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3D-Polarized Light Imaging (3D-PLI) enables high-resolution three-dimensional mapping of the nerve fiber architecture in unstained histological brain sections based on the intrinsic birefringence of myelinated nerve fibers. The interpretation of the measured birefringent signals comes with conjointly measured information about the local fiber birefringence strength and the fiber orientation. In this study, we present a novel approach to disentangle both parameters from each other based on a weighted least squares routine (ROFL) applied to oblique polarimetric 3D-PLI measurements. This approach was compared to a previously described analytical method on simulated and experimental data obtained from a post mortem human brain. Analysis of the simulations revealed in case of ROFL a distinctly increased level of confidence to determine steep and flat fiber orientations with respect to the brain sectioning plane. Based on analysis of histological sections of a human brain dataset, it was demonstrated that ROFL provides a coherent characterization of cortical, subcortical, and white matter regions in terms of fiber orientation and birefringence strength, within and across sections. Oblique measurements combined with ROFL analysis opens up new ways to determine physical brain tissue properties by means of 3D-PLI microscopy.

Derivation of Fiber Orientations From Oblique Views Through Human Brain Sections in 3D-Polarized Light Imaging