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使用基于光片的荧光显微镜对赤拟谷盗胚胎发育进行实时成像。

Live imaging of Tribolium castaneum embryonic development using light-sheet-based fluorescence microscopy.

机构信息

Physical Biology/Physikalische Biologie (IZN, FB 15), Buchmann Institute for Molecular Life Sciences (BMLS), Cluster of Excellence Frankfurt-Macromolecular Complexes (CEF-MC), Goethe University-Frankfurt am Main (Campus Riedberg), Frankfurt am Main, Germany.

出版信息

Nat Protoc. 2015 Oct;10(10):1486-507. doi: 10.1038/nprot.2015.093. Epub 2015 Sep 3.

Abstract

Tribolium castaneum has become an important insect model organism for evolutionary developmental biology, genetics and biotechnology. However, few protocols for live fluorescence imaging of Tribolium have been reported, and little image data is available. Here we provide a protocol for recording the development of Tribolium embryos with light-sheet-based fluorescence microscopy. The protocol can be completed in 4-7 d and provides procedural details for: embryo collection, microscope configuration, embryo preparation and mounting, noninvasive live imaging for up to 120 h along multiple directions, retrieval of the live embryo once imaging is completed, and image data processing, for which exemplary data is provided. Stringent quality control criteria for developmental biology studies are also discussed. Light-sheet-based fluorescence microscopy complements existing toolkits used to study Tribolium development, can be adapted to other insect species, and requires no advanced imaging or sample preparation skills.

摘要

赤拟谷盗已成为进化发育生物学、遗传学和生物技术的重要昆虫模式生物。然而,目前报道的赤拟谷盗活体荧光成像的方法很少,可用的图像数据也很少。在这里,我们提供了一种基于光片的荧光显微镜记录赤拟谷盗胚胎发育的方法。该方案可在 4-7 天内完成,并提供了以下方面的详细过程:胚胎收集、显微镜配置、胚胎准备和安装、长达 120 小时的多个方向的非侵入性活体成像、成像完成后对活体胚胎进行回收,以及图像数据处理,其中提供了示例数据。还讨论了严格的发育生物学研究质量控制标准。基于光片的荧光显微镜补充了现有的用于研究赤拟谷盗发育的工具包,可适应于其他昆虫物种,并且不需要先进的成像或样本制备技能。

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