Fig. 10: Bioenergetic profiles, NB expression, enzymatic activities, and oxygen consumption rates in HEK-293T cells.
From: Nanobodies as tools for studying human frataxin biology
A Aconitase and B succinate dehydrogenase activities (ACO and SDH, respectively) were quantified 48 h after transfection. OCR profiles are shown for cells transfect with C NB_4A7, D NB_6B1, E NB_16C10, and F NB_28F6. G The OCR parameters. After basal O2 consumption recording, oligomycin (an ATP synthase inhibitor) is added, and the coupling to the ATP synthesis is measured. Then, the addition of FCCP (a protonophore that uncouples oxygen consumption from ATP synthesis) allows us to measure the maximum respiratory capacity to use OXPHOS. This also enables the measurement of the spare respiratory capacity (SRC) as the difference between maximal and basal OCR. Finally, rotenone and antimycin A (Complex I and Complex III inhibitors, respectively) are added to block the electron transport chain. For panels A and B, three independent experiments were analyzed (n = 3), each with two technical replicates. For C–F, two independent experiments were analyzed (n = 2), including three technical replicates for each. Statistical analysis was performed using one-way ANOVA, followed by Dunnett´s test. Differences were considered significant when P ≤ 0.05. Error bars represent the standard errors.
