Chondroitin sulfate is involved in the hypercalcification of the organic matrix of bovine peritubular dentin

• Chondroitin sulfate B is the major matrix component of bovine peritubular dentin.
• Laser capture of PTD and ITD sections allowed mapping of elemental content; Ca; S ;P.
• PTD and ITD differed quantitatively in elemental composition: SPTD > SITD; CaPTD > CaITD.
• The PTD-ITD boundary is marked by enhanced PTD topographical surface roughness.
• We demonstrate the EDS mapping of mineralized tissue with sub-nanometer resolution.

Apatitic mineral of dentin forms within the collagenous matrix (intertubular dentin, ITD) secreted from the odontoblastic processes (OP). Highly calcified mineral (peritubular dentin, PTD) is deposited at the interface between the ITD and each process membrane, creating a tubular system penetrating the dentin that extends from the dentino–enamel junction to the predentin–dentin junction. We focus on determining the composition of the PTD both with regard to its organic matrix and the inorganic phase. A laser capture technique has been adapted for the isolation of the mineralized PTD free from the ITD, and for the analysis of the PTD by SEM, TEM, and energy dispersive spectrometry (EDS), these data were subsequently compared with similar analyses of intact dentin slices containing ITD bounded-PTD annuli. Elemental line scans reveal clearly marked boundaries between ITD, PTD, and OP components, and illustrate the differences in composition, and topographical surface roughness. The organic matrix of the PTD was shown to be sulfur rich, and further antibody labeling showed the sulfated organic component to be chondroitin sulfate B. In this PTD organic matrix the S/Ca and Ca/P ratios were distinctly higher than in the ITD, indicating that polysaccharide bound S supplies the anionic counterion facilitating the formation of the apatitic PTD mineral.