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An Extremely Highly Sensitive ELISA in pg mL(−1) Level Based on a Newly Produced Monoclonal Antibody for the Detection of Ochratoxin A in Food Samples
In this study, an extremely highly sensitive enzyme-linked immunosorbent assay (ELISA) based on a newly produced monoclonal antibody (mAb) for the detection of ochratoxin A (OTA) in food samples was developed. OTA-Bovine serum albumin (BSA) conjugate was prepared and used as the immunogen for the pr...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10420233/ https://www.ncbi.nlm.nih.gov/pubmed/37570711 http://dx.doi.org/10.3390/molecules28155743 |
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author | Ren, Yexuan Tian, Ruwen Wang, Ting Cao, Junlin Li, Jianguo Deng, Anping |
author_facet | Ren, Yexuan Tian, Ruwen Wang, Ting Cao, Junlin Li, Jianguo Deng, Anping |
author_sort | Ren, Yexuan |
collection | PubMed |
description | In this study, an extremely highly sensitive enzyme-linked immunosorbent assay (ELISA) based on a newly produced monoclonal antibody (mAb) for the detection of ochratoxin A (OTA) in food samples was developed. OTA-Bovine serum albumin (BSA) conjugate was prepared and used as the immunogen for the production of the mAb. Among four hybridoma clones (8B10, 5C2, 9B7, and 5E11), the antibody from 8B10 displayed the highest affinity recognition for OTA. Based on the mAb (8B10), the IC(50) and LOD of the ELISA for OTA were 34.8 pg mL(−1) and 1.5 pg mL(−1), respectively, which was 1.53~147 times lower than those in published ELISAs, indicating the ultra-sensitivity of our assay. There was no cross-reactivity of the mAb with the other four mycotoxins (AFB(1), ZEN, DON, and T-2). Due to the high similarity in molecular structures among OTA, ochratoxin B (OTB), and ochratoxin C (OTC), the CR values of the mAb with OTB and OTC were 96.67% and 22.02%, respectively. Taking this advantage, the ELISA may be able to evaluate total ochratoxin levels in food samples. The recoveries of the ELISA for OTA in spiked samples (corn, wheat, and feed) were 96.5–110.8%, 89.5–94.4%, and 91.8–113.3%; and the RSDs were 5.2–13.6%, 8.2–13.0%, and 7.7–13.7% (n = 3), respectively. The spiked food samples (corn) were measured by ELISA and HPLC-FLD simultaneously. A good correlation between ELISA (x) and HPLC-FLD (y) with the linear regression equation of y = 0.918x − 0.034 (R(2) = 0.985, n = 5) was obtained. These results demonstrated that the newly produced mAb-based ELISA was a feasible and ultra-sensitive analytical method for the detection of OTA in food samples. |
format | Online Article Text |
id | pubmed-10420233 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-104202332023-08-12 An Extremely Highly Sensitive ELISA in pg mL(−1) Level Based on a Newly Produced Monoclonal Antibody for the Detection of Ochratoxin A in Food Samples Ren, Yexuan Tian, Ruwen Wang, Ting Cao, Junlin Li, Jianguo Deng, Anping Molecules Article In this study, an extremely highly sensitive enzyme-linked immunosorbent assay (ELISA) based on a newly produced monoclonal antibody (mAb) for the detection of ochratoxin A (OTA) in food samples was developed. OTA-Bovine serum albumin (BSA) conjugate was prepared and used as the immunogen for the production of the mAb. Among four hybridoma clones (8B10, 5C2, 9B7, and 5E11), the antibody from 8B10 displayed the highest affinity recognition for OTA. Based on the mAb (8B10), the IC(50) and LOD of the ELISA for OTA were 34.8 pg mL(−1) and 1.5 pg mL(−1), respectively, which was 1.53~147 times lower than those in published ELISAs, indicating the ultra-sensitivity of our assay. There was no cross-reactivity of the mAb with the other four mycotoxins (AFB(1), ZEN, DON, and T-2). Due to the high similarity in molecular structures among OTA, ochratoxin B (OTB), and ochratoxin C (OTC), the CR values of the mAb with OTB and OTC were 96.67% and 22.02%, respectively. Taking this advantage, the ELISA may be able to evaluate total ochratoxin levels in food samples. The recoveries of the ELISA for OTA in spiked samples (corn, wheat, and feed) were 96.5–110.8%, 89.5–94.4%, and 91.8–113.3%; and the RSDs were 5.2–13.6%, 8.2–13.0%, and 7.7–13.7% (n = 3), respectively. The spiked food samples (corn) were measured by ELISA and HPLC-FLD simultaneously. A good correlation between ELISA (x) and HPLC-FLD (y) with the linear regression equation of y = 0.918x − 0.034 (R(2) = 0.985, n = 5) was obtained. These results demonstrated that the newly produced mAb-based ELISA was a feasible and ultra-sensitive analytical method for the detection of OTA in food samples. MDPI 2023-07-29 /pmc/articles/PMC10420233/ /pubmed/37570711 http://dx.doi.org/10.3390/molecules28155743 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Ren, Yexuan Tian, Ruwen Wang, Ting Cao, Junlin Li, Jianguo Deng, Anping An Extremely Highly Sensitive ELISA in pg mL(−1) Level Based on a Newly Produced Monoclonal Antibody for the Detection of Ochratoxin A in Food Samples |
title | An Extremely Highly Sensitive ELISA in pg mL(−1) Level Based on a Newly Produced Monoclonal Antibody for the Detection of Ochratoxin A in Food Samples |
title_full | An Extremely Highly Sensitive ELISA in pg mL(−1) Level Based on a Newly Produced Monoclonal Antibody for the Detection of Ochratoxin A in Food Samples |
title_fullStr | An Extremely Highly Sensitive ELISA in pg mL(−1) Level Based on a Newly Produced Monoclonal Antibody for the Detection of Ochratoxin A in Food Samples |
title_full_unstemmed | An Extremely Highly Sensitive ELISA in pg mL(−1) Level Based on a Newly Produced Monoclonal Antibody for the Detection of Ochratoxin A in Food Samples |
title_short | An Extremely Highly Sensitive ELISA in pg mL(−1) Level Based on a Newly Produced Monoclonal Antibody for the Detection of Ochratoxin A in Food Samples |
title_sort | extremely highly sensitive elisa in pg ml(−1) level based on a newly produced monoclonal antibody for the detection of ochratoxin a in food samples |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10420233/ https://www.ncbi.nlm.nih.gov/pubmed/37570711 http://dx.doi.org/10.3390/molecules28155743 |
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