A quantitative luminescence assay for nonradioactive nucleic acid probes

Vladimir V. Didenko, Peter J. Hornsby

Research output: Contribution to journalArticle

9 Scopus citations

Abstract

In histochemical work with digoxigenin- or biotin-labeled nucleic acid probes, reproducibility of in situ hybridization depends on accurate measurement of the amount of non-radioactive label being used. We describe a rapid and sensitive assay for nonradioactive label incorporated into nucleic acids employing a luminogenic substrate for alkaline phosphatase, CSPD (disodium 3-(4-methoxyspirol[1,2-dioxetane-3,2'-(5'- chloro)tricyclo[3,3.1.13,7]decan]-4-yl)phenyl phosphate). An alkaline phosphatase antibody conjugate was bound to digoxigenin-labeled nucleic acids spotted on nylon membranes. Light emission from the reaction of the bound alkaline phosphatase with CSPD was measured with a luminometer. This method allows an accurate determination of digoxigenin incorporated into nucleic acid probes in the range of 0.5-500 fmol of nonradioactive label.

Original languageEnglish (US)
Pages (from-to)657-660
Number of pages4
JournalJournal of Histochemistry and Cytochemistry
Volume44
Issue number6
DOIs
StatePublished - Jan 1 1996
Externally publishedYes

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Keywords

  • CSPD
  • Digoxigenin
  • Luminescence
  • Nucleic acid probes

ASJC Scopus subject areas

  • Anatomy
  • Histology

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