Analytical Sciences

Sensitive Detection of Glycyrrhizinand Evaluation of the Affinity Constants by a Surface PlasmonResonance-based Immunosensor

Takatoshi SAKAI,* Katsura SHINAHARA,* Akira TORIMARU,* Hiroyuki TANAKA,** Yukihiro SHOYAMA,** and Kiyoshi MATSUMOTO* 

*Department of Bioscience and Biotechnology, Faculty of Agriculture, Kyushu University, Fukuoka 812-8581, Japan
**Department of Chemo-Pharmaceutical Sciences, Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka 812-8582, Japan

A sensitive and selective determination of glycyrrhizin (GC) based on surface plasmon resonance (SPR) was performed by using an anti-GC monoclonal antibody (GC-MAb) and GC-bovine serum albumin (GC-BSA) conjugate (antigen). GC-BSA was immobilized on an Au thin film of the SPR sensor chip by physical adsorption, and GC determinations were performed by an indirect competitive method. The addition of GC into the GC-MAb solution (5 µg/ml) was found to decrease the incident-angle shift sharply because of an inhibition effect of GC. The RSDs (n = 3) of each point were less than 4%. The lowest detection limit for GC by SPR was almost the same as that by ELISA, 60 - 75 ng/ml. An evaluation of the affinity constant between GC-MAb and GC using the data from ELISA and those from SPR measurements was performed. The values of the association constant (K_A) from three different analyses of ELISA data and from SPR measurements are discussed in detail. As a whole, the affinity constant (K_A) between GC-MAb and GC was on the order of 10⁷ M^-1.

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