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Detection of programmed cell death in cells exposed to genotoxic agents using a caspase activation assay

Michael E Gehring1, Patrick P Koty

  • 1Department of Pediatrics, Wake Forest University School of Medicine, Winston-Salem, NC, USA.

Methods in Molecular Biology (Clifton, N.J.)
|October 27, 2004
PubMed

Insights

Environmental toxins can cause DNA damage, leading cells to repair or undergo programmed cell death (PCD). This study introduces a method to detect activated caspases, key to PCD execution, in intact cells using fluorescent microscopy.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Toxicology

Background:

  • Environmental toxins induce DNA damage, triggering cellular responses.
  • Cells attempt DNA repair before replication to prevent mutations.
  • If repair fails, cells initiate programmed cell death (PCD) to eliminate damage.

Purpose of the Study:

  • To detail a method for detecting activated caspases in intact cells.
  • To utilize fluorochrome-conjugated caspase inhibitors for visualization.
  • To enable early detection of PCD execution via fluorescent microscopy.

Main Methods:

  • Employing fluorochrome-conjugated inhibitors specific to caspases.
  • Applying these inhibitors to intact cells.
  • Utilizing fluorescent microscopy for detection and analysis.

Main Results:

  • The protocol allows for the detection of activated caspases.
  • Activated caspases are identified as an early marker in the PCD pathway.
  • This method visualizes caspase activation in intact cells prior to other PCD indicators.

Conclusions:

  • This method provides a sensitive approach to study PCD.
  • Early detection of caspase activation is crucial for understanding cell fate decisions.
  • The protocol facilitates research into the role of caspases in response to DNA damage.

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