<title>Multiphoton excitation microscopy of human skin in vivo: early development of an optical biopsy</title>

Abstract

This paper presents several milestones in the development of techniques to be used as an optical biopsy for dermatology. The instrumentation for multiphoton excitation microscopy as applied to in vivo human skin is described. We compare multi-photon excitation microscopy and reflected light confocal microscopy for the microscopic observation of human skin in vivo. Multi-photon excitation is induced by a 80 MHz pulse train of femtosecond laser pulses at 780 nm wavelength. This nonlinear microscopic technique is inherently suitable for tissue fluorescence imaging because of its deeper penetration depth and lower specimen photodamage. This technique has non-invasively obtained tissue structural information in human epidermis and dermis. Confocal light microscopy based on a white light source, or a laser source can provide video-rate image acquisition with high resolution and high contrast. The relative merits of these two techniques can be identified by comparing 3D resolved images obtained from the forearm skin of the same volunteer. Optical low coherence reflectometry is another technique to image human skin in vivo. The application of Fourier transform analysis to the surface structure of skin is described as a method to characterize the skin surface topography.