Published: Journal of Analytical Toxicology, Volume 22, Number 6, October 1998, pp.418-424.

Amphetamines in Hair by Enzyme-Linked Immunosorbent Assay
Stacy A. Sweeney, Raymond C. Kelly, James A. Bourland, Ted Johnson, W. Craig Brown, Howard Lee, and Ernie Lewis

Human hair was collected from the occipital crown region of the head from several subjects; these hair samples were presumptively positive for amphetamines by a previously evaluated immunoassay. Hair was washed briefly with methanol to remove external contamination, then extracted with hot methanol for 2 h to recover the drugs. The extracts were evaporated to dryness, reconstituted in buffer, and analyzed using a new enzyme-linked immunosorbent assay (ELISA) technique adapted for the detection of amphetamines in hair. Gas chromatographyÐmass spectrometry was used as the reference technique. Cross-reactivity of several related compounds was evaluated by equating the inverse of the ligand concentration at 50% antibody binding to the affinity constant for each compound. The ratio of a compoundÕs affinity constant to that for d-methamphetamine was used to derive percent crossreactivity. These experiments yielded values of 30.8% for d-amphetamine, 7.4% for l-methamphetamine, 4.3% for phentermine, 2.9% for l-amphetamine, and <1% for ephedrine, methylenedioxyamphetamine, and methylenedioxymethamphetamine. Cross-reactivity of unrelated compounds was found to be non-existent. The optimum cutoff concentration was determined by receiver operating characteristic curve analysis to be 300 pg/mg and the observed limit of detection was 60 pg/mg. Intra-assay precision at 300 pg/mg was 3.3% (coefficient of variation, CV), and the interassay CV was 10.5%. The sensitivity and specificity of the method were 83% and 92%, respectively.

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