Direct measurement of oxygen consumption rates from attached and unattached cells in a reversibly sealed, diffusionally isolated sample chamber

Abstract

Oxygen consumption is a fundamental component of metabolic networks, mitochondrial function, and global carbon cycling. To date there is no method available that allows for replicate measurements on attached and unattached biological samples without compensation for extraneous oxygen leaking into the system. Here we present the Respiratory Detection System, which is compatible with virtually any biological sample. The RDS can be used to measure oxygen uptake in microliter-scale volumes with a reversibly sealed sample chamber, which contains a porphyrin-based oxygen sensor. With the RDS, one can maintain a diffusional seal for up to three hours, allowing for the direct measurement of respiratory function of samples with fast or slow metabolic rates. The ability to easily measure oxygen uptake in small volumes with small populations or dilute samples has implications in cell biology, environmental biology, and clinical diagnostics.

Regulation of mammalian nucleotide metabolism and biosynthesis

“…Supplementary Table S2) and the overall metabolic demand. The oxygen consumption rate of some cancer cells has been measured as 4 fmol/min/cell under normoxia (21% oxygen), to 2 fmol/min/cell under hypoxia (1% O 2 ) (117–120). Such cells can thus generate ca.…”

Probing the metabolic phenotype of breast cancer cells by multiple tracer stable isotope resolved metabolomics

“…Although the set of genetic lesions, the origin of the transformed cells, and their microenvironment differ among cancer cells (Fan, Lane et al 2009, Marin-Valencia, Yang et al 2012, Giussani M., Merlino G. et al 2015), their metabolism is generally enhanced to provide sufficient metabolic energy and anabolic substrates to drive proliferation, which requires macromolecular biosynthesis. Often, cancer cell metabolism is also adapted to deal with an increasingly hostile extracellular (Gatenby and Gillies 2008) and intracellular environments, including ROS production from accelerated respiration (Fantin, St-Pierre et al 2006, Telang 2007, Strovas, McQuaide et al 2010, Weinberg, Hamanaka et al 2010, Koppenol, Bounds et al 2011). …”

“…Figure 1). Moreover, glucose is metabolized in the mitochondrial Krebs cycle as mitochondria remain active in many cancer cells, and in fact may respire at a higher rate than the untransformed cell counterparts (Telang 2007, Strovas, McQuaide et al 2010). The respiratory activity is often, though not always perfectly, coupled to ATP synthesis (Gogvadze, Zhivotovsky et al 2009) to supplement glycolytically-derived energy.…”

Respiration Response Imaging for Real-Time Detection of Microbial Function at the Single-Cell Level

“…strain LW4 cultures was performed and compared with RSG staining. We used flow cytometry to estimate the number of respiring cells (stained with RSG) and employed a phosphorescence-based assay to monitor oxygen consumption (17). Samples with the source of electrons eliminated displayed a low rate of O 2 consumption (see Table S1 in the supplemental material), and fewer cells exhibited green fluorescence (see Fig.…”