Supatto Willy, McMahon Amy, Fraser Scott E, Stathopoulos Angelike
Division of Biology and Beckman Institute, California Institute of Technology, Pasadena, California, USA.
Nat Protoc. 2009;4(10):1397-412. doi: 10.1038/nprot.2009.130. Epub 2009 Sep 10.
This protocol describes imaging and computational tools to collect and analyze live imaging data of embryonic cell migration. Our five-step protocol requires a few weeks to move through embryo preparation and four-dimensional (4D) live imaging using multi-photon microscopy, to 3D cell tracking using image processing, registration of tracking data and their quantitative analysis using computational tools. It uses commercially available equipment and requires expertise in microscopy and programming that is appropriate for a biology laboratory. Custom-made scripts are provided, as well as sample datasets to permit readers without experimental data to carry out the analysis. The protocol has offered new insights into the genetic control of cell migration during Drosophila gastrulation. With simple modifications, this systematic analysis could be applied to any developing system to define cell positions in accordance with the body plan, to decompose complex 3D movements and to quantify the collective nature of cell migration.
本方案描述了用于收集和分析胚胎细胞迁移实时成像数据的成像和计算工具。我们的五步方案需要几周时间，从胚胎准备和使用多光子显微镜进行四维（4D）实时成像，到使用图像处理进行三维（3D）细胞追踪、追踪数据的配准以及使用计算工具进行定量分析。它使用市售设备，需要适合生物学实验室的显微镜和编程专业知识。提供了定制脚本以及样本数据集，以使没有实验数据的读者能够进行分析。该方案为果蝇原肠胚形成过程中细胞迁移的遗传控制提供了新的见解。通过简单修改，这种系统分析可应用于任何发育系统，以根据身体蓝图确定细胞位置、分解复杂的三维运动并量化细胞迁移的集体性质。
