Published: Journal of Analytical Toxicology, Volume 21, Number 3, May/June 1997, pp. 190–196.
Detection of Flunixin
in Greyhound Urine by a Kinetic Enzyme-Linked Immunosorbent Assay
T.C. Brady,
T.J. Yang, W.G. Hyde, A.J. Kind, and D.W. Hill
A two-step kinetic enzyme-linked immunosorbent assay was developed to detect the presence of flunixin in the urine of greyhound dogs. The assay system was developed using polyclonal antiflunixin antisera, a rabbit albumin-flunixin conjugate adsorbed onto polystyrene microtiter strips, and flunixin reference standards for calibration. The assay parameters were optimized and the performance characteristics were determined. The quantitative intra- and inter-run precisons (%CV) of the analysis of replicate (n = 10) flunixin-spiked urine samples were 9.9–12.5% and 10.2–13.6%, respectively. The linear dynamic range was 1–100 ng/mL, and the quantitative accuracy, as determined by calculation of percent error of measured flunixin in flunixin-spiked drug-free greyhound urine, was –16% to +14% over this range. The I50 of the ELISA was 17.3 ng/mL. The limit of detection was 25 ng/mL in greyhound urine. The reactivity in the assay system relative to flunixin (100%) was 147% for flunixin glucuronide, 25% for clonixin, and 5% for niflumic acid. The ELISA was capable of detecting total flunixin for up to 72 h in dogs administered flunixin at 0.55 mg/kg orally and up to 96 h in a dog that was administered flunixin at 1.0 mg/kg orally.
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