Determining Maximum Glycolytic Capacity Using Extracellular Flux Measurements
Fig 6
Effects of attenuating ATP demand using cycloheximide on assay of glycolytic capacity in HEK293 fibroblasts.
Raw traces of oxygen consumption rate (OCR) (a, c) and extracellular acidification rate (ECAR) (b, d) after sequential addition using ports A-D of 10 mM glucose, followed by vehicle or 10 μM cycloheximide (CHX), and then 2 μg/mL oligomycin (black), or 1 μM rotenone plus 1 μM myxothiazol (red), and then vehicle (black) or 200 μM monensin plus 1 μM FCCP (red). One representative experiment is shown. e: Respiratory (open column sections) and glycolytic (blue column sections) proton production rates (PPR) of the experiment exemplified in a-d calculated using Eq 1. Data are means ± SEM of n = 4 independent biological replicates. n.s.: not significant; *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.005. Statistical analysis was of glycolytic proton production rates only (blue column sections). w, well; A, B, C, D, addition ports.