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The measurement of PO2 and VO2 in microscopic volumes of living organs presents technical and interpretational challenges. The method of phosphorescence quenching by oxygen opens the possibility of non-invasive measurement of [O2] and PO2 in organs and tissues of a normally functioning organism. Information on the processes of oxygen transport and consumption in a microscopic volume of living tissue may be obtained with the excitation of an oxygen-sensitive probe inside and outside of a microvessel. However, in order to obtain an assayable phosphorescent signal from the microscopic volume of tissue, it is necessary to increase the probe concentration and the intensity of the excitation light, which leads to significant consumption of dissolved oxygen by the method itself. It is necessary to isolate internal oxygen flows from the influence of ambient air, provide temperature stabilization of the tissue and protect it from excessive illumination. The localization of the signal in the tissue depends not only on the size of the excitation spot, but also on the distribution of the probe in the various tissue compartments. The existence of steep gradients of PO2 at the microscopic level leads to the formation of heterogeneous phosphorescence decay curves, requiring a special approach for analysis. In this chapter we briefly discuss the key problems that have to be considered in experimental situations to provide reliable information on tissue oxygenation and detailed features of oxygen transport from arterial blood to cells.

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