Simultaneous enzyme immunoassay of two thyroid hormones.

We describe an enzyme immunoassay in which the two thyroid hormones, triiodothyronine and thyroxin, are measured simultaneously in a single tube. The method involves labeling the two with separate enzymes (beta-galactosidase and alkaline phosphatase, respectively), whose catalyzed reactions can easily be distinguished from each other by absorption spectrophotometry, with o-nitrophenyl-beta-galactoside and phenolphthalein monophosphate as substrates. Performance of this dual assay method compares well with that of conventional single-hapten enzyme-labeled assays, and results compare well with those by two single-hapten radioimmunoassays. The dual assay has certain advantages over single-hapten methods: smaller sample volume, lower reagent cost, and shorter overall assay time. As presented here, the use of enzyme labels to measure two (or more) haptens simultaneously represents a significant advance in the use of immunoassay techniques.