Multiplexing fluorogenic esterase-based viability assay with luciferase assays

Kenji Ohgane, Hiromasa Yoshioka, Yuichi Hashimoto

Research output: Contribution to journalArticlepeer-review

2 Citations (Scopus)

Abstract

Luciferase-based reporter assays are one of the most common cell-based screening formats for drug discovery, and simultaneous evaluation of the cytotoxic effect of test compounds is of great value in reducing false-positives. Here we share a multiplex assay protocol that allows sequential measurement of cell viability (cell number) and luciferase activity of the same sample in a multi-well-plate format. The viability assay employs a fluorogenic esterase substrate, CytoRed. • This protocol allows sequential measurement of endogenous esterase activity (as a surrogate for cell number) and then luciferase activity in a single sample. • The protocol eliminates the need for parallel viability assay or protein assay using separate aliquots of the lysate. • This protocol is especially useful for assays with cells stably expressing a luciferase construct, for which co-transfection of another reporter gene is not a viable option.

Original languageEnglish
Pages (from-to)2013-2020
Number of pages8
JournalMethodsX
Volume6
DOIs
Publication statusPublished - 2019

Keywords

  • CytoRed
  • CytoRed-luciferase multiplex assay
  • Esterase
  • Fluorogenic substrate
  • Luciferase
  • Multiplex assay
  • Viability

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