Chemiluminescent enzyme immunoassay of alpha-fetoprotein based on an adamantyl dioxetane phenyl phosphate substrate.

We have evaluated a new chemiluminescent substrate for the alkaline phosphatase (EC 3.1.3.1) label used in a Hybritech Tandem-E immunoassay of alpha-fetoprotein (AFP). The new substrate, adamantyl 1,2-dioxetane phenyl phosphate (AMPPD), emits light at 477 nm when acted upon by the enzyme. Detection limits for AFP with this method were 33 ng/L (mean of 20 replicates of the zero standard + 2 SD) and 470 ng/L (twice background). Between-batch CVs ranged from 4.31% to 9.60% for AFP in the range 29.1-132.0 micrograms/L. Comparison of results for 49 specimens assayed with use of the chemiluminescent kit and a colorimetric version of the AFP assay gave statistical values as follows: slope = 0.88, intercept = 4.19, and r = 0.94.