Tubule and microbranch variations in human dentine: A quantitative 3D study with serial block‐face scanning electron microscopy

OBJECTIVE: Microbranches exist within dentine, providing a network of fluid filled connections between tubules. This study used serial block-face scanning electron microscopy to generate 3D volumes of dentine from different tooth locations, allowing for investigations of the microbranch morphology, the microbranch network characteristics and regional variations in the tooth. METHODS: Three human teeth were sectioned to generate 21 dentine samples extracted from 7 identical regions of interest around the tooth. Serial block-face scanning electron microscopy was performed using a pixel size of 10 nm, a slice thickness of 50 nm, and a field of view of 20.48 µm × 20.48 µm. Tubules and microbranches were automatically segmented and the following quantified: tubule diameter and density; microbranch diameter, density, and connectivity; tissue porosity. RESULTS: The microbranches were successfully captured at sufficient resolution to create a 3D volume from sequential 2D images. The microbranches move tortuously through the intertubular dentine and travelled a variety of distances before connecting to other tubules. The microbranch density varied by location, with more found in areas of lower tubule density. Microbranches significantly increased tissue porosity (p < 0.001). Microbranch diameter did not vary regionally, with most approximately 140-210 nm, regardless of sample location. Tubule density and diameter varied radially-both largest closest to the pulp cavity, then decreasing to the dentinoenamel junction. CONCLUSIONS: This study provides new three-dimensional insight into the morphology and network of the dentinal microbranch network, demonstrating a variability in the density of the microbranches with location around the tooth and a significant increase in tissue porosity.