FULL ARTICLE Multiphoton imaging of the dentine-enamel junction Thierry Cloitre 1; 2 , Ivan V. Panayotov 3 , Herve ´ Tassery 3; 4 , Csilla Gergely 1; 2 , Bernard Levallois 3 , and Fre ´de ´ric J. G. Cuisinier 3 1 Universite ´ Montpellier 2, Laboratoire Charles Coulomb UMR 5221, F-34095, Montpellier, France 2 CNRS, Laboratoire Charles Coulomb UMR 5221, F-34095, Montpellier, France 3 EA 4203, UFR Odontologie, Universite ´ Montpellier 1, 34193 Montpellier Cedex 5, France 4 Aix Marseille Universite ´ ; Assistance Public Ho ˆpitaux de Marseille, 13354 Marseille, France Received 9 April 2012, revised 21 June 2012, accepted 22 June 2012 Published online 23 July 2012 Key words: dentine-enamel junction, multiphoton microscopy, second harmonic generation, two photon exited fluorescence, histology, human 1. Introduction Attempts to image the dentine-enamel junction (DEJ) follow the history of microscopy. Kohliker (1855) was the one who provided a first view of the DEJ histology, which appears as a simple line when imaged microscopically [1]. This apparently sharp in- terface, the ‘optical DEJ’, is thought to represent the original position of the basal membrane between ameloblasts and odontoblasts, where they contact in # 2012 by WILEY-VCH Verlag GmbH& Co. KGaA, Weinheim Journal of BIOPHOTONICS Early View publication on www.wileyonlinelibrary.com (issue and page numbers not yet assigned; citable using Digital Object Identifier – DOI) Multiphoton microscopy has been used to reveal struc- tural details of dentine and enamel at the dentin-enamel junction (DEJ) based on their 2-photon excited fluores- cence (2PEF) emission and second harmonic generation (SHG). In dentine tubule 2PEF intensity varies due to protein content variation. Intertubular dentin produces both SHG and 2PEF signals. Tubules are surrounded by a thin circular zone with a lower SHG signal than the bulk dentine and the presence of collagen fibers perpen- dicular to the tubule longitudinal axis is indicated by strong SHG responses. The DEJ appears as a low inten- sity line on the 2PEF images and this was never pre- viously reported. The SHG signal is completely absent for enamel and aprismatic enamel shows a homogeneous low 2PEF signal contrary to prismatic enamel. The SHG intensity of mantle dentine is increasing from the den- tine-enamel junction in the first 12 mm indicating a pro- gressive presence of fibrillar collagen and corresponding to the more external part of mantle dentine where ma- trix metallo-proteases accumulate. The high information content of multiphoton images confirms the huge poten- tial of this method to investigate tooth structures in phy- siological and pathological conditions. Dentine-Enamel Junction. Superposition of SHG and 2PEF images showing the relationship of enamel and dentine. * Corresponding author: e-mail: fre´ de´ ric.cuisinier@univ-montp1.fr, Phone: +33 411 759 225, Fax: +00 411 759 20 J. Biophotonics 1–9 (2012) / DOI 10.1002/jbio.201200065