Development of a latex agglutination inhibition reaction test for amphetamines in urine

A simple and rapid immunoassay of amphetamines based on latex agglutination inhibition reaction has been developed. N-(3-aminopropyl) amphetamine, a novel amphetamine derivative, and its BSA conjugate were synthesized and characterized. The hapten density in the conjugate was determined spectroscopically to be 62.52 mol/mol of BSA. Two other immunogens, amphetamine-BSA and amphetamino succinamide-BSA, were also synthesized and studied. It was found that N-(3-aminopropyl) amphetamine-BSA exhibits favorable features in terms of immunogenicity and immunochemical specificity when compared to the other two amphetamine immunogens. A latex agglutination inhibition reaction test (LAIRT) using DEAE-cellulose purified rabbit IgG against N-(3-aminopropyl) amphetamine-BSA was found to give a sensitivity of 0.6 microgram/ml and 4.0 microgram/ml of amphetamine and metamphetamine, respectively. Various commonly used drugs and narcotics at concentrations 0.8 mg/ml or less, did not interfere with the test. Interference by normal urine was observed but it could be eliminated by the inclusion of 0.78% normal rabbit serum. The sensitized latex was stable at 4 degrees C for at least 6 months. It was also stable to lyophilization and to at least four cycles of freezing and thawing. The total test time was 35 min. Comparison was made between the LAIRT and EMIT-d.a.u. on 56 urine samples collected from truck drivers. While the EMIT showed 47 positives and nine negatives, the LAIRT gave 38 positives and 18 negatives. The two tests showed no statistical significant difference (P < 0.05).