Cell invasion through basement membrane (BM) barriers is crucial in development, leukocyte trafficking and the spread of cancer. The mechanisms that direct invasion, despite their importance in normal and disease states, are poorly understood, largely because of the inability to visualize dynamic cell-BM interactions in vivo. This protocol describes multichannel time-lapse confocal imaging of anchor-cell invasion in live Caenorhabditis elegans. Methods presented include outline-slide preparation and worm growth synchronization (15 min), mounting (20 min), image acquisition (20-180 min), image processing (20 min) and quantitative analysis (variable timing). The acquired images enable direct measurement of invasive dynamics including formation of invadopodia and cell-membrane protrusions, and removal of BM. This protocol can be combined with genetic analysis, molecular-activity probes and optogenetic approaches to uncover the molecular mechanisms underlying cell invasion. These methods can also be readily adapted by any worm laboratory for real-time analysis of cell migration, BM turnover and cell-membrane dynamics.

中文翻译：

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秀丽隐杆线虫通过基底膜的锚定细胞入侵的活细胞共聚焦显微镜和定量4D图像分析。

通过基底膜（BM）屏障的细胞入侵在发育，白细胞运输和癌症扩散中至关重要。尽管它们在正常状态和疾病状态中很重要，但直接入侵的机制却鲜为人知，这主要是因为无法在体内可视化动态的细胞-BM相互作用。该协议描述了活的秀丽隐杆线虫中锚细胞入侵的多通道延时共聚焦成像。提出的方法包括轮廓幻灯片准备和蠕虫生长同步（15分钟），安装（20分钟），图像获取（20-180分钟），图像处理（20分钟）和定量分析（可变时间）。所获取的图像能够直接测量侵袭动力学，包括侵入足和细胞膜突起的形成以及去除BM。该协议可以与遗传分析，分子活性探针和光遗传学方法相结合，以揭示细胞入侵的分子机制。这些方法也可以由任何蠕虫实验室轻易修改，以实时分析细胞迁移，BM周转和细胞膜动力学。