The Seahorse XFe96 Extracellular Flux Analyzer (Agilent, Santa Clara, CA) was used to analyze cellular bioenergetics. Cells were plated in XF 96-well plates (Agilent) at a density of 25,000 cells/well and adhered to the plate at 37 °C, 5% CO2, for 4–8 hours before treatment with OT-82 or DMSO for 20 hours. The Glycolytic Rate Assay was performed according to manufacturer’s instructions and the Mito Stress Test was performed as previously described.(14 (link),43 (link)) Each assay was performed at least three times with at least three biological replicates.
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Cellular Bioenergetics Analysis with Seahorse XFe96
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Corresponding Organization : National Cancer Institute
Other organizations : National Center for Advancing Translational Sciences, Frederick National Laboratory for Cancer Research
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Variable analysis
independent variables
- OT-82
- Cellular bioenergetics
- Glycolytic rate
- Mitochondrial function
- Cell density (25,000 cells/well)
- Incubation time (4-8 hours for cell adherence, 20 hours for treatment)
- Incubation conditions (37 °C, 5% CO2)
- Positive control: Not explicitly mentioned.
- Negative control: DMSO
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