title: Development of Fluorescence-Linked Immunosorbent Assay for Paeoniflorin.
abstract: In this study, we developed a fluorescent immunoassay approach to detect paeoniflorin (PF) using a fluorescently labelled monoclonal antibody. The PF-specific antibody was purified by the caprylic acid-ammonium sulfate method and protein G Sepharose 4 Fast Flow column and then labelled with fluorescein isothiocyanate (FITC). The FITC-labelled monoclonal antibody was highly specific for PF, with less than 0.076 % cross-reactivity to seven structurally related compounds. The FITC-labelled monoclonal antibody was then used to develop an indirect competitive enzyme-linked immunosorbent assay (icELISA) and indirect competitive fluorescence-linked immunosorbent assay (icFLISA), respectively. FLISA is simple, rapid and sensitive, with a 500-fold lower limit of detection (LOD) compared with conventional ELISA. Finally, using a variety of standards, FLISA was validated. We observed a strong correlation between the results determined by either FLISA or conventional HPLC for the quantification of PF levels (R(2) = 0.9927). Collectively, this study shows that the icFLISA method can be successfully applied for the detection and quantification of PF in medicines and biological samples.
authors.affiliation: School of Basic Medical Sciences, Beijing University of Chinese Medicine, 11 Beisanhuandong Road, Chaoyang District, Beijing, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhao
authors.forename: Huihua
authors.initials: H
authors.lastname: Qu
authors.forename: Xueqian
authors.initials: X
authors.lastname: Wang
authors.forename: Yue
authors.initials: Y
authors.lastname: Zhang
authors.forename: Jinjun
authors.initials: J
authors.lastname: Cheng
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhao
authors.forename: Qingguo
authors.initials: Q
authors.lastname: Wang
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology*; D004365:Drugs, Chinese Herbal / Q000032:analysis*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005453:Fluorescence; D005455:Fluorescent Antibody Technique; D005960:Glucosides / Q000032:analysis* / Q000276:immunology; D007164:Immunosorbents / Q000737:chemistry*; D057230:Limit of Detection; D039821:Monoterpenes / Q000032:analysis* / Q000276:immunology; D006027:Glycosides
chemical_list: D000911:Antibodies, Monoclonal; D004365:Drugs, Chinese Herbal; D005960:Glucosides; D007164:Immunosorbents; D039821:Monoterpenes; C015423:peoniflorin; D006027:Glycosides