29 December 1997 Bioluminescence microscopy: application to ATP measurements in single living cells
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Bioluminescence microscopy can be used to measure intracellular cofactors and ionic concentrations (Ca2+, K+, ATP, NADH), as an alternative to micro- spectrophotometry and micro-fluorimetry, due to the development of sensitive detectors (cooled photomultipliers tubes and CCD). The main limitation comes from the very small and brief intensity of the emitted light. Our instrumentation based on an inverted microscope, equipped with high aperture immersion lenses is presented. Light intensity measurements are carried out through a photomultiplier sorted for low dark current and cooled at -5 degree(s)C to reduce thermal noise. Our first aim is to quantify ATP on single living cells using the firefly luciferin-luciferase couple. Experimental and kinetic aspects are presented to emphasize the potentialities of the technique.
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Frederic Brau, Frederic Brau, Pierre Helle, Pierre Helle, Jean Claude Bernengo, Jean Claude Bernengo, "Bioluminescence microscopy: application to ATP measurements in single living cells", Proc. SPIE 3197, Optical Biopsies and Microscopic Techniques II, (29 December 1997); doi: 10.1117/12.297967; https://doi.org/10.1117/12.297967

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