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Development of an enzyme‐linked immunosorbent assay and immunoaffinity chromatography for glycyrrhizic acid using an anti‐glycyrrhizic acid monoclonal antibody
Author(s) -
Zhang Yue,
Qu Huihua,
Zeng Wenhao,
Zhao Yan,
Shan Wenchao,
Wang Xueqian,
Wang Qingguo,
Zhao Yan
Publication year - 2015
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.201500242
Subject(s) - monoclonal antibody , chemistry , affinity chromatography , chromatography , splenocyte , conjugate , aminopterin , monoclonal , antibody , enzyme , bovine serum albumin , biochemistry , microbiology and biotechnology , in vitro , biology , immunology , methotrexate , mathematical analysis , mathematics
In this work, a new monoclonal antibody specific for glycyrrhizic acid was prepared and characterized. A hybridoma secreting an anti‐glycyrrhizic acid monoclonal antibody was produced by fusing splenocytes from a mouse immunized against a glycyrrhizic acid–bovine serum albumin conjugate with the hypoxanthine–aminopterin–thymidine‐sensitive mouse myeloma cell line (Sp2/0‐Ag14). Subsequently, an indirect, competitive enzyme‐linked immunosorbent assay for glycyrrhizic acid was developed using the monoclonal antibody. In this assay, we detected an effective measuring range of 78.12–2500 ng/mL. Both intra‐assay and inter‐assay repeatability and precision were achieved, with relative standard deviations lower than 10%. In addition, glycyrrhizic acid levels in both formulated Chinese medicines and biological samples were determined with high sensitivity and efficiency. We then successfully developed a reliable immunoaffinity chromatography to separate glycyrrhizic acid completely from its parent medicine. These methods will contribute to further research investigations to better understand the interactions of glycyrrhizic acid with other drugs in the complex system of traditional Chinese medicine.