A modified fluorescence polarization immunoassay method incorporating fat emulsion (FE-FPIA) to determine cyclosporin A concentrations in rat skin.
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.
Kurosaki
Y
Y
Department of Clinical Pharmaceutical Science, Graduate School of Pharmaceutical Sciences, Okayama University, Japan.
Asada
T
T
Yasumoto
N
N
Nakamura
T
T
Masada
M
M
Kawasaki
H
H
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
D007166:Immunosuppressive Agents; D011933:Reagent Kits, Diagnostic; D016572:Cyclosporine