Functional microscopic imaging of in vivo tissues aims at characterizing parameters at the level of the unitary cellular components under normal conditions, in the presence of blood flow, to understand and monitor phenomena that lead to maintaining homeostatic balance. Of principal interest are the setting of shear stress on the endothelium; formation of the plasma layer, where the balance between nitric oxide production and scavenging is established; and formation of the oxygen gradients that determine the distribution of oxygen from blood into the tissue. Optical techniques that enable the analysis of functional microvascular processes are the measurement of blood vessel dimensions by image shearing, the photometric analysis of the extent of the plasma layer, the dual-slit methodology for measuring blood flow velocity, and the direct measurement of oxygen concentration in blood and tissue. Each of these technologies includes the development of paired, related mathematical approaches that enable characterizing the transport properties of the blood tissue system. While the technology has been successful in analyzing the living tissue in experimental conditions, deployment to clinical settings remains an elusive goal, due to the difficulty of obtaining optical access to the depth of the tissue.