Development of ELISA for detection of Rh1 and Rg2 and potential method of immunoaffinity chromatography for separation of epimers

作者：Qu, Huihua; Wang, Yan; Shan, Wenchao; Zhang, Yue; Feng, Huibin; Sai, Jiayang; Wang, Qingguo; Zhao, Yan^*

来源：Journal of Chromatography B-Analytical Technologies in the Biomedical and Life Sciences, 2015, 985: 197-205.

DOI：10.1016/j.jchromb.2015.01.037

• Ginsenoside-Rh1
• 20(R,S)-ginsenoside-Rg2
• Monoclonal antibody
• Indirect competitive enzyme-linked immunosorbent assay
• Immunoaffinity chromatography

In this work, hybridomas producing anti-ginsenoside-Rh1 monoclonal antibodies (MAbs) were generated. These MAbs were subsequently used to create indirect competitive enzyme-linked immunosorbent assays (icELISAs). A linear correlation was obtained for G-Rh1 concentrations in the range from 26 to 512 ng/mL. The regression equation was y=1.979-0.201 Log(2)(X) with a regression coefficient of 0.9898. Precision and accuracy of the icELISA method were evaluated by the variations between replicates from well to well (intra-assay) and plate to plate (inter-assay). The recovery rates ranged from 93.16% to 108.43%. Testing with the icELISA demonstrated that the MAbs were specific for 20(S)-Rh1 and 20(S)-Rg2 with no cross-reactivity against 20(R)-Rhl and 20(R)-Rg2. The immunoaffinity chromatography column (IAC) was constructed by covalently coupling monoclonal antibody (MAb) against G-Rh1 to CNB-activated Sepharose 4B. When 20(R)-type-Rg2 passed through the IAC column, it was adsorbed, but the amount adsorbed was lower than that when 20(S)-type-Rg2 ran through the column. The differences in adsorption between the 20(S) and 20(R) type ginsenosides bring a new approach or method to separate 20(S)-Rg2 and 20(R)-Rg2 by IAC. Our results indicate that the icELISA is a sensitive and efficient approach for the identification of epimers, and the application of IAC using MAbs against small molecules provides a totally new thought and potential method for resolving epimers.

• 出版日期2015-3-15
• 单位北京中医药大学