AbstractDentin hypersensitivity is an oral health concern affecting a large percentage of the world's adult population. Occlusion of the exposed dentinal tubules is among the treatment options available, and silver diammine fluoride (SDF) is an occluding agent used for interrupting or dampening the stimulus of the dental pulp nerves that produce pain. In addition to dentin permeability testing, the evaluation of desensitizing agents occluding dentinal tubules strongly relies on microscopic techniques, such as scanning electron microscopy (SEM). Limitations of SEM are that it provides only surface images that lack detailed information on the depth of penetration and amount of material present within the treated specimen, and it is prone to sample preparation artifacts. Here, we present high-resolution X-ray computed tomography (nano-CT) as a potential method for investigating dentin specimens with occluded tubules. We studied human dentin treated with SDF as an exemplary dentinal occlusion treatment option. We evaluated the silver deposits formed on the dentin surface region near the dentinal tubules and in the tubular regions using cross-section SEM, Energy Dispersive X-ray (EDX) analysis, and nano-CT. The resulting images obtained by SEM and nano-CT had comparable resolutions, and both techniques produced images of the tubules' occlusion. Nano-CT provided three-dimensional images adequate to quantitate tubule size and orientation in space. Moreover, it enabled clear visualization of dentinal tubules in any virtual plane and estimation of the amount and depth of occluding material. Thus, nano-CT has the potential to be a valuable technique for evaluating the occluding effects of virtually any material applied to dentinal tubules, supporting deciding between the best occluding treatment options.