DEVELOPMENT OF A HOMOGENEOUS PHOSPHORESCENT IMMUNOASSAY FOR THE DETECTION OF POLYCHLORINATED DIBENZO-p-DIOXINS

Abstract

A homogeneous immunoassay for dioxins was developed using a phosphorescent label for detection. A dioxin derivative was conjugated to Pt-coproporphyrin. In the assay, when the antibodies against dioxin (DD3) were bound to the phosphorescent conjugate, the signal from Pt-coproporphyrin was quenched. The interaction between antibody and the conjugate was studied by time-resolved luminescence spectroscopy. As concentrations of dioxin standards increased from 39 pg/well to 2.5 ng/well the lifetime of the phosphorescence of the shortlived component increased from 25.6 microseconds to 58.9 microseconds. This increase in half-life was associated with a dose dependent quenching of the phosphorescence. The inhibition obtained is similar to that for a reported enzyme-based immunoassay, but the data were obtained in minutes instead of hours.

Keywords:

• Phosphorescence
• Pt-coproporphyrin
• Homogeneous immunoassay
• Polychlorinated dibenzo-p-dioxins
• Time-resolved luminescence spectroscopy

ACKNOWLEDGMENTS

The research described in this publication was made possible in part by Award No. RN 1–426 of the U.S. Civilian Research & Development Foundation for the Independent States of the Former Soviet Union (CRDF). U.S. investigators were funded in part by the National Institute of Environmental Health Sciences Superfund Basic Research Program ES04699, U.S. Environmental Protection Agency Center grant CR819658, and the National Institute of Environmental Health Sciences Center ES05707.