Abstract:
Apoptosis biomarkers were investigated in actinomycin D-treated SiHa cervical cancer
cells using a benchtop flow cytometer. Early biomarkers (Annexin V and mitochondrial
membrane potential) and late biomarkers (caspases 3 and 7, and DNA damage)
of apoptosis were measured in experimental and control cultures. Cultures were
incubated for 24 hours in a humidified incubator at 37 °C with 5% CO2. The cells were
then detached using trypsin and enumerated using a flow cytometric cell count assay.
Cells were further analyzed for apoptosis using an Annexin V assay, a mitochondrial
electrochemical transmembrane potential assay, a caspase 3/7 assay, and a DNA
damage assay. This article provides an overview of apoptosis and traditional flow
cytometry, and elaborates flow cytometric protocols for processing and analyzing SiHa
cells. The results describe positive, negative, and sub-optimal experimental data. Also
discussed are interpretation and caveats in performing flow cytometric analysis of
apoptosis using this analytical platform. Flow cytometric analysis provides an accurate
measurement of early and late biomarkers for apoptosis.
