Conventional imaging for three-dimensional (3D) ultra-architectural analysis of collagen fibers and fibroblasts is time-consuming and requires numerous ultrathin sections to search the target area. Currently, no method allows 3D ultra-architectural analysis of predetermined areas including spatial relationships between collagen fibers and fibroblasts in vitro. Herein, we developed a new method for in vitro analysis of the 3D ultrastructure of fibroblasts and collagen fibers using CLEM optimized for picrosirius red staining and FIB/SEM tomography. Collagen fibers were observed between, rather than on top of, stacked cells. This method offers the advantage of mesoscopic and ultrastructural analysis, thus minimizing bias and ensuring accurate observation.

中文翻译：

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使用针对天狼星红染色和 FIB/SEM 断层扫描优化的 CLEM 对胶原纤维和成纤维细胞进行相关成像

用于对胶原纤维和成纤维细胞进行三维 (3D) 超结构分析的常规成像非常耗时，并且需要大量超薄切片来搜索目标区域。目前，没有任何方法允许对预定区域进行 3D 超结构分析，包括体外胶原纤维和成纤维细胞之间的空间关系。在此，我们开发了一种使用针对天狼星红染色和 FIB/SEM 断层扫描优化的 CLEM 对成纤维细胞和胶原纤维的 3D 超微结构进行体外分析的新方法。在堆叠细胞之间而不是在堆叠细胞顶部观察到胶原纤维。这种方法提供了细观和超微结构分析的优势，从而最大限度地减少偏差并确保准确观察。