A Quantitative Method for Detecting Ara h 2 by Generation and Utilization of Monoclonal Antibodies

Chen, Huifang; Zou, Zehong; Tao, Ailin

doi:10.1155/2018/4894705

Cited by 4 publications

(2 citation statements)

References 39 publications

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“…We also compared the sensitivities and target antigens of the developed indirect ELISA to immunoassays reported in other reports [ 36 - 38 ] ( Table 2 ). The lateral flow immunoassay is specific to native purified Ara h 1, and sandwich ELISA based on a monoclonal antibody specific to recombinant Ara h 2 of peanuts has been reported previously.…”

Section: Resultsmentioning

confidence: 99%

Detection of a Thermal Stable-Soluble Protein (TSSP) as a Marker of Peanut Adulteration Using a Highly Sensitive Indirect Enzyme-Linked Immunosorbent Assay based on Monoclonal Antibodies

Kim

Toushik

Lee

et al. 2023

J. Microbiol. Biotechnol.

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Food allergy represents a severe problem for many societies, including sensitive populations, academies, health authorities, and the food industry. Peanut allergy occupies a special place in the food allergy spectrum. To prevent consumption by consumers suffering from a peanut allergy, a rapid and sensitive detection method is essential to identify unintended peanut adulteration in processed foods. In this study, we produced four monoclonal antibodies (MAbs; RO 3A1-12, PB 4C12-10, PB 5F9-23, and PB 6G4-30) specific to thermo-stable and soluble proteins (TSSPs) of peanut and developed an enzyme-linked immunosorbent assay (ELISA) based on the MAbs. Among them, PB 5F9-23 MAb was firmly bound to Ara h 1, and other MAbs strongly reacted to Ara h 3 in the Western blot analysis. An antibody cocktail solution of the MAbs was used to enhance the sensitivity of an indirect ELISA, and the limit of detection of the indirect ELISA based on the antibody cocktail solution was 1 ng/ml and improved compared to the indirect ELISA based on the single MAb (11 ng/ml). The cross-reaction analysis revealed the high specificity of developed MAbs to peanut TSSPs without cross-reaction to other food allergens, including nuts. Subsequently, analyzing processed foods by indirect ELISA, all foods labeled as containing peanuts in the product description were confirmed to be positive. The results indicate that the developed antibodies exhibit high specificity and sensitivity to peanuts and can be used as bio-receptors in immunoassays or biosensors to detect intentional or unintentional adulteration of peanuts in processed foods, particularly heat-processed foods.

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Section: Resultsmentioning

confidence: 99%

Detection of a Thermal Stable-Soluble Protein (TSSP) as a Marker of Peanut Adulteration Using a Highly Sensitive Indirect Enzyme-Linked Immunosorbent Assay based on Monoclonal Antibodies

Kim

Toushik

Lee

et al. 2023

J. Microbiol. Biotechnol.

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“…In ELISA, allergens can be detected by the colorimetric reaction after binding of the antigen with the specific enzyme-labeled antibody. Earlier studies reported the allergenic proteins quantification in peanut-based products through ELISA and HPLC technique (29, 33–35), however, no information on peanut seeds. The key issues in ELISA are the sample preparation and unacceptable variability in multiple measurements of the same sample.…”

Section: Introductionmentioning

confidence: 99%

An Improved Enzyme-Linked Immunosorbent Assay (ELISA) Based Protocol Using Seeds for Detection of Five Major Peanut Allergens Ara h 1, Ara h 2, Ara h 3, Ara h 6, and Ara h 8

et al. 2019

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Peanut allergy is an important health concern among many individuals. As there is no effective treatment to peanut allergy, continuous monitoring of peanut-based products, and their sources is essential. Precise detection of peanut allergens is key for identification and development of improved peanut varieties with minimum or no allergens in addition to estimating the levels in peanut-based products available in food chain. The antibody based ELISA protocol along with sample preparation was standardized for Ara h 1, Ara h 2, Ara h 3, Ara h 6, and Ara h 8 to estimate their quantities in peanut seeds. Three different dilutions were optimized to precisely quantify target allergen proteins in peanut seeds such as Ara h 1 (1/1,000, 1/2,000, and 1/4,000), Ara h 2 and Ara h 3 (1/5,000, 1/10,000, and 1/20,000), Ara h 6 (1/40,000, 1/80,000, and 1/1,60,000), and Ara h 8 (1/10, 1/20, and 1/40). These dilutions were finalized for each allergen based on the accuracy of detection by achieving <20% coefficient of variation in three technical replicates. This protocol captured wide variation of allergen proteins in selected peanut genotypes for Ara h 1 (77–46,106 μg/g), Ara h 2 (265–5,426 μg/g), Ara h 3 (382–12,676 μg/g), Ara h 6 (949–43,375 μg/g), and Ara h 8 (0.385–6 μg/g). The assay is sensitive and reliable in precise detection of five major peanut allergens in seeds. Deployment of such protocol allows screening of large scale germplasm and breeding lines while developing peanut varieties with minimum allergenicity to ensure food safety.

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Development of validated sandwich ELISA for detecting peanut allergen Ara h 3 in food

Lin,

Chang,

Hsieh

et al. 2024

Food Chemistry

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A Quantitative Method for Detecting Ara h 2 by Generation and Utilization of Monoclonal Antibodies

Cited by 4 publications

References 39 publications

Detection of a Thermal Stable-Soluble Protein (TSSP) as a Marker of Peanut Adulteration Using a Highly Sensitive Indirect Enzyme-Linked Immunosorbent Assay based on Monoclonal Antibodies

Detection of a Thermal Stable-Soluble Protein (TSSP) as a Marker of Peanut Adulteration Using a Highly Sensitive Indirect Enzyme-Linked Immunosorbent Assay based on Monoclonal Antibodies

An Improved Enzyme-Linked Immunosorbent Assay (ELISA) Based Protocol Using Seeds for Detection of Five Major Peanut Allergens Ara h 1, Ara h 2, Ara h 3, Ara h 6, and Ara h 8

Development of validated sandwich ELISA for detecting peanut allergen Ara h 3 in food

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