Development of a confocal and two-photon endomicroscope – Preliminary results of qualitative evaluation

Abstract

Optical methods, particularly confocal microscopy (CFM) and two-photon microscopy (TPM), have the potential to support or even replace the traditional histological diagnosis of cell-altering diseases. The high axial and lateral resolution of these techniques enables 3D imaging of even intracellular details. Using optical fibers, these systems can be miniaturized to a size sufficiently small to access the surface cells of a hollow organ through the working channel of an endoscope. In contrast to CFM, the TPM technique employs pulsed laser sources in order to nonlinearly stimulated fluorescence confined to the focal spot. Especially in cases of high power density in the optical fiber, and in addition to the dispersion effects, a nonlinear interaction of the light pulses with the fiber material will distort the pulses and therefore reduce the detectable signal. This paper presents the development of a confocal and a two-photon endomicroscope. In the experimental set-up, both a fs- and a cw-laser were coupled into a fiber bundle in order to compare qualitative aspects of confocal and two-photon imaging. It was possible to resolve cellular structure with a high axial resolution using this system. However, further developments are needed to improve the efficiency of the fluorescence excitation.