We present an imaging technique for the correction of geometrical effects in fluorescence measurement of optically thick, turbid media such as human tissue. Specifically, we use the cross-polarization method to reject specular reflection and enhance the diffusive backscattering of polarized fluorescence excitation light from the turbid media. We correct the nonuniformity of the image field caused by the excitation-and-collection geometry of a fluorescence imaging system by normalizing the fluorescence image to the cross-polarized reflection image. The ratio image provides a map of relative fluorescence yield, defined as the ratio of emerging fluorescence power to incident excitation, over the surface of an imaged homogeneous turbid medium when fluorescence excitation-and-collection geometries vary in a wide range. We investigate the mechanism of ratio imaging by using Monte Carlo modeling. Our findings show that this technique could have a potential use in the detection of early cancer, which usually starts from a superficial layer of tissue, based on the contrast in the tissue fluorescence of an early lesion and of the surrounding normal tissue.
A calibrated fluorescence imaging system utilizing a combination of fluorescence and cross-polarization imaging technology is described and applied to tissue examination in vivo. The results show that the inhomogeneity of fluorescence excitation and collection across the irregular surface of the examined tissue is calibrated to a great extent by taking the ratio of the raw fluorescence image to the cross-polarized reflection image. The effects of optical properties of tissue on the calibrated fluorescence signals are studies on simulated tissue phantoms systematically. Using the calibrated fluorescence imaging technology, we demonstrate that different tissue can be clearly separated endoscopically and in vivo based on the calibrated fluorescence signal.
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