Establishment of an indirect immunofluorescence assay for detection of murine norovirus
10.3969/j.issn.1671.7856.2014.006.013
- VernacularTitle:小鼠诺如病毒间接免疫荧光检测方法的建立
- Author:
Shengnan TIAN
;
Wei TONG
;
Lifang ZHANG
;
Hui CHANG
;
Yuhan LI
;
Jingfen SU
;
Xianju LIU
;
Zhiguang XIANG
;
Yunbo LIU
- Publication Type:Journal Article
- Keywords:
Indirect immunofluorescence detection;
Murine Norovirus
- From:
Chinese Journal of Comparative Medicine
2014;(6):58-62
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish an indirect immunofluorescence assay for detection of murine norovirus ( MNV) .Methods Mouse leukaemic monocyte macrophage cell line RAW 264.7 cells were infected with MNV-1 and cultured for 36 hours to collect the virus and uninfected cells , and to make antigen glass slides .BALB/c mice were gavaged with MNV-1 (107 TCID50) and infected sera were collected as positive control .The serum was 1:10 diluted and used for measuring MNV antibody by immunofluorescence assay ( IFA ) .80 serum samples were tested using the two methods , IFA and ELISA, and the discrepant samples were validated by Western blotting .Results RAW264.7 cells were infected with MNV-1 for 36-48 h, showing an infection rate of 60% of the cells, and the cells infected for 36 h were preferred.IFA method was used to detect the serum with MNV-1 infection and showed that the antibody content was gradually increased at one week after infection , reaching a maximum antibody concentration at 4 weeks after infection , and maintained a stable level later .The mouse serum at four weeks after MNV-1infection was used as positive quality control . Among the 80 serum samples , 27 positive and 53 negative cases were detected by IFA method , and 32 positive and 48 negative cases were detected by ELISA .The five discrepant samples were verified by Western blotting , resulted in 3 positive and 2 negative cases . The coincidence rate of IFA was 96.0% and that of ELISA methods was 97.5%. Conclusions Basically, immunofluorescence assay can be used to detect the MNV-1 infection in mice, although false negative result may occur occasionally .IFA and ELISA detection can be selected as initial screening measures , and use Western blot assay to verify the discrepant samples .
