Abstract
Spectroscopic and microscopic methods for probing mitochondrial malfunction were established using cultivated endothelial cells from the calf aorta and various inhibitors of the respiratory chain, which is located at the inner mitochondrial membrane. Time-gated fluorescence spectroscopy was used to measure autofluorescence of the coenzyme NADH as well as “energy transfer efficacy” from excited NADH molecules (energy donor) to the mitochondrial marker rhodamine-123 (energy acceptor). Autofluorescence usually exhibited a weak increase after specific inhibition of enzyme complexes of the respiratory chain. In contrast, a pronounced increase in energy transfer efficacy was observed after inhibition of the same enzyme complexes. The detection of donor (NADH) and acceptor (R123) fluorescence in different nanosecond time gates following the exciting laser pulses enhances selectivity and improves quantification of energy transfer measurements. Therefore, timegated energy transfer spectroscopy is suggested to be an appropriate tool for probing mitochondrial malfunction.
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Schneckenburger, H., Gschwend, M.H., Strauss, W.S.L. et al. Time-gated microscopic energy transfer measurements for probing mitochondrial metabolism. J Fluoresc 7, 3–10 (1997). https://doi.org/10.1007/BF02764571
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DOI: https://doi.org/10.1007/BF02764571