Publication: Preparation of a highly specific single chain variable fragment antibody targeting miroestrol and its application in quality control of Pueraria candollei by enzyme-linked immunosorbent assay
Issued Date
2019-11-01
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10991565
09580344
09580344
DOI
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2-s2.0-85065016571
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Mahidol University
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SCOPUS
Bibliographic Citation
Phytochemical Analysis. Vol.30, No.6 (2019), 600-608
Suggested Citation
Benyakan Pongkitwitoon, Panitch Boonsnongcheep, Tharita Kitisripanya, Gorawit Yusakul, Seiichi Sakamoto, Hiroyuki Tanaka, Satoshi Morimoto, Waraporn Putalun Preparation of a highly specific single chain variable fragment antibody targeting miroestrol and its application in quality control of Pueraria candollei by enzyme-linked immunosorbent assay. Phytochemical Analysis. Vol.30, No.6 (2019), 600-608. doi:10.1002/pca.2832 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/49715
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Title
Preparation of a highly specific single chain variable fragment antibody targeting miroestrol and its application in quality control of Pueraria candollei by enzyme-linked immunosorbent assay
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Abstract
© 2019 John Wiley & Sons, Ltd. Introduction: Miroestrol is the potent phytoestrogen isolated from White Kwao Krua (Pueraria candollei var. mirifica (Airy Shaw & Suvat.) Niyomdham, a Thai traditional medicinal plant. Nowadays, various health supplementary products featuring White Kwao Krua are available worldwide. A sensitive and rapid analytical method for quantification of miroestrol is necessary for quality control of these products. Objectives: To prepare a single-chain variable fragment (scFv) antibody specific to miroestrol and develop a scFv-based enzyme-linked immunosorbent assay (ELISA) for quantitative analysis of miroestrol in plant materials and health supplementary products. Methods: A gene encoding anti-miroestrol scFv antibody was constructed and expressed in Escherichia coli SHuffle T7 strain. Anti-miroestrol scFv antibody was characterised and applied to ELISA. The developed scFv-based ELISA method was validated for its sensitivity, specificity, accuracy and precision. Results: Anti-miroestrol scFv antibody was highly specific to miroestrol. The scFv-based ELISA was applied to determine miroestrol in the range 0.06–7.81 μg/mL, with the limit of quantification of 0.06 μg/mL miroestrol. The accuracy of the assay was validated by its 95.08–103.99% recovery from the spiked miroestrol recovery experiment and in good correlation with the results from the monoclonal antibody-based ELISA. The relative standard deviation of the intra- and inter-assay were less than 6.0%. Conclusion: The developed scFv-based ELISA was sensitive, specific, accurate, and precise for determination of miroestrol and useful for quality control of P. candollei plant raw materials and supplementary products.