Development and Application of a Nanobody-Based Competitive ELISA for Detecting Antibodies against Hepatitis E Virus from Humans and Domestic Animals
ABSTRACT Hepatitis E virus (HEV) is a zoonotic pathogen that is widespread worldwide. At present, most enzyme-linked immunosorbent assay (ELISA) kits only detect antibodies against human HEV. In this study, a nanobody-horseradish peroxidase (HRP) fusion protein-based competitive ELISA (cELISA) with...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
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American Society for Microbiology
2023-08-01
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| Series: | Microbiology Spectrum |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/spectrum.03607-22 |
| _version_ | 1797741788294807552 |
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| author | Yiyang Chen Meimei Zhang Tianxiang Chen Jiaxi Wang Qin Zhao En-Min Zhou Baoyuan Liu |
| author_facet | Yiyang Chen Meimei Zhang Tianxiang Chen Jiaxi Wang Qin Zhao En-Min Zhou Baoyuan Liu |
| author_sort | Yiyang Chen |
| collection | DOAJ |
| description | ABSTRACT Hepatitis E virus (HEV) is a zoonotic pathogen that is widespread worldwide. At present, most enzyme-linked immunosorbent assay (ELISA) kits only detect antibodies against human HEV. In this study, a nanobody-horseradish peroxidase (HRP) fusion protein-based competitive ELISA (cELISA) with more convenience and spectral characteristics for HEV antibody detection was developed and used to detect HEV IgG in various species. First, 6 anti-swine HEV capsid protein nanobodies were screened using phage display technology from an immunized Bactrian camel. Then, HEV-Nb67-HRP fusions were expressed and used as a probe for developing a cELISA. The cutoff value of the cELISA was 17.8%, and there was no cross-reaction with other anti-swine virus antibodies, suggesting that the cELISA had good specificity. The intra-assay and interassay coefficients of variation (CVs) were 1.33 to 5.06% and 1.52 to 6.84%, respectively. The cELISA and Western blot showed a higher coincidence rate (97.14%, kappa value = 0.927) than cELISA and indirect ELISA (95.00%, kappa value = 0.876) in clinical swine serum samples. Finally, the seroprevalence of HEV IgG in humans, pigs, rabbits, cows, and goats was 30.67%, 19.26%, 8.75%, 27.59%, and 18.08%, respectively, suggesting that cELISA may have a broader scale for mammalian HEV antibody detection. These results suggest that the newly developed cELISA was rapid, low-cost, reliable, and useful for the serological evaluation of current HEV. IMPORTANCE HEV is thought to be a zoonotic infection and is widespread worldwide; it is beneficial to establish a more convenient and spectral method for HEV antibody detection. In this study, a convenient, time-saving, reproducible, highly sensitive, specific, and novel nanobody-based cELISA was developed and can be used to detect IgG antibodies against mammalian HEV. It provides a new technique for serological evaluation and ELISA-based diagnosis of HEV infection. |
| first_indexed | 2024-03-12T14:31:44Z |
| format | Article |
| id | doaj.art-c740c98872374d47b622560d8ae41f37 |
| institution | Directory Open Access Journal |
| issn | 2165-0497 |
| language | English |
| last_indexed | 2024-03-12T14:31:44Z |
| publishDate | 2023-08-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | Microbiology Spectrum |
| spelling | doaj.art-c740c98872374d47b622560d8ae41f372023-08-17T13:04:14ZengAmerican Society for MicrobiologyMicrobiology Spectrum2165-04972023-08-0111410.1128/spectrum.03607-22Development and Application of a Nanobody-Based Competitive ELISA for Detecting Antibodies against Hepatitis E Virus from Humans and Domestic AnimalsYiyang Chen0Meimei Zhang1Tianxiang Chen2Jiaxi Wang3Qin Zhao4En-Min Zhou5Baoyuan Liu6Department of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, ChinaDepartment of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, ChinaDepartment of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, ChinaDepartment of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, ChinaDepartment of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, ChinaDepartment of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, ChinaDepartment of Preventive Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, ChinaABSTRACT Hepatitis E virus (HEV) is a zoonotic pathogen that is widespread worldwide. At present, most enzyme-linked immunosorbent assay (ELISA) kits only detect antibodies against human HEV. In this study, a nanobody-horseradish peroxidase (HRP) fusion protein-based competitive ELISA (cELISA) with more convenience and spectral characteristics for HEV antibody detection was developed and used to detect HEV IgG in various species. First, 6 anti-swine HEV capsid protein nanobodies were screened using phage display technology from an immunized Bactrian camel. Then, HEV-Nb67-HRP fusions were expressed and used as a probe for developing a cELISA. The cutoff value of the cELISA was 17.8%, and there was no cross-reaction with other anti-swine virus antibodies, suggesting that the cELISA had good specificity. The intra-assay and interassay coefficients of variation (CVs) were 1.33 to 5.06% and 1.52 to 6.84%, respectively. The cELISA and Western blot showed a higher coincidence rate (97.14%, kappa value = 0.927) than cELISA and indirect ELISA (95.00%, kappa value = 0.876) in clinical swine serum samples. Finally, the seroprevalence of HEV IgG in humans, pigs, rabbits, cows, and goats was 30.67%, 19.26%, 8.75%, 27.59%, and 18.08%, respectively, suggesting that cELISA may have a broader scale for mammalian HEV antibody detection. These results suggest that the newly developed cELISA was rapid, low-cost, reliable, and useful for the serological evaluation of current HEV. IMPORTANCE HEV is thought to be a zoonotic infection and is widespread worldwide; it is beneficial to establish a more convenient and spectral method for HEV antibody detection. In this study, a convenient, time-saving, reproducible, highly sensitive, specific, and novel nanobody-based cELISA was developed and can be used to detect IgG antibodies against mammalian HEV. It provides a new technique for serological evaluation and ELISA-based diagnosis of HEV infection.https://journals.asm.org/doi/10.1128/spectrum.03607-22hepatitis E virus (HEV)cELISAantibodieshumansdomestic animals |
| spellingShingle | Yiyang Chen Meimei Zhang Tianxiang Chen Jiaxi Wang Qin Zhao En-Min Zhou Baoyuan Liu Development and Application of a Nanobody-Based Competitive ELISA for Detecting Antibodies against Hepatitis E Virus from Humans and Domestic Animals Microbiology Spectrum hepatitis E virus (HEV) cELISA antibodies humans domestic animals |
| title | Development and Application of a Nanobody-Based Competitive ELISA for Detecting Antibodies against Hepatitis E Virus from Humans and Domestic Animals |
| title_full | Development and Application of a Nanobody-Based Competitive ELISA for Detecting Antibodies against Hepatitis E Virus from Humans and Domestic Animals |
| title_fullStr | Development and Application of a Nanobody-Based Competitive ELISA for Detecting Antibodies against Hepatitis E Virus from Humans and Domestic Animals |
| title_full_unstemmed | Development and Application of a Nanobody-Based Competitive ELISA for Detecting Antibodies against Hepatitis E Virus from Humans and Domestic Animals |
| title_short | Development and Application of a Nanobody-Based Competitive ELISA for Detecting Antibodies against Hepatitis E Virus from Humans and Domestic Animals |
| title_sort | development and application of a nanobody based competitive elisa for detecting antibodies against hepatitis e virus from humans and domestic animals |
| topic | hepatitis E virus (HEV) cELISA antibodies humans domestic animals |
| url | https://journals.asm.org/doi/10.1128/spectrum.03607-22 |
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