DEVELOPMENT OF AN ENZYME-LINKED-IMMUNOSORBENT-ASSAY FOR AZADIRACHTINS

An enzyme-linked immunosorbent assay for azadirachtin A, a bioactive t etranor-triterpenoid from the neem tree (Azadirachta indica A. Juss), was developed. The conjugates were synthesized by employing the hemisu ccinate method and an ester activation method, which were previously d eveloped. Four rabbits were immunized with either 7-hemisuccinylazadir achtin-keyhole limpet hemocyanin (KLH) or 3-hemisuccinylazadirachtin-K LH. All sera were tested against the coating antigens 7-hemisuccinylaz adirachtin-bovine serum albumin (BSA) and 3-hemisuccinylazadirachtin-B SA. From this screening procedure, an assay was developed using antise rum 7056. The assay showed an IC50 value of about 100 ppb, with a rang e of detection from 10 to 1000 ppb for azadirachtin A and related comp ounds. Thus, the method presented is a step forward in terms of inexpe nsive, fast, and sensitive (improvement by a factor of 100) analysis o f azadirachtin and should provide a method that can easily be used mor e widely.