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14 March 2016 Large field of view multiphoton microscopy of human skin
Mihaela Balu, Hideharu Mikami, Jue Hou, Eric O. Potma, Bruce J. Tromberg
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Proceedings Volume 9712, Multiphoton Microscopy in the Biomedical Sciences XVI; 97121F (2016) https://doi.org/10.1117/12.2216163
Event: SPIE BiOS, 2016, San Francisco, California, United States
Abstract
Clinical examination crucially relies on the ability to quickly examine large tissue areas and rapidly zoom in to regions of interest. Skin lesions often show irregularity in color and appearance in general, especially when they start to progress towards malignancy. Large field of view (FOV) and automatic translation of the imaging area are critical in the assessment of the entire lesion. Imaging of limited FOVs of the lesion can easily result in false negative diagnosis. We present a multiphoton microscope based on two-photon excited fluorescence and second-harmonic generation that images FOVs of about 0.8 mm2 (without stitching adjacent FOVs) at speeds of 10 frames/second (800 x 800 pixels) with lateral and axial resolutions of 0.5 μm and 2.5 μm, respectively. The main novelty of this instrument is the design of the scan head, which includes a fast galvanometric scanner, relay optics, a beam expander and a high NA objective lens. We optimized the system based on the Olympus 25x, 1.05NA water immersion lens, that features a long working distance of 1 mm. Proper tailoring of the beam expander, which consists of the scan and tube lens elements, enables scaling of the FOV. The design criteria include a flat wavefront of the beam, minimum field curvature, and suppressed spherical aberrations. All aberrations in focus are below the Marechal criterion of 0.07λ rms for diffraction-limited performance. We demonstrate the practical utility of this microscope by ex-vivo imaging of wide FOVs in normal human skin.
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Mihaela Balu, Hideharu Mikami, Jue Hou, Eric O. Potma, and Bruce J. Tromberg "Large field of view multiphoton microscopy of human skin", Proc. SPIE 9712, Multiphoton Microscopy in the Biomedical Sciences XVI, 97121F (14 March 2016); https://doi.org/10.1117/12.2216163
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KEYWORDS
Skin
Microscopes
Objectives
Imaging systems
Monochromatic aberrations
Relays
Scanners
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