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2 protocols using n 2 hydroxyethylpiperazine n 2 ethanesulfonic acid hepes potassium salt

1

Mitochondrial Bioenergetics Profiling Protocol

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Mannitol, sucrose, bovine serum albumin (BSA), N-2-hydroxyethylpiperazine-N’-2-ethanesulfonic acid (HEPES) potassium salt, potassium phosphate monobasic anhydrous (KH2PO4), magnesium chloride (MgCl2), pyruvate, malate, adenosine -5′-diphosphate (ADP), oligomycin A, carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP), rotenone, succinate, dimethyl sulfoxide (DMSO), N-acetylcysteine (NAC), cyclosporine A (CsA), metaphosphoric acid, paraformaldehyde were purchased from Sigma-Aldrich (St Louis, MO, USA). BCA protein assay kit was purchased from pierce (Rockford,IL). N-acetylcysteine amide (NACA) was a gift from Sentient Lifesciences Inc, New York, NY 10021.
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2

Mitochondrial Bioenergetics Assay

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Mannitol, sucrose, bovine serum albumin (BSA), N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid (HEPES) potassium salt, potassium phosphate monobasic anhydrous (KH2PO4), magnesium chloride (MgCl2), ethylene-diamine-tetra-acetic acid (EDTA), ethylene-glycol-tetra-acetic acid (EGTA), pyruvate, malate, glutamate, succinate, β-hydroxybutyrate, α-ketoglutarate, adenosine-5′-diphosphate (ADP), oligomycin A, carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP), rotenone, succinate, dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich (St. Louis, MO, USA). BCA protein assay kit was purchased from Pierce (Rockford, IL). Both NAD(t) and FAD(t) content assessments kits were purchased from Sigma-Aldrich (St. Louis, MO, USA). Mitochondrial substrates and inhibitors stock solutions were prepared and aliquots stored at −80°C.
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