Chemiluminescence imaging ELISA using an imprinted polymer as the recognition element instead of an antibody

An imaging assay analogous to competitive enzyme immunoassays has been deve loped using a molecularly imprinted polymer instead of an antibody. The ant igen 2,4-dichlorophenoxyacetic acid (2,4-D) was labeled with tobacco peroxi dase, and the chemiluminescence reaction of luminol was used for detection. Microtiter plates (96 or 384 wells) were coated with polymer microspheres imprinted with 2,4-D, which were fixed in place by using poly(vinyl alcohol ) as glue. In a competitive mode, the analyte-peroxidase conjugate was incu bated with the free analyte in the microtiter plate, after which the bound fraction of the conjugate was quantified. After addition of the chemilumine scent substrates, light emission was measured in a high-throughput imaging format with a CCD camera. Calibration curves corresponding to analyte conce ntrations ranging from 0.01 to 100 mug/mL were obtained.