Light sheet luminescence imaging with Cherenkov excitation in thick scattering media.

Light scattering leads to a severe loss of axial and transverse resolution with depth into tissue, limiting accuracy and value of biomedical luminescence imaging techniques. High-resolution imaging beyond a few-millimeter depth is prohibited because diffusive transport dominates beyond a few scattering distances. In this study, light sheet imaging through scattering media is demonstrated using a radiotherapy linear accelerator to deliver well-defined thin scanned sheets of x-rays. These sheets produce Cherenkov light within the medium, which in turn excites luminescence of an optical probe across the sheet plane. This luminescence can then be imaged by an intensified camera positioned perpendicular to the sheet plane. The precise knowledge of the light sheet position within the medium allowed for efficient attenuation correction of the signal with depth as well as spatial deconvolution of the excitation light. Together these methods allowed for the first time, to the best of our knowledge, high-resolution imaging of tissue-equivalent phantoms up to 3 cm thick, yielding the precise position and shape of luminescent lesions located deep in tissue without the need for nonlinear image reconstruction.