Mitochondrial Membrane Potential Assay

After contact inhibition or treatment of HepG2 cells with vitamin C, cells were resuspended in 1 mL PBS at approximately 1 × 10⁶ cells/ml. TMRM was added to a final concentration of 20 nM and incubated for 30 mins at 37 °C, 5% CO₂. For CCCP control samples, CCCP was added to a final concentration of 50 nM to the cells, incubated for 5 mins at 37 °C, 5% CO₂ and then treated with 20 nM TMRM reagent for 30 mins. Cells were analyzed on a BD Symphony flow cytometer with 561 nm excitation.

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Occean J.R., Yang N., Sun Y., Dawkins M.S., Munk R., Belair C., Dar S., Anerillas C., Wang L., Shi C., Dunn C., Bernier M., Price N.L., Kim J.S., Cui C.Y., Fan J., Bhattacharyya M., De S., Maragkakis M., deCabo R., Sidoli S, & Sen P. (2023). Gene body DNA hydroxymethylation restricts the magnitude of transcriptional changes during aging. bioRxiv.

Publication Preprint 2023

BD Symphony flow cytometer

Cccp Cells Contact inhibition Hepg2 cells Vitamin c

Corresponding Organization : National Institute on Aging

Other organizations : Albert Einstein College of Medicine, Yale University

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Variable analysis

independent variables

Contact inhibition treatment
Vitamin C treatment

dependent variables

Mitochondrial membrane potential

control variables

Cell density (approximately 1 × 10^6 cells/ml)
TMRM concentration (20 nM)
Incubation time (30 mins)
Incubation temperature (37 °C)
Incubation atmosphere (5% CO2)

controls

Negative control, CCCP-treated cells

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