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Cell-Based High Content Analysis of Cell Proliferation and Apoptosis.

Bhaskar S Mandavilli1, Michelle Yan2, Scott Clarke2

  • 1Thermo Fisher Scientific, 29851 Willow Creek Road, Eugene, OR, 97402, USA. Bhaskar.mandavilli@Thermofisher.com.

Methods in Molecular Biology (Clifton, N.J.)
|October 31, 2017
PubMed
Summary

This study details methods for analyzing cell cycle progression using 5'-Ethynyl-2'-deoxyuridine (EdU) incorporation and detecting programmed cell death via caspase 3 activation. These techniques aid in evaluating compound toxicity and anti-cancer drug efficacy.

Keywords:
5-Ethynyl-2-deoxyuridineApoptosisCell cycleClick chemistry

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Area of Science:

  • Cell biology
  • Molecular biology
  • Pharmacology

Background:

  • High-content imaging enables multiplexed cell cycle analysis, including DNA content, synthesis, and proliferation markers.
  • 5 '-Ethynyl-2 '-deoxyuridine (EdU) incorporation offers a sensitive and convenient alternative to BrdU for detecting DNA synthesis.
  • Caspase 3 activation is a key indicator of programmed cell death, triggered by intrinsic or extrinsic pathways.

Purpose of the Study:

  • To describe methods for detecting S-phase cell cycle progression using EdU incorporation.
  • To outline procedures for detecting caspase 3 activation with the CellEvent caspase 3/7 detection reagent.
  • To provide tools for phenotypic analysis of compound toxicity and anti-cancer drug effects.

Main Methods:

  • Utilizing high-content imaging for multiplexed cell cycle analysis.
  • Employing 5 '-Ethynyl-2 '-deoxyuridine (EdU) incorporation for S-phase detection.
  • Using the CellEvent caspase 3/7 detection reagent to assess apoptosis.

Main Results:

  • Demonstrated the utility of EdU incorporation for sensitive DNA synthesis detection.
  • Showcased the effectiveness of CellEvent reagent for monitoring caspase 3 activation.
  • Established methods for simultaneous cell cycle and apoptosis analysis.

Conclusions:

  • EdU incorporation and caspase 3 detection are valuable tools for cell cycle and apoptosis studies.
  • These methods facilitate the phenotypic analysis of drug toxicity and anti-cancer drug development.
  • The described techniques enhance the understanding of cellular responses to chemical compounds.