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Related Experiment Videos

Multiparametric staining to identify apoptotic human cells

C Negri1, M Donzelli, R Bernardi

  • 1Istituto di Genetica Biochimica ed Evoluzionistica C.N.R., Pavia, Italy.

Experimental Cell Research
|July 10, 1997
PubMed
Summary
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A new tricolor assay detects apoptosis by analyzing chromatin condensation, DNA degradation, and poly(ADP-ribose) synthesis. This method reveals stimulated poly(ADP-ribose) production in apoptotic cells treated with antitumoral drugs.

Area of Science:

  • Cell Biology
  • Biochemistry
  • Molecular Biology

Background:

  • Apoptosis is a critical cellular process.
  • Detecting apoptosis accurately is vital for understanding disease and drug efficacy.
  • Existing methods may not capture all key apoptotic events simultaneously.

Purpose of the Study:

  • To develop and validate a novel "tricolor" assay for simultaneous analysis of key apoptosis indicators.
  • To investigate the role of poly(ADP-ribose) synthesis during etoposide-induced apoptosis in HeLa and HL60 cells.

Main Methods:

  • Development of a "tricolor" assay combining Hoechst staining (chromatin condensation), TUNEL assay (DNA degradation), and anti-poly(ADP-ribose) immunoreaction (poly(ADP-ribose) synthesis).
  • Simultaneous single-cell analysis of these three parameters.

Related Experiment Videos

  • Application to etoposide-treated HeLa and HL60 cell lines.

    • Main Results:

    • The tricolor assay successfully visualized multiple apoptotic features concurrently.
    • Demonstrated for the first time that endogenous poly(ADP-ribose) production is stimulated in cells undergoing apoptosis after antitumoral drug treatment.
    • Poly(ADP-ribose) synthesis is linked to DNA damage and chromatin condensation during apoptosis.

    Conclusions:

    • The developed tricolor assay is an effective tool for comprehensive apoptosis detection.
    • Monitoring poly(ADP-ribose) synthesis alongside morphological and biochemical markers aids in apoptotic cell identification.
    • This assay provides new insights into the biochemical events of drug-induced apoptosis.