Chemiluminescent measurement of peroxidase activity and its application using a lucigenin CT-complex

Luminescence. 2000 Jan-Feb;15(1):1-7. doi: 10.1002/(SICI)1522-7243(200001/02)15:1<1::AID-BIO561>3.0.CO;2-7.

Abstract

Luminol (3-aminophthaloylhydrazine) is well known as a chemiluminescent reagent that can be used to measure peroxidase (POD) activity. We have developed a more sensitive assay for POD using a new chemiluminescent reagent based on 10,10'-dimethyl-9, 9'-biacridinium nitrate (lucigenin). This new reagent was obtained from lucigenin by light irradiation in an organic polar solvent such as N,N-dimethylacetamide (DMAc). The detection limit for POD activity using this reagent is 10(-19) mol/assay. In an application for enzyme immunoassay based on a POD label, we were able to detect human alpha-fetoprotein (AFP) at 1pg/mL. The characterization of this chemiluminescent reagent was obtained by optical measurements, and the formation of lucigenin charge transfer complex (CT-complex) was confirmed. We also investigated the stability of lucigenin CT-complex as a chemiluminescent reagent for the measurement of POD activity, and confirmed that the reagent was stable for more than a year.

MeSH terms

  • Acridines*
  • Antibodies, Monoclonal
  • Electron Spin Resonance Spectroscopy
  • Humans
  • Immunoenzyme Techniques
  • Luminescent Measurements
  • Luminol
  • Peroxidases / analysis*
  • Sensitivity and Specificity
  • alpha-Fetoproteins / analysis*

Substances

  • Acridines
  • Antibodies, Monoclonal
  • alpha-Fetoproteins
  • 10,10'-dimethyl-9,9'-biacridinium
  • Luminol
  • Peroxidases