Development of a Molecular Assay for Rapid Screening of Chemopreventive Compounds Targeting Nrf2

Zhaohui Wang, Vinay Gidwani, Zheng Sun, Donna D. Zhang, Pak Kin Wong

Research output: Contribution to journalArticlepeer-review

11 Scopus citations

Abstract

Emerging molecular studies have shown that the transcription factor NF-E2-related factor (Nrf2) plays an essential role in cancer chemoprevention. Here, we report the development of a molecular biosensor for rapid detection of antioxidant-responsive element (ARE)-bound Nrf2 protein. The development will provide a molecular assay for high-throughput screening of chemopreventive compounds. Specifically, a double-stranded DNA probe is designed based on the ARE sequence. One of the DNA strands is labeled with a fluorophore on the 5′ end and the complementary strand is labeled with a quencher on the 3′ end. A single-stranded DNA competitor is also designed. The existence of the Nrf2 stabilizes the fluorescent probes and delays the competitor from separating the fluorophore-quencher complex. Therefore, the concentration of the Nrf2 proteins can be measured quantitatively based on the fluorescence intensity. The molecular binding scheme was demonstrated using purified p50 and the detection of endogenous Nrf2 was demonstrated using whole-cell lysates treated with sulforaphane. The assay can easily be incorporated into an automated platform for high-throughput screening of chemopreventive compounds targeting Nrf2.

Original languageEnglish (US)
Pages (from-to)243-248
Number of pages6
JournalJournal of Laboratory Automation
Volume13
Issue number4
DOIs
StatePublished - Aug 2008

Keywords

  • Nrf2
  • biosensor
  • chemoprevention
  • chemotherapy
  • high-throughput screening

ASJC Scopus subject areas

  • Computer Science Applications
  • Medical Laboratory Technology

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