Slow imaging with quicksilver: high pressure mercury intrusion in dental tissues studied with BSE-SEM and XMT.

Abstract

Mercury intrusion under very high pressures is used to study the distributions of dimensions of pores in permeable solids. We noted that archival calcified tissue samples which had been tested to 750 bar were permanently stained, had therefore retained mercury, and decided to investigate whether this could be a valuable morphological space stain. 40 yBP porosimetry samples were embedded in poly-methyl-methacrylate. Blocks were cut and polished for imaging, uncoated, with 20kV backscattered electron scanning electron microscopy (BSE-SEM): they were also imaged using X-ray microtomography (XMT) to study the 3D distribution of the mercury. Samples in this study included human bone, deciduous molars and permanent incisors, premolars and molars, rat upper and lower incisors, African elephant tusk dentine and cementum, and freeze-dried bovine teeth with immature enamel. In all the calcified tissues studied, small, unmineralized space compartments were brought into great prominence: in bone, osteocyte lacunae and canaliculi: in cementum, non-mineralised cores of Sharpey fibres and cementocyte lacunae and canaliculi, but we discovered previously unrecognised mineralisation defect zones in innermost primary acellular distal root cementum in human teeth. Dentine tubules were extremely marked and often showed an astonishing number of fine side branches at a size just below the resolution limit of conventional optical microscopy: their continuity across the enamel dentine junction with enamel tubules and spindles was clearly shown. Interglobular dentine zones in human dentine and incremental layering in elephant dentine were heavily impregnated. We obtained detailed information concerning the tubule curvature (odontoblast movement) pattern responsible for the beautiful chequer board arrangement in elephant tusk [which has led to the endangerment of the species]. In enamel, prism boundary discontinuities (‘sheaths’) were prominent in the known hypomineralisation tracts - tufts and lamellae - but also in developmental hypomaturation regions and ‘white spot’ caries. In rat incisor, defect non-mineralised zones were seen in innermost enamel. Because of the extreme high density and signal levels from Hg, both XMT and BSE-SEM demonstrate its presence at below their normal spatial resolution limits giving rise to apparently higher resolution. Ethics statement: all samples studied antedated the UK Human Tissues Act 2004 and CITES regulations.