Abstract
A competitive enzyme immunoassay for cinobufagin with alkaline phosphatase-labeled cinobufagin and polyethylene glycol as a precipitant was developed. The anti-cinobufagin antisera produced in rabbits by immunization with cinobufagin 3-hemisuccinate-bovine serum albumin conjugate efficiently recognized 14β, 15β-epoxy, 16β-acetoxy and 17β-six-membered unsaturated lactone structures, and therefore showed high affinity for cinobufagin and cinobufotalin. The range of cinobufagin measurable by the enzyme immunoassay was 10-500 pg/tube. Plasma cinobufagin levels of dogs and cats were determined after i.v. and p.o. administrations.
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