Establishment and evaluation of receptor binding domain (RBD)-based ELISA for Middle East respiratory syndrome coronavirus (MERS-CoV) antibody detection
10.3760/cma.j.issn.0254-5101.2019.10.007
- VernacularTitle: 基于受体结合区(RBD)的ELISA在MERS-CoV抗体检测中的应用评价
- Author:
Wenling WANG
1
;
Ren YANG
1
;
Qianqian SONG
2
;
Huijuan WANG
1
;
Yao DENG
1
;
Li ZHAO
1
;
Wenjie TAN
1
,
3
Author Information
1. National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Key Laboratory of Biosafety, Ministry of Health of the People′s Republic of China, Beijing 102206, China
2. Zhejiang Provincial Key Laboratory of Medical Genetics, School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, Wenzhou 325035, China
3. Zhejiang Provincial Key Laboratory of Medical Genetics, School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, Wenzhou 325035, China
- Publication Type:Journal Article
- Keywords:
Middle East respiratory syndrome coronavirus (MERS-CoV);
Receptor binding domain (RBD);
Antibody;
Enzyme-linked immunosorbent assay (ELISA)
- From:
Chinese Journal of Microbiology and Immunology
2019;39(10):763-770
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish an indirect enzyme-linked immunosorbent assay (ELISA)and to compare the efficiency of receptor binding domain (RBD) proteins in different forms for Middle East respiratory syndrome coronavirus (MERS-CoV) antibody detection.
Methods:The monomeric and trimeric forms of MERS-CoV RBD were expressed in Bac-insect cells, 293T cells and ExpiCHO-S™ expression system and then purified. The purified RBD proteins were identified with native gel electrophoresis and Western blot. Then, an equal amount of each RBD protein was used as coating antigen to establish an ELISA for detecting MERS-CoV IgG titer. For comparison, the newly developed ELISA and the commercial MERS-CoV IgG antibody detection kit (Euroimmune with S1 as the coating antigen) were used to measure the MERS-CoV antibody reference panel supplied by World Health Organization (WHO).
Results:The purified monomeric and trimeric MERS-CoV RBD were successfully prepared using 293T cells and ExpiCHO-S™ system. RBD antigens of different forms and from different systems could recognize MERS-CoV specific antibody without having any cross reaction with the sera from healthy adults. The in-house RBD-based ELISA had good detection consistency with the Euroimmune commercial kit. The positive samples showed higher and more concentrated values based on the RBD trimer than the monomer.
Conclusions:Novel indirect ELISA methods based on the monomeric and trimeric forms of RBD protein were established. The trimetric form-based ELISA achieved higher detection efficiency than the one using the monomer antigen, suggesting that it could be uses as a competent alternative to the commercial kit.
