A Quantitative Method for Detecting Ara h 2 by Generation and Utilization of Monoclonal Antibodies
Peanut (Arachis hypogaea) is one of the most common food allergens that can induce fatal anaphylaxis, and Ara h 2 is one of the major allergen components involved in peanut allergy. The aim of this study was to develop a quantitative method for detecting peanut allergen using monoclonal antibodies a...
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| Format: | Article |
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Wiley
2018-01-01
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| Series: | Journal of Immunology Research |
| Online Access: | http://dx.doi.org/10.1155/2018/4894705 |
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| author | Huifang Chen Zehong Zou Ailin Tao |
| author_facet | Huifang Chen Zehong Zou Ailin Tao |
| author_sort | Huifang Chen |
| collection | DOAJ |
| description | Peanut (Arachis hypogaea) is one of the most common food allergens that can induce fatal anaphylaxis, and Ara h 2 is one of the major allergen components involved in peanut allergy. The aim of this study was to develop a quantitative method for detecting peanut allergen using monoclonal antibodies against Ara h 2. The splenocytes of immunized mice were fused with myeloma cells (SP2/0), and stable mAb-producing clones were obtained by limiting dilution. mAbs against Ara h 2 were isolated from mouse ascites, and specificity was confirmed by immunoblotting. Five mAbs with high purity and specific reactivity were obtained, which were referred to as 1-2E10, 2-1D5, 3-1C5, 4-1C2, and 5-1G4, respectively. After screening different mAb combinations for development of a sandwich ELISA, we selected 5-1G4 as the capture antibody and 1-2E10 as the detection antibody for the measurement of Ara h 2 from which an optimal correlation between the Ara h 2 concentration and the OD value was obtained. This sandwich ELISA could specifically detect Ara h 2 in peanut extract at concentrations as low as 5 ng/mL and up to 10 μg/mL. These mAbs can, therefore, serve as quantitative diagnostic reagents for peanut and peanut product risk assessment. |
| format | Article |
| id | doaj-art-2b708fa20dab4bfb8aefbcfacfd02bba |
| institution | Kabale University |
| issn | 2314-8861 2314-7156 |
| language | English |
| publishDate | 2018-01-01 |
| publisher | Wiley |
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| series | Journal of Immunology Research |
| spelling | doaj-art-2b708fa20dab4bfb8aefbcfacfd02bba2025-02-03T06:06:07ZengWileyJournal of Immunology Research2314-88612314-71562018-01-01201810.1155/2018/48947054894705A Quantitative Method for Detecting Ara h 2 by Generation and Utilization of Monoclonal AntibodiesHuifang Chen0Zehong Zou1Ailin Tao2The Second Affiliated Hospital, The State Key Laboratory of Respiratory Disease, Guangdong Provincial Key Laboratory of Allergy & Clinical Immunology, Guangzhou Medical University, Guangzhou, ChinaThe Second Affiliated Hospital, The State Key Laboratory of Respiratory Disease, Guangdong Provincial Key Laboratory of Allergy & Clinical Immunology, Guangzhou Medical University, Guangzhou, ChinaThe Second Affiliated Hospital, The State Key Laboratory of Respiratory Disease, Guangdong Provincial Key Laboratory of Allergy & Clinical Immunology, Guangzhou Medical University, Guangzhou, ChinaPeanut (Arachis hypogaea) is one of the most common food allergens that can induce fatal anaphylaxis, and Ara h 2 is one of the major allergen components involved in peanut allergy. The aim of this study was to develop a quantitative method for detecting peanut allergen using monoclonal antibodies against Ara h 2. The splenocytes of immunized mice were fused with myeloma cells (SP2/0), and stable mAb-producing clones were obtained by limiting dilution. mAbs against Ara h 2 were isolated from mouse ascites, and specificity was confirmed by immunoblotting. Five mAbs with high purity and specific reactivity were obtained, which were referred to as 1-2E10, 2-1D5, 3-1C5, 4-1C2, and 5-1G4, respectively. After screening different mAb combinations for development of a sandwich ELISA, we selected 5-1G4 as the capture antibody and 1-2E10 as the detection antibody for the measurement of Ara h 2 from which an optimal correlation between the Ara h 2 concentration and the OD value was obtained. This sandwich ELISA could specifically detect Ara h 2 in peanut extract at concentrations as low as 5 ng/mL and up to 10 μg/mL. These mAbs can, therefore, serve as quantitative diagnostic reagents for peanut and peanut product risk assessment.http://dx.doi.org/10.1155/2018/4894705 |
| spellingShingle | Huifang Chen Zehong Zou Ailin Tao A Quantitative Method for Detecting Ara h 2 by Generation and Utilization of Monoclonal Antibodies Journal of Immunology Research |
| title | A Quantitative Method for Detecting Ara h 2 by Generation and Utilization of Monoclonal Antibodies |
| title_full | A Quantitative Method for Detecting Ara h 2 by Generation and Utilization of Monoclonal Antibodies |
| title_fullStr | A Quantitative Method for Detecting Ara h 2 by Generation and Utilization of Monoclonal Antibodies |
| title_full_unstemmed | A Quantitative Method for Detecting Ara h 2 by Generation and Utilization of Monoclonal Antibodies |
| title_short | A Quantitative Method for Detecting Ara h 2 by Generation and Utilization of Monoclonal Antibodies |
| title_sort | quantitative method for detecting ara h 2 by generation and utilization of monoclonal antibodies |
| url | http://dx.doi.org/10.1155/2018/4894705 |
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