An Effective, Versatile, and Inexpensive Device for Oxygen Uptake Measurement

Abstract

In the last ten years, the use of fluorescent probes developed to measure oxygen has resulted in several marketed devices, some unreasonably expensive and with little flexibility. We have explored the use of the effective, versatile, and inexpensive Redflash technology to determine oxygen uptake by a number of different biological samples using various layouts. This technology relies on the use of an optic fiber equipped at its tip with a membrane coated with a fluorescent dye (). This oxygen-sensitive dye uses red light excitation and lifetime detection in the near infrared. So far, the use of this technology has mostly been used to determine oxygen concentration in open spaces for environmental studies, especially in aquatic media. The oxygen uptake determined by the device can be easily assessed in small volumes of respiration medium and combined with the measurement of additional parameters, such as lactate excretion by intact cells or the membrane potential of purified mitochondria. We conclude that the performance of by this technology should make it a first choice in the context of both fundamental studies and investigations for respiratory chain deficiencies in human samples.

Mitochondria are physiologically maintained at close to 50 °C

“…Its fluorescence at 562 nm (essentially unchanged by the pH of the solution buffer in the range 7.2–9.5) ( S2 Fig ), decreased in a reversible and nearly linear fashion as temperature was increased: a temperature rise from 34 to 60 °C decreased fluorescence by about 50%, whilst 82% of the response to a 3 °C shift at 38 °C was preserved at 50 °C ( Fig 1Ba and 1Bb ). Using a thermostated, magnetically stirred, closable 750-μl quartz-cuvette fitted with an oxygen-sensitive optode device [ 2 ], we simultaneously studied oxygen consumption (or tension) and changes in MTY fluorescence ( Fig 1C ). Adherent cells were loaded for a minimum of 10 min with 100 nM MTY, harvested, and washed, then kept as a concentrated pellet at 38 °C for 10 min, reaching anaerobiosis in <1 min.…”

Mitochondria are physiologically maintained at close to 50 °C

“…1B, a, b). Using a thermostated, magnetically stirred, closable 750 µl quartz-cuvette fitted with an oxygen sensitive optode device [2] we simultaneously studied oxygen consumption (or tension) and changes in MTY fluorescence (Fig. 1C).…”

A PSII photosynthetic control is activated in anoxic cultures of green algae following illumination

“…In order to pin-point the cause of the apparent decrease in linear electron flow, we evaluated the effect of anoxia on the activity of isolated PSII. To that aim, we purified PSII complexes from C. reinhardtii wild-type strain CC124 cells, and examined O 2 evolution rates, using a Pyroscience FireSting O 2 probe 32 (Fig. 3a ).…”