KEYWORDS: Microscopy, Depth of field, Human subjects, Diseases and disorders, Tolerancing, Real time imaging, Imaging systems, Image deconvolution, Fluorescence microscopy, Eye models
Conjunctival goblet cells (CGCs) are specialized epithelial cells that secrete mucins to form the mucous layer of the protective tear film and by suppressing inflammation. Although CGCs are an important biomarker for diagnosing ocular surface diseases, rapid and noninvasive CGC examination methods have not been available. We have developed a new imaging system, high-speed extended depth-of-field wide-field microscopy with surface tracking, to enable non-contact large-area CGC imaging in human subjects. A novel long-range surface detection method was developed for rapid large-area mosaic imaging with lateral translation. Large-area CGC imaging and density quantification in human subjects was demonstrated. This new imaging system could be useful for noninvasive CGC examination in humans, which could be valuable for precision diagnosis and optimal treatment of ocular surface diseases.
Nondestructive three-dimensional (3D) pathology based on high-throughput 3D microscopy holds promise as a complement to traditional hematoxylin and eosin (H&E) stained slide-based two-dimensional (2D) pathology by providing rapid 3D pathological information. However, conventional techniques provided superficial information only due to shallow imaging depth. Herein, we developed open-top two-photon light sheet microscopy (OT-TPLSM) for intraoperative 3D pathology. A two-photon excitation light sheet, generated by 1D scanning of a Bessel beam illuminated the sample and planar imaging was conducted at 400 frames/s max. An imaging depth of 60-100 μm was achieved with long excitation wavelengths, and the image throughput was up to 1 cm2 per 7 min. Cells and extra-cellular matrix were visualized using extrinsic fluorescence and intrinsic second harmonic generation, respectively. OTTPLSM was tested in various human cancer specimens and cancer structures were detected via 3D visualization. OT-TPLSM may have the potential for rapid and precise 3D histopathological examination.
Mohs Micrographic Surgery (MMS) needs optical biopsy methods for tumor margin determination. Although confocal microscopy CM) has been used, CM has poor contrast to detect cancer cells with reflection contrast. We developed combined reflectance confocal (RC) & Moxifloxacin based two-photon (MB-TP) microscopy for high contrast. Here, combined microscopy was tested in various skin cancer tissues. Combined microscopy visualized both cell & extracellular matrix. Basal cell carcinoma nests were detected and distinguished from glands. Squamous cell carcinoma was detected with some features. This study showed combined microscopy has potentials for guiding MMS.
Surgical resection is the primary treatment for malignant brain tumors. This procedure has a dilemma—aggressive surgical resection tends to extend patient survival; however, it also increases the risk of neurological deficiencies. Current medical imaging methods are not sensitive and their interpretation largely depend on surgeon’s impression. High-speed cellular imaging method by using clinically applicable moxifloxacin was demonstrated for fast and sensitive tumor-detection. The detailed cytoarchitecture of brain tumor mouse model and malignant human brain tumors was revealed. This study showed the potential and feasibility of moxifloxacin-based confocal microscopy as a surgery-guiding method for tumor removal.
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