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Establishment of a Blocking ELISA Detection Method for Against African Swine Fever Virus p30 Antibody

African swine fever (ASF) is a highly lethal hemorrhagic viral disease of domestic pigs caused by African swine fever virus (ASFV). A sensitive and reliable serological diagnostic assay is required, so laboratories can effectively and quickly detect ASFV infection. The p30 protein is abundantly expr...

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Autores principales: Yu, Xuexiang, Zhu, Xianjing, Chen, Xiaoyu, Li, Dongfan, Xu, Qian, Yao, Lun, Sun, Qi, Ghonaim, Ahmed H., Ku, Xugang, Fan, Shengxian, Yang, Hanchun, He, Qigai
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8718596/
https://www.ncbi.nlm.nih.gov/pubmed/34977214
http://dx.doi.org/10.3389/fvets.2021.781373
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author Yu, Xuexiang
Zhu, Xianjing
Chen, Xiaoyu
Li, Dongfan
Xu, Qian
Yao, Lun
Sun, Qi
Ghonaim, Ahmed H.
Ku, Xugang
Fan, Shengxian
Yang, Hanchun
He, Qigai
author_facet Yu, Xuexiang
Zhu, Xianjing
Chen, Xiaoyu
Li, Dongfan
Xu, Qian
Yao, Lun
Sun, Qi
Ghonaim, Ahmed H.
Ku, Xugang
Fan, Shengxian
Yang, Hanchun
He, Qigai
author_sort Yu, Xuexiang
collection PubMed
description African swine fever (ASF) is a highly lethal hemorrhagic viral disease of domestic pigs caused by African swine fever virus (ASFV). A sensitive and reliable serological diagnostic assay is required, so laboratories can effectively and quickly detect ASFV infection. The p30 protein is abundantly expressed early in cells and has excellent antigenicity. Therefore, this study aimed to produce and characterize p30 monoclonal antibodies with an ultimate goal of developing a monoclonal antibody-based enzyme-linked immunosorbent assay (ELISA) for ASFV antibody detection. Three monoclonal antibodies against p30 protein that were expressed in E. coli were generated, and their characterizations were investigated. Furthermore, a blocking ELISA based on a monoclonal antibody was developed. To evaluate the performance of the assay, 186 sera samples (88 negative and 98 positive samples) were analyzed and a receiver-operating characteristic (ROC) analysis was applied to determine the cutoff value. Based on the ROC analysis, the area under the curve (AUC) was 0.997 (95% confidence interval: 99.2 to 100%). Besides, a diagnostic sensitivity of 97.96% (95% confidence interval: 92.82 to 99.75%) and a specificity of 98.96% (95% confidence interval: 93.83 to 99.97%) were achieved when the cutoff value was set to 38.38%. Moreover, the coefficients of inter- and intra-batches were <10%, indicating the good repeatability of the method. The maximum dilution of positive standard serum detected by this ELISA method was 1:512. The blocking ELISA was able to detect seroconversion in two out of five pigs at 10 Dpi and the p30 response increasing trend through the time course of the study (0–20 Dpi). In conclusion, the p30 mAb-based blocking ELISA developed in this study demonstrated a high repeatability with maximized diagnostic sensitivity and specificity. The assay could be a useful tool for field surveillance and epidemiological studies in swine herd.
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spelling pubmed-87185962022-01-01 Establishment of a Blocking ELISA Detection Method for Against African Swine Fever Virus p30 Antibody Yu, Xuexiang Zhu, Xianjing Chen, Xiaoyu Li, Dongfan Xu, Qian Yao, Lun Sun, Qi Ghonaim, Ahmed H. Ku, Xugang Fan, Shengxian Yang, Hanchun He, Qigai Front Vet Sci Veterinary Science African swine fever (ASF) is a highly lethal hemorrhagic viral disease of domestic pigs caused by African swine fever virus (ASFV). A sensitive and reliable serological diagnostic assay is required, so laboratories can effectively and quickly detect ASFV infection. The p30 protein is abundantly expressed early in cells and has excellent antigenicity. Therefore, this study aimed to produce and characterize p30 monoclonal antibodies with an ultimate goal of developing a monoclonal antibody-based enzyme-linked immunosorbent assay (ELISA) for ASFV antibody detection. Three monoclonal antibodies against p30 protein that were expressed in E. coli were generated, and their characterizations were investigated. Furthermore, a blocking ELISA based on a monoclonal antibody was developed. To evaluate the performance of the assay, 186 sera samples (88 negative and 98 positive samples) were analyzed and a receiver-operating characteristic (ROC) analysis was applied to determine the cutoff value. Based on the ROC analysis, the area under the curve (AUC) was 0.997 (95% confidence interval: 99.2 to 100%). Besides, a diagnostic sensitivity of 97.96% (95% confidence interval: 92.82 to 99.75%) and a specificity of 98.96% (95% confidence interval: 93.83 to 99.97%) were achieved when the cutoff value was set to 38.38%. Moreover, the coefficients of inter- and intra-batches were <10%, indicating the good repeatability of the method. The maximum dilution of positive standard serum detected by this ELISA method was 1:512. The blocking ELISA was able to detect seroconversion in two out of five pigs at 10 Dpi and the p30 response increasing trend through the time course of the study (0–20 Dpi). In conclusion, the p30 mAb-based blocking ELISA developed in this study demonstrated a high repeatability with maximized diagnostic sensitivity and specificity. The assay could be a useful tool for field surveillance and epidemiological studies in swine herd. Frontiers Media S.A. 2021-12-17 /pmc/articles/PMC8718596/ /pubmed/34977214 http://dx.doi.org/10.3389/fvets.2021.781373 Text en Copyright © 2021 Yu, Zhu, Chen, Li, Xu, Yao, Sun, Ghonaim, Ku, Fan, Yang and He. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Veterinary Science
Yu, Xuexiang
Zhu, Xianjing
Chen, Xiaoyu
Li, Dongfan
Xu, Qian
Yao, Lun
Sun, Qi
Ghonaim, Ahmed H.
Ku, Xugang
Fan, Shengxian
Yang, Hanchun
He, Qigai
Establishment of a Blocking ELISA Detection Method for Against African Swine Fever Virus p30 Antibody
title Establishment of a Blocking ELISA Detection Method for Against African Swine Fever Virus p30 Antibody
title_full Establishment of a Blocking ELISA Detection Method for Against African Swine Fever Virus p30 Antibody
title_fullStr Establishment of a Blocking ELISA Detection Method for Against African Swine Fever Virus p30 Antibody
title_full_unstemmed Establishment of a Blocking ELISA Detection Method for Against African Swine Fever Virus p30 Antibody
title_short Establishment of a Blocking ELISA Detection Method for Against African Swine Fever Virus p30 Antibody
title_sort establishment of a blocking elisa detection method for against african swine fever virus p30 antibody
topic Veterinary Science
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8718596/
https://www.ncbi.nlm.nih.gov/pubmed/34977214
http://dx.doi.org/10.3389/fvets.2021.781373
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