Truong Quang Binh-An, Lenne Pierre-François
Institute for Developmental Biology of Marseille (IBDM), CNRS & Aix-Marseille Université, Campus de Luminy, Marseille Cedex, France.
Methods Cell Biol. 2015;125:119-42. doi: 10.1016/bs.mcb.2014.10.015. Epub 2015 Jan 8.
Visualization and quantification of supramolecular assemblies in cells are essential to understand the design principles of cells and tissues. The advent of photoactivated localization microscopy (PALM) and related techniques has offered unprecedented information on protein supramolecular assemblies in 3-D with a spatial resolution of a few tens of nanometers. Yet application of PALM microscopy for in vivo studies remains challenging. This chapter describes how to implement PALM microscopy for quantitative analysis of intercellular adhesion in the Drosophila embryo. Our protocol describes the sample preparation, the imaging setup, and the acquisition procedure. We also discuss how to proceed with quantitative analysis of data. Initially designed and implemented for Drosophila embryo imaging of intercellular adhesion, this protocol can be readily adapted to other structures than adhesions and other organisms such as Zebrafish or Caenorhabditis elegans.
细胞中超分子组装体的可视化和定量分析对于理解细胞和组织的设计原理至关重要。光激活定位显微镜（PALM）及相关技术的出现，提供了关于蛋白质超分子组装体三维结构的前所未有的信息，其空间分辨率可达几十纳米。然而，将PALM显微镜应用于体内研究仍然具有挑战性。本章描述了如何将PALM显微镜用于果蝇胚胎细胞间黏附的定量分析。我们的方案介绍了样品制备、成像设置和采集程序。我们还讨论了如何进行数据的定量分析。该方案最初设计并应用于果蝇胚胎细胞间黏附的成像，它可以很容易地适用于除黏附结构之外的其他结构，以及斑马鱼或秀丽隐杆线虫等其他生物体。
