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A microplate assay for measuring cell death in C2C12 cells
Tanes I Lima1,2, Leonardo R Silveira1,3
1a Obesity and Comorbidities Research Center (OCRC), Institute of Biology, University of Campinas (UNICAMP), Campinas, São Paulo, Brazil.
Summary
This study presents a rapid microplate assay for measuring propidium iodide (PI) to assess cell viability in C2C12 cells. The assay is sensitive, reproducible, and effective for evaluating palmitate-induced cell death.
Area of Science:
- Cell Biology
- Biochemistry
- Toxicology
Background:
- Propidium iodide (PI) is a fluorescent intercalating agent that binds to DNA.
- PI is commonly used to identify dead cells in cell viability assays.
- Assessing cell viability is crucial in toxicological and pharmacological studies.
Purpose of the Study:
- To develop a rapid and straightforward microplate assay for quantifying propidium iodide (PI) in C2C12 cells.
- To establish a reproducible method for assessing cell viability and palmitate-induced cell death.
- To validate the sensitivity of the PI assay in detecting subtle changes in cell health.
Main Methods:
- C2C12 cells were treated with varying concentrations of palmitate for 24 hours.
- Propidium iodide (PI) fluorescence was measured using a microplate reader.
- Mitochondrial function and reactive oxygen species (ROS) production were evaluated.
- Catalase overexpression was used to assess assay sensitivity.
Main Results:
- PI fluorescence increased in a concentration-dependent manner with increasing palmitate levels.
- Palmitate exposure led to decreased mitochondrial function and increased ROS production.
- The microplate PI assay detected minor changes in cell viability following catalase overexpression.
Conclusions:
- A rapid, reproducible, and sensitive microplate assay for PI measurement in C2C12 cells was successfully developed.
- The assay is effective for quantifying cell death induced by cytotoxic agents like palmitate.
- This method provides an accurate tool for in-vitro cell viability studies.
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