(SEMI-)QUANTITATIVE ANALYSIS OF REDUCED NICOTINAMIDE ADENINE-DINUCLEOTIDE FLUORESCENCE IMAGES OF BLOOD-PERFUSED RAT-HEART

In vivo analysis of the metabolic state of tissue by means of reduced nicotinamide adenine dinucleotide (NADH) fluorimetry is disturbed by t issue movements and by hemodynamic and oximetric effects. These factor s cause changes in the absorption of ultraviolet (UV) excitation light by the tissue. Many different methods have been used in the literatur e to compensate measured NADH fluorescence intensities for these effec ts. In this paper we show on theoretical grounds that the ratio of NAD H fluorescence intensity and UV diffuse reflectance intensity provides a (semi-)quantitative measure of tissue NADH concentrations. This res ult is corroborated by experiments with tissue phantoms in which absor ption and backscattering properties were varied. Furthermore, we have verified the validity of this compensation method in isolated Langendo rff-perfused rat heart preparations. In this preparation oximetric eff ects (of blood and tissue) are the major determinants of the metabolis m-dependent UV diffuse reflectance change. Hemodynamic effects accompa nying compensatory vasodilation are negligible. Movement artifacts wer e eliminated by simultaneously recording fluorescence and reflectance images, using a CCD camera with a biprism configuration. The results s how that the NADH fluorescence/UV reflectance ratio can be used to mon itor the mitochondrial redox state of the surface of intact blood-perf used myocardium.