Radiography has long been the standard approach to characterize carious lesions. Spatially resolved X-ray diffraction, specifically small-angle X-ray scattering (SAXS), has recently been applied to caries research. The aims of this combined SAXS and micro computed tomography (µCT) study were to locally characterize and compare the micro-and nanostructures of one natural carious lesion and of one artificially induced enamel lesion; and demonstrate the feasibility of an automated approach to

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... ned SAXS and µCT data in segmenting affected and unaffected enamel. Enamel, demineralized by natural or artificial caries, exhibits a significantly reduced X-ray attenuation compared to sound enamel and gives rise to a drastically increased small-angle scattering signal associated with the presence of nanometer-size pores. In addition, X-ray scattering allows the assessment of the overall orientation and the degree of anisotropy of the nanostructures present. Subsequent to the characterization with µCT, specimens were analyzed using synchrotron radiation-based SAXS in transmission raster mode. The bivariate histogram plot of the projected data combined the local scattering signal intensity with the related X-ray attenuation from µCT measurements. These histograms permitted the segmentation of anatomical features, including the lesions, with micrometer precision. The natural and artificial lesions showed comparable features, but they also exhibited size and shape differences. The clear identification of the affected regions and the characterization of their nanostructure allow the artificially induced lesions to be verified against selected natural carious lesions, offering the potential to optimize artificial demineralization protocols. Analysis of joint SAXS and µCT histograms objectively segmented sound and affected enamel.