Use of a regenerable immobilized second antibody to determine azidothymidine in a flow system

Abstract

A regenerable immobilized second-antibody reactor was used to measure azidothymidine (AZT) by competitive enzyme immunoassay in a flow system. The immobilized antibody was regenerated 4 times with little loss of immunospecificity. A residual enzymatic activity of about 7% of the total response was obtained with horseradish peroxidase as label. AZT was measured below the nanomolar level by allowing competition to proceed for about 2 min. A limit of detection of 6.57 x 10(-11) +/- 1.56 x 10(-11) M AZT was obtained when the concentration of AZT-peroxidase conjugate was 0.125 ng/mL. This system was used to determined azidothymidine in Retrovir capsule with good results. In replicate measurements, RSDs of 5.51% and 2.44%, n = 6, were obtained at 7.48 x 10(-11) and 3.74 x 10(-9) M AZT, respectively.