title: A modified fluorescence polarization immunoassay method incorporating fat emulsion (FE-FPIA) to determine cyclosporin A concentrations in rat skin.
abstract: We have developed a simple, sensitive and reliable assay procedure for cyclosporin A (CyA), a modified fluorescence polarization immunoassay method incorporating fat emulsion (FE-FPIA), to determine the CyA content in rat skin. The conventional fluorescence polarization immunoassay (FPIA) method for CyA using a commercially available FPIA kit, TDX cyclosporine monoclonal whole blood, was modified. A fat emulsion for intravenous infusion, Intralipos, was incorporated for dissolving the CyA extracted from the skin tissue, and a mixture of MeOH/purified water was used as the sample pretreatment medium instead of the precipitation reagent in the conventional FPIA kit intended for whole blood samples. These modifications enabled us to produce a reliable and the sensitive assay of CyA in skin tissue. The reproducibility (coefficient of variation), detection limit, and assay time for FE-FPIA were below 2%, 25 ng/ml, and about 24 min/24 samples, respectively, and were comparable with those for the whole blood samples determined by the conventional FPIA. Pre-purification of samples required by the HPLC assay is not needed in the FE-FPIA method. The usefulness of the FE-FPIA method in evaluating the topical pharmacokinetics of CyA in skin is discussed.
authors.affiliation: Department of Clinical Pharmaceutical Science, Graduate School of Pharmaceutical Sciences, Okayama University, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Kurosaki
authors.forename: T
authors.initials: T
authors.lastname: Asada
authors.forename: N
authors.initials: N
authors.lastname: Yasumoto
authors.forename: T
authors.initials: T
authors.lastname: Nakamura
authors.forename: M
authors.initials: M
authors.lastname: Masada
authors.forename: H
authors.initials: H
authors.lastname: Kawasaki
mesh_terms: D000818:Animals; D016572:Cyclosporine / Q000032:analysis* / Q000493:pharmacokinetics; D016231:Fluorescence Polarization Immunoassay / Q000379:methods*; D007166:Immunosuppressive Agents / Q000032:analysis* / Q000493:pharmacokinetics; D008297:Male; D011786:Quality Control; D051381:Rats; D017208:Rats, Wistar; D011933:Reagent Kits, Diagnostic; D015203:Reproducibility of Results; D012867:Skin / Q000737:chemistry* / Q000378:metabolism
chemical_list: D007166:Immunosuppressive Agents; D011933:Reagent Kits, Diagnostic; D016572:Cyclosporine