A Near-Infrared HyperSpectral Reflective Confocal Microscopy (NIHS-RCM) is proposed in order to get high resolution images of deep biological tissues such as skin. The microscopy system uses a super-continuum laser for illumination, an acousto-optic tunable filter (AOTF) for rapid selection of near-infrared spectrum, a resonant galvanometer scanner for high speed imaging (15f/s) and near-infrared avalanche diode as detector. Porcine skin and other experiments show that the microscopy system could get deep tissue images (180 μm), and show the different ingredients of tissue with different wavelength of illumination. The system has the ability of selectively imaging of multiple ingredients at deep tissue which can be used in skin diseases diagnosis and other fields.
Laser scanning confocal microscope has been widely used in biology, medicine and material science owing to its advantages of high resolution and tomographic imaging. Based on a set of confirmatory experiments and system design, a novel confocal microscopic imaging system is developed. The system is composed of a conventional fluorescence microscope and a confocal scanning unit. In the scanning unit a laser beam coupling module provides four different wavelengths（405nm，488nm，561nm and 638nm）which can excite a variety of dyes. The system works in spot-to-spot scanning mode with a two-dimensional galvanometer. A 50 microns pinhole is used to guarantee that stray light is blocked and only the fluorescence signal from the focal point can be received . The three-channel spectral splitter is used to perform fluorescence imaging at three different working wavelengths simultaneously. The rat kidney tissue slice is imaged using the developed confocal microscopic imaging system. Nucleues labeled by DAPI and kidney spherule curved pipe labeled by Alexa Fluor 488 can be imaged clearly and respectively, realizing the distinction between the different components of mouse kidney tissue. The three-dimensional tomographic imaging of mouse kidney tissue is reconstructed by several two-dimensional images obtained in different depths. At last the resolution of the confocal microscopic imaging system is tested quantitatively. The experimental result shows that the system can achieve lateral resolution priority to 230nm.