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Although neuronal density analysis on human brain slices is available from stereological studies, data on the spatial distribution of neurons in 3D are still missing. Since the neuronal organization is very inhomogeneous in the cerebral cortex, it is critical to map all neurons in a given volume rather than relying on sparse sampling methods. To achieve this goal, we implement a new tissue transformation protocol to clear and label human brain tissues and we exploit the high-resolution optical sectioning of two-photon fluorescence microscopy to perform 3D mesoscopic reconstruction. We perform neuronal mapping of 100mm^3 human brain samples and evaluate the volume and density distribution of neurons from various areas of the cortex originating from different subjects (young, adult, and elderly, both healthy and pathological). The quantitative evaluation of the density in combination with the mean volume of the thousands of neurons identified within the specimens, allow us to determine the layer-specific organization of the cerebral architecture.