Goblet cells (GCs) in the conjunctiva are specialized epithelial cells producing mucins on the ocular surface. GCs play important roles in maintaining homeostasis of the ocular surface, and GC dysfunction is associated with various complications including dry eye diseases. Current GC examination methods, which are conjunctival impression cytology and confocal reflection microscopy, have limitations for routine examination. Fluorescence microscopy using moxifloxacin was recently introduced as a non-invasive and high-contrast imaging method, but further development is needed to be used for GC examination. Here we developed a non-invasive high-speed high-contrast GC imaging method, called moxifloxacin based axially swept wide-field fluorescence microscopy (MBAS-WFFM). This method acquired multiple fluorescence images with the axial sweeping of the focal plane to capture moxifloxacin labeled GCs on the tilted conjunctival surface in focus and generated all-in-focus images by combining the acquired images. The imaging field of view and imaging speed were increased to 1.6 mm × 1.6 mm and 30 fps. An image processing method was developed for the analysis of GC density. MBAS-WFFM was applied to alkali burn mouse models and detected GC damage and recovery via longitudinal imaging. MBAS-WFFM could assess the status of GCs rapidly and non-invasively. We anticipate MBAS-WFFM to be a starting point for non-invasive GC examination and the diagnosis of GC associated diseases. For example, MBAS-WFFM could be used to classify dry eye diseases into detail categories for effective treatment.

中文翻译：

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基于莫西沙星的轴向扫频广域荧光显微镜用于结膜杯状细胞的高速成像

结膜中的杯状细胞（GC）是专门的上皮细胞，在眼表产生黏蛋白。气相色谱在维持眼表稳态方面起着重要作用，气相色谱功能障碍与包括干眼病在内的各种并发症有关。当前的GC检查方法，即结膜印象细胞学检查和共聚焦反射显微镜检查，在常规检查中存在局限性。最近引入了使用莫西沙星的荧光显微镜作为一种非侵入性的高对比度成像方法，但需要进一步的发展以用于GC检查。在这里，我们开发了一种非侵入性的高速高对比度GC成像方法，称为基于莫西沙星的轴向扫频宽场荧光显微镜（MBAS-WFFM）。该方法通过焦平面的轴向扫掠获取多个荧光图像，以在焦点上倾斜的结膜表面上捕获莫西沙星标记的GC，并通过组合获取的图像生成全焦点图像。成像视野和成像速度增加到1.6 mm×1.6 mm和30 fps。开发了一种图像处理方法来分析GC密度。MBAS-WFFM用于碱烧小鼠模型，并通过纵向成像检测GC损伤和恢复。MBAS-WFFM可以快速，无创地评估GC的状态。我们预计MBAS-WFFM将成为无创GC检查和GC相关疾病诊断的起点。例如，MBAS-WFFM可用于将干眼病分为有效治疗的详细类别。