Microelectrode and optical measurements of oxygen in biological systems in vitro and in vivo

The oxygenation status of tissue is important in several medical applications. It has long been appreciated that hypoxic tumors are resistant to radiation therapy, while in photodynamic therapy (PDT), the dependence on oxygen is even more pronounced. Our laboratory studies several problems in PDT that originate with this ³O ₂-dependence. Use of ³O ₂-sensitive electrodes with small (≤ 10 μm) tip diameters enables accurate measurement of metabolic and photochemical ³O ₂ consumption in photosensitized tumor spheroids in vitro. The temporal response of these devices (<1 s) is sufficient to capture rapid depletion of ³O ₂ during laser irradiation of individual spheroids. An important aspect of this research has been the development of appropriate mathematical models with which to interpret the microelectrode data and determine photophysical parameters. Recently, we have been establishing optical methods of measuring hemoglobin ³O ₂-saturation in order to monitor oxygenation in rodent tumors during PDT. We have adopted a steady-state diffuse reflectance technique, in which white light is injected into the tumor via an optical fiber, and the spatially-resolved diffuse reflectance is monitored using appropriately spaced detection fibers. The signals are spectrally dispersed onto a CCD camera by a grating spectrograph, providing simultaneous acquisition of spatially-resolved diffuse reflectance data for a 160 nm range of wavelengths. Absorption spectra reconstructed using a diffusion theory approximation are then fit to linear combinations of oxy- and deoxyhemoglobin spectra to obtain the ³O ₂-saturation.