Nicotine Antibodies: Comparison of Ligand Specificities of Antibodies Produced against Two Nicotine Conjugates

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Abstract
Conjugates between bovine serum albumin and (R,S)-2-aminonicotine were produced, and these conjugates were employed in rabbits and goats for the production of nicotine antibodies. In the assay of nicotine, an 125I-tyrosine methyl ester derivative of (R,S)-6-aminonicotine was employed as radioligand. The antibody-bound derivative was separated from the free derivative by charcoal adsorption (0.5% charcoal, 0.1% dextran T-70, 0.1% bovine serum albumin, pH 7.3). Among the 25 nicotine derivatives and metabolites examined, (R,S)-6-aminonicotine gave the highest cross-reaction. Cross-reaction with cotinine, a major mammalian metabolite of nicotine, was < 0.1% for both the rabbit-derived and goat-derived antisera. Cross-reaction by other metabolites, such as (S)-nicotine-N''-oxide, (S)-nornicotine, and N-methylpyrrolidine was < 1%. The antibodies produced were thus highly specific to nicotine. The radioimmunosassay for nicotine showed a maximum sensitivity of 10 ng/ml in 50-.mu.l plasma samples for both antisera. After the smoking of a single cigarette (1.2 mg nicotine content in mainstream) the peak blood plasma level of nicotine in the subjects varied from 20-104 ng/ml, and high levels of nicotine were not necessarily found in heavy smokers.

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