Burdyniuk Mariia, Wesolowska Natalia, Fleszar Michal, Karreman Matthia A, Machado Pedro, Borrego-Pinto Joana, Ruthensteiner Bernhard, Schwab Yannick, Lénárt Péter
Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
Electron Microscopy Core Facility, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
Methods Cell Biol. 2018;145:293-313. doi: 10.1016/bs.mcb.2018.03.031. Epub 2018 Apr 26.
The rapid and synchronous divisions of large and transparent oocytes, eggs, and embryos of marine species are exceptionally well suited for microscopic observation. Consequently, these cells have been models for cell division research since its beginnings and contributed some of its first and most fundamental discoveries. While large size and rapid transitions render these cells ideal specimens for light microscopy, the same features constitute a challenge for electron microscopy. Here, we describe example protocols from our work on starfish oocyte meiosis, where we overcome these challenges by using live imaging of fluorescently labeled structures in combination with correlated electron microscopy. In this work, we demonstrate how: (i) to capture a rapid, transient event in time and (ii) to localize a small structure within the large oocyte. These techniques are applicable with minor modifications to oocytes and embryos of other species and, possibly, to other cell types.
海洋物种的大型透明卵母细胞、卵子和胚胎能够快速同步分裂，这使其非常适合进行显微镜观察。因此，自细胞分裂研究伊始，这些细胞就一直是该领域的研究模型，并为一些最早且最基础的发现做出了贡献。尽管细胞体积大且变化迅速使其成为光学显微镜观察的理想标本，但这些特性对电子显微镜而言却是一项挑战。在此，我们描述了我们在海星卵母细胞减数分裂研究中的示例方案，通过结合荧光标记结构的实时成像与相关电子显微镜技术，我们克服了这些挑战。在这项工作中，我们展示了如何：（i）及时捕捉快速、短暂的事件，以及（ii）在大型卵母细胞内定位小结构。这些技术只需进行少量修改，即可应用于其他物种的卵母细胞和胚胎，甚至可能适用于其他细胞类型。
