title: Preparation and application of a monoclonal antibody against the isoflavone glycoside daidzin using a mannich reaction-derived hapten conjugate.
abstract: BACKGROUND
  Daidzin and its aglycone daidzein are major pharmacologically active compounds of soybean (Glycine max), kudzu (Pueraria lobata), and kwao kruea khao (P. mirifica). Pharmacological activities of daidzin are mediated by its more potent metabolites daidzein and equol; however, daidzin is the predominant compound found in these medicinal plants, and the efficacy and safety of equol depend on the amount of daidzin consumed.
  
  
  OBJECTIVE
  To develop a specific monoclonal antibody (MAb)-based indirect competitive enzyme-linked immunosorbent assay (icELISA) for standardisation of daidzin content in herbal medicines or nutraceuticals.
  
  
  METHODS
  The Mannich reaction was used for the synthesis of a highly immunogenic conjugate between daidzin and a cationised carrier protein. Splenocytes of hyperimmunised mice were fused with myeloma cells to generate a hybridoma secreting antibody against daidzin.
  
  
  RESULTS
  The icELISA showed high selectivity and acceptable sensitivity for daidzin determination (1.56-100 ng/mL) with high reproducibility (coefficients of variation were < 5%). The icELISA was a reliable analytical method for daidzin in Glycine max, Pueraria lobata and P. mirifica, for which daidzin recoveries from spiked samples were 98.99-104.94%. Daidzin content of these plant-derived products determined using the icELISA were in close agreement with those determined by a HPLC-UV method.
  
  
  CONCLUSIONS
  The icELISA is useful for specific daidzin determination because of its reliability, low cost, speed and high throughput.
authors.affiliation: Department of Pharmacognosy, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka, 812-8582, Japan.
authors.forename: Gorawit
authors.initials: G
authors.lastname: Yusakul
authors.affiliation: Department of Pharmacognosy, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka, 812-8582, Japan.
authors.forename: Seiichi
authors.initials: S
authors.lastname: Sakamoto
authors.affiliation: Faculty of Pharmaceutical Sciences, Ubon Rachathani University, Ubon Ratchathani, 34190, Thailand.
authors.forename: Thaweesak
authors.initials: T
authors.lastname: Juengwatanatrakul
authors.affiliation: Faculty of Pharmaceutical Sciences, Khon Kaen University, Khon Kaen, 40002, Thailand.
authors.forename: Waraporn
authors.initials: W
authors.lastname: Putalun
authors.affiliation: Department of Pharmacognosy, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka, 812-8582, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Tanaka
authors.affiliation: Department of Pharmacognosy, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka, 812-8582, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Morimoto
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005504:Food Analysis / Q000379:methods; D006241:Haptens / Q000737:chemistry*; D007529:Isoflavones / Q000032:analysis* / Q000737:chemistry / Q000276:immunology*; D008297:Male; D008807:Mice, Inbred BALB C; D029908:Pueraria / Q000737:chemistry; D015203:Reproducibility of Results; D012710:Serum Albumin, Bovine / Q000737:chemistry; D013025:Glycine max / Q000737:chemistry; D013154:Spleen / Q000166:cytology / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D006241:Haptens; D007529:Isoflavones; D012710:Serum Albumin, Bovine; C013908:daidzin
keywords: ELISA; Glycine max; Pueraria lobata; Pueraria mirifica; daidzin, soybean; monoclonal antibody