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Updated: Feb 16, 2026
07:34
Modified Annexin V/Propidium Iodide Apoptosis Assay For Accurate Assessment of Cell Death
Published on: April 24, 2011
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Assessment of Apoptosis (Programmed Cell Death) by Flow Cytometry
Cold Spring Harbor Protocols
|January 4, 2018
Summary
This study presents a high-throughput flow cytometry assay using Annexin V and propidium iodide to measure early apoptosis. This method efficiently characterizes therapeutic antibody candidates by assessing their cell-killing capabilities.
Area of Science:
- Immunology
- Cell Biology
- Biotechnology
Background:
- Therapeutic antibodies can eliminate target cells through various mechanisms, including apoptosis.
- Characterizing these cell-killing effects is crucial for evaluating lead antibody candidates.
- High-throughput assays are needed for efficient, multidose candidate evaluation.
Purpose of the Study:
- To describe a simple, multiplexed flow cytometry assay for measuring early apoptosis.
- To enable rapid characterization of therapeutic antibody candidates.
- To assess antibody-mediated cell killing.
Main Methods:
- Utilized Annexin V and propidium iodide for a flow cytometry assay.
- Exploited the calcium-dependent binding of Annexin V to phosphatidyl serine (PS) on apoptotic cell surfaces.
- Incorporated EDTA to confirm Annexin V staining specificity.
Main Results:
- The assay effectively measures an early marker of apoptosis.
- Annexin V binding specificity was confirmed through calcium dependence and EDTA inhibition.
- Propidium iodide allowed differentiation of later apoptotic stages and cell death.
Conclusions:
- This multiplexed flow assay provides a rapid and efficient method for characterizing therapeutic antibody-mediated cell killing.
- The assay is suitable for high-throughput screening of antibody candidates.
- The described method aids in the development of effective therapeutic antibodies.
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