Polyclonal-based ELISA for the identification of cyclohexanedione analogs that inhibit maize acetyl coenzyme-A...

Cyclohexanedione herbicides inhibit monocotyledonous acetyl coenzyme-A carboxylase (ACCase; E.C. 6.4.1.2.), which catalyzes the first committed step in fatty acid biosynthesis. Although the target site has been identified, little is known about the mechanisms involved in herbicide binding. An immunological study was undertaken to create a model to better characterize the herbicide-enzyme interaction. Cyclohexanedione-specific antiserum was raised in New Zealand white rabbits by immunizing them with a cyclohexanedione analog-bovine serum albumin conjugate. Two indirect enzyme-linked immunosorbent assays (ELISA) were developed using 2 different cyclohexanedione analogs conjugated to ovalbumin as coating conjugates. Nineteen cyclohexanedione analogs, 13 active ACCase inhibitors, and 6 inactive analogs were tested for their ability to compete with both coating conjugates for antiserum binding. All active ACCase inhibitors were observed to compete with both coating conjugates, whereas all inactive analogs failed to compete with at least one coating conjugate. On the basis of these results, the immunological model could be used to distinguish all active ACCase inhibitors from inactive analogs using the 2 ELISAs sequentially.