Development of an enzyme-linked immunosorbent assay and immunoaffinity chromatography for glycyrrhizic acid using an anti-glycyrrhizic acid monoclonal antibody.
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.
Department of Biopharmaceutics, School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing, China.
Yue
Y
Zhang
Department of Molecular Biology, Centre of Scientific Experiment, Beijing University of Chinese Medicine, Beijing, China.
Huihua
H
Qu
Department of Chemistry of TCM, School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing, China.
Wenhao
W
Zeng
Department of Basic TCM Clinical Science, School of Basic Medical Sciences, Beijing University of Chinese Medicine, Beijing, China.
Yan
Y
Zhao
Department of Chemistry of TCM, School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing, China.
Wenchao
W
Shan
Department of Basic TCM Clinical Science, School of Basic Medical Sciences, Beijing University of Chinese Medicine, Beijing, China.
Xueqian
X
Wang
Department of Basic TCM Clinical Science, School of Basic Medical Sciences, Beijing University of Chinese Medicine, Beijing, China.
Qingguo
Q
Wang
Department of Basic TCM Clinical Science, School of Basic Medical Sciences, Beijing University of Chinese Medicine, Beijing, China.
Yan
Y
Zhao
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D002846:Chromatography, Affinity / Q000379:methods*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D019695:Glycyrrhizic Acid / Q000032:analysis* / Q000276:immunology; D057230:Limit of Detection; D051379:Mice; D008807:Mice, Inbred BALB C
D000911:Antibodies, Monoclonal; D019695:Glycyrrhizic Acid
Glycyrrhizic acid; Immunoaffinity chromatography; Indirect competitive enzyme-linked immunosorbent assay; Monoclonal antibodies