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Modified Annexin V/Propidium Iodide Apoptosis Assay For Accurate Assessment of Cell Death
Published on: April 24, 2011
Rationale for the real-time and dynamic cell death assays using propidium iodide
Hong Zhao1, Jadwiga Oczos, Pawel Janowski
1Brander Cancer Research Institute and Department of Pathology, New York Medical College, Valhalla, New York, USA.
Summary
Propidium iodide (PI) can be safely used in real-time cell viability assays. This study confirms PI has minimal impact on cell cycle, proliferation, and DNA damage, supporting its use in kinetic assays.
Area of Science:
- Cell Biology
- Biotechnology
- Molecular Toxicology
Background:
- Charged fluorochromes like propidium iodide (PI) are essential for cell viability assays.
- Real-time, dynamic assays require markers with minimal cellular impact.
Purpose of the Study:
- To provide a mechanistic rationale for using PI in kinetic cell viability assays.
- To investigate the effects of PI on live cells, including cell cycle, proliferation, and DNA damage.
Main Methods:
- Laser scanning microscopy and cytometry were used to analyze PI uptake and cellular effects.
- 5'-ethynyl-2-deoxyuridine (EdU) incorporation and histone gammaH2AX phosphorylation were measured.
- Confocal imaging assessed PI localization within various cell types.
Main Results:
- PI exposure (1.5-7.5 microM for 24-48h) minimally affected cell cycle progression and DNA replication.
- No significant increase in DNA damage markers (gammaH2AX) was observed.
- PI accumulated in cytoplasm and nucleoli, with no impact on cellular response to anticancer drugs.
Conclusions:
- PI demonstrates minimal toxicity and cellular disruption at concentrations used in kinetic viability assays.
- The findings support the reliable application of PI in real-time, dynamic cell viability assessments.
- PI's mechanistic profile validates its use as a cell permeability marker in kinetic studies.
