV in this area, so as to timely judge the infection status of SPF mice.

　　Methods Three MHV strains, MHV1, MHV-JHM and MHV-A59, were selected, purified and concentrated antigens were obtained through L929 cell expansion and culture, and the types of antigens suitable for coating were screened; BALB/C mice were immunized to obtain high titer immunopositive serum; optimized The ELISA reaction system established a standardized ELISA kit and used it for the detection of clinical mouse serum samples.

　　RESULTS: The type of MHV coating antigen suitable for this region was screened out as MHV1, and the virus amplification, concentration and purification method was established. The prepared MHV antiserum reached a large number of high titers in the same batch, and could be used as a standardized quality control serum. The optimal working concentrations of the coated antigen, the serum to be tested and the enzyme conjugate were 4.0 μg/mL (10-7.73/0.1 mL TCID50), 1:40 and 1:4000 dilution, respectively; the average coefficient of variation within and between batches were 5.13% and 5.57%, respectively; the detection sensitivity was 1:4000 dilution; with mouse Sendai virus (SV), mouse pneumonia virus (PVM), reovirus type III (Reo3), mouse parvovirus (MVM) No cross-reaction with mousepox virus (Ect) positive serum. The relative deviation of the stability test is less than 10%. A total of 165 serum samples were tested, the coincidence rate of positive serum was 97.37% (37/38), and the coincidence rate of negative serum was 92.19% (118/127).

　　Conclusion The ELISA method established in this study has high specificity, sensitivity and reproducibility for the detection of serum MHV antibodies in mice, and can be used for the daily etiological monitoring of MHV in this area to accurately judge the infection status of mice.