Confocal and multi-photon microscopy of dental hard tissues and biomaterials

Confocal microscopy is a technique that can be used both in the clinic and the high-resolution microscopy suite. This form of optical microscopy enables high-resolution images to be made of samples with minimum requirements for specimen preparation. Images may be made of either reflections from the sample surface or, if an immersion medium is used to optically couple the objective lens, then sub-surface images can be produced of reflective or fluorescent structures within semi transparent materials such as cells and dental hard tissues. These images are like optical sections, giving thin (> 0.35 microm) slices up to 200 microm below the surface of a mineralized tissue. The technique generates significant improvements in resolution, lying somewhere between that of conventional light microscopy and TEM/SEM. Instruments that work at video-rate allow high-speed events to be examined, such as in vivo clinical studies, cutting of dental tissues and fracture of adhesive interfaces. New dyes offer many exciting prospects for labeling changes in chemical composition in materials or biological tissues, while new imaging techniques such as multi-photon laser excitation of dyes give the potential of greater depth penetration and improved resolution. As with all new techniques the inexperienced should be aware of some of the artifacts inherent to the system. However, the widespread availability of conventional confocal microscopes should give ample opportunity for dental researchers to capitalize on this new technology.