Establishment and application of a luciferase immunosorbent assay for the detection of antibodies to Crimean-Congo hemorrhagic fever virus
10.3969/j.issn.1002-2694.2025.00.042
- VernacularTitle:克里米亚—刚果出血热病毒抗体荧光素酶免疫吸附法检测方法的建立与应用
- Author:
Qi CHEN
1
;
Jin-zhe MA
1
;
Li-tai XU
1
;
Xin-yue LI
1
;
Yu-ting FANG
1
;
Cheng-song WAN
1
Author Information
1. 南方医科大学公共卫生学院,广州 510515
- Publication Type:Journal Article
- Keywords:
Crimean-Congo hemorrhagic fever virus;
Gc protein;
luciferase immunosorbent assay;
IgG antibody;
detection
- From:
Chinese Journal of Zoonoses
2025;41(3):290-296
- CountryChina
- Language:Chinese
-
Abstract:
The purpose of this study was to establish a luciferase immunosorbent assay(LISA)using the Crimean-Congo hemorrhagic fever virus(CCHFV)glycoprotein C(Gc),a specific antigen,for the detection of CCHFV IgG antibodies.Three antigenic fragments based on CCHFV glycoprotein C were designed,and three recombinant plasmids were constructed by liga-tion with the NanoLuc luciferase(NLuc)expression vector pNLF1-N through molecular cloning.The accuracy of the sequences in the recombinant plasmids was confirmed through sequencing.The recombinant plasmids were transfected into eukaryotic cells to obtain fusion proteins containing specific antigens and luciferase,and the expression of the fusion proteins was verified by western blotting,thereby facilitating the establishment of the CCHFV-LISA detection technique.The assay's sensitivity,specificity,and stability were evaluated and compared with those of a commercial CCHFV IgG antibody test kit.Three recom-binant antigen fragments of CCHFV Gc—NLuc-Gc-Full,NLuc-Gc-C1,and NLuc-Gc-C2—were expressed,with molecular weights of 80.1 kDa,62.8 kDa,and 53.9 kDa,respectively.The optimal fragment for CCHFV detection was NLuc-Gc-C2.The sensitivity of the CCHFV-LISA was 90.9％,and the specificity was 100％;the findings were highly concordant with those for the commercial CCHFV enzyme-linked immunosorbent assay kit.Repeatability tests indicated no statistically significant differ-ences in inter-and intra-assay variability within the same batch.The LISA was highly specific,sensitive,and user-friendly in detecting IgG antibodies against the CCHFV.Therefore,this method may facilitate serological diagnosis and epidemiological studies in endemic regions,and provide essential technical support for disease surveillance and early warning.
