Dache et al. (FASEB J 34: 3616-3630, 2020) recently reported the presence of respiratory-competent cell-free mitochondria in human blood (up to 3.7 × 106 per mL of blood), providing exciting perspectives on the potential role of these extracellular mitochondria. Although their evidence for the presence of cell-free mitochondria in human blood is compelling, their conclusion that these cell-free mitochondria are respiratory competent or functional has to be reevaluated. To this end, we evaluated the functionality of cell-free mitochondria in human blood using high-resolution respirometry and mitochondria extracted from platelets of the same blood samples as positive controls. Although cell-free mitochondria were present in human plasma (i.e., significant MitoTracker Green fluorescence and complex IV activity), there was no evidence suggesting that their mitochondrial electron transport system (ETS) was functional (i.e., respiration rate not significantly different from 0; no significant responses to ADP, uncoupler, or mitochondrial inhibitors oligomycin and antimycin A). Yet, in vitro complex IV activity was detectable and even slightly higher than levels found in mitochondria extracted from platelets, suggesting that cell-free mitochondria in human blood are likely to only retain a nonfunctional part of the ETS. Despite being unlikely to be fully functional in the narrow sense (i.e., capable of oxidative phosphorylation), circulating cell-free mitochondria may have significant physiological roles that remain to be elucidated.NEW & NOTEWORTHY The recently reported cell-free mitochondria in human blood have been thought to be respiratory competent, giving rise to speculation about their biological function(s). By characterizing their bioenergetics in vitro, we show that circulating cell-free mitochondria are unlikely to be functional in vivo since they display no potential for oxidative phosphorylation.