4 trifluoromethoxy phenylhydrazone fccp

Mitochondrial function was analyzed using a Seahorse XF96 extracellular flux analyzer (Agilent Technologies, Carlsbad, CA, USA). After a 14-day culture, SOTRs were dissociated into a single-cell suspension by stirring for 30 min in protease solution, followed by cell seeding onto a cell culture microplate (Agilent Technologies) at a density of 20,000 cells per XF96 well. OCR assays were performed 3 days after seeding, and the culture medium was changed to the base medium (Seahorse XF assay media supplemented with 1 mM sodium pyruvate; Life Technologies, Carlsbad, CA, USA). Substrates and inhibitors were injected during measurements at a final concentration of 3.5 μM 4-(trifluoromethoxy) phenylhydrazone (FCCP; Seahorse Bioscience, Billerica, MA, USA), 1 μM oligomycin, 0.5 μM antimycin, and 0.5 μM rotenone for the MitoStress assay.

Mitochondrial function was analyzed using a Seahorse XF96 extracellular flux analyzer (Agilent Technologies, Carlsbad, CA, USA). After culturing for 14 days, the hiPSC-CM tissue was dissociated into a single-cell suspension using 0.25% trypsin/EDTA (Thermo Fisher) and then seeded in a microplate (XF96, Agilent Technologies) at a density of 2×10⁴ cells/well. After 3 days of culture, the culture medium was replaced with base medium (Seahorse XF assay media; Agilent Technologies, Carlsbad, CA, USA) supplemented with 1 mM sodium pyruvate. Substrates and inhibitors were added during measurements to attain a final concentration of 3.5 μM 4-(trifluoromethoxy) phenylhydrazone (FCCP; Seahorse Bioscience, Billerica, MA, USA), 1 μM oligomycin, 0.5 μM antimycin, and 0.5 μM rotenone for the MitoStress assay.