The development of a sensitive and specific enzyme immunoassay for FK480, a novel cholecystokinin type-A receptor antagonist, in human plasma.
A sensitive and specific enzyme immunoassay for FK480, a novel cholecystokinin type-A (CCK-A) receptor antagonist, was developed to study the pharmacokinetics of the drug at low-dose administration using a specific monoclonal antibody. The high performance liquid chromatography (HPLC) method had been used for studying toxicokinetics, but its determination limit (2.5 ng ml-1) was too high for use in clinical studies. Subsequently we developed an enzyme immunoassay (EIA) using rabbit anti-FK480 serum (polyclonal antibody). It had higher sensitivity (0.1 ng ml-1) when 0.5 ml of plasma was used but its specificity was low because of the cross-reactivity of the metabolites of FK480. Therefore we produced several monoclonal antibodies for FK480 by cell fusion, and selected the antibody which was least cross-reactive for the isolated metabolites of FK480. Finally we developed a sensitive and specific EIA using this monoclonal antibody. The lower limit of quantification of this method was 0.2 ng ml-1 when 0.2 ml of human plasma was used. The coefficient of variation over the calibration range (0.2-10 ng ml-1) was less than 15%. We used this method for clinical studies, and it showed a good correlation to the HPLC method when plasma concentration was 2.5 ng ml-1 or more.
Nozaki
K
K
Department of Pharmaceutical, Fujisawa Pharmaceutical Co., Ltd., Osaka, Japan.
Suzuki
S
S
Maeda
F
F
Takagaki
S
S
Suzuki
A
A
Hata
T
T
D000818:Animals; D000911:Antibodies, Monoclonal; D001570:Benzodiazepinones / Q000097:blood* / Q000276:immunology / Q000378:metabolism / Q000493:pharmacokinetics; D002851:Chromatography, High Pressure Liquid; D003429:Cross Reactions; D004285:Dogs; D005260:Female; D006727:Hormone Antagonists / Q000097:blood* / Q000276:immunology / Q000378:metabolism / Q000493:pharmacokinetics; D006801:Humans; D007124:Immunoenzyme Techniques; D007211:Indoles / Q000097:blood* / Q000276:immunology / Q000378:metabolism / Q000493:pharmacokinetics; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D011817:Rabbits; D051381:Rats; D017207:Rats, Sprague-Dawley; D011949:Receptors, Cholecystokinin / Q000037:antagonists & inhibitors*; D012680:Sensitivity and Specificity
D000911:Antibodies, Monoclonal; D001570:Benzodiazepinones; D006727:Hormone Antagonists; D007211:Indoles; D011949:Receptors, Cholecystokinin; C082966:FR 120480