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固定和活原生小胶质细胞中观察微管和微管末端的方案。

Protocol for observing microtubules and microtubule ends in both fixed and live primary microglia cells.

机构信息

Center for Life Nano- & Neuro-Science, Istituto Italiano di Tecnologia, 00161 Rome, Italy.

Center for Life Nano- & Neuro-Science, Istituto Italiano di Tecnologia, 00161 Rome, Italy; Department of Physiology and Pharmacology, Sapienza University, 00185 Rome, Italy.

出版信息

STAR Protoc. 2023 Sep 15;4(3):102499. doi: 10.1016/j.xpro.2023.102499. Epub 2023 Aug 12.

DOI:10.1016/j.xpro.2023.102499
PMID:37573502
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC10448205/
Abstract

Microtubule dynamics and orientation have crucial roles in many vital cellular processes. However, functional live imaging of microtubules and/or microtubule ends in primary microglia can be challenging. Here, we present a protocol for observing microtubules and microtubule ends in both fixed and live primary microglia cells. We describe steps for microglia culture and in vitro stimulation, SiR-tubulin labeling, lentivirus preparation, live imaging, immunostaining, and image acquisition. We also provide procedures for SiR-tubulin, EB3-EGFP, and EB1 analyses. For complete details on the use and execution of this protocol, please refer to Rosito et al. (2023)..

摘要

微管动力学和取向在许多重要的细胞过程中起着关键作用。然而，对原代小胶质细胞中的微管和/或微管末端进行功能性活体成像可能具有挑战性。在这里，我们提供了一种在固定和活原生小胶质细胞中观察微管和微管末端的方案。我们描述了小胶质细胞培养和体外刺激、SiR-微管蛋白标记、慢病毒制备、活体成像、免疫染色和图像采集的步骤。我们还提供了 SiR-微管蛋白、EB3-EGFP 和 EB1 分析的程序。有关该方案使用和执行的完整详细信息，请参阅 Rosito 等人。（2023）。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/ecb8467263c2/gr6.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/dda5f62f383d/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/7fe29a5b54d2/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/972f5a3aef3a/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/813f85ba09b6/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/b71d996cef6f/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/8f3b8ff65e48/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/ecb8467263c2/gr6.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/dda5f62f383d/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/7fe29a5b54d2/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/972f5a3aef3a/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/813f85ba09b6/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/b71d996cef6f/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/8f3b8ff65e48/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/782b/10448205/ecb8467263c2/gr6.jpg

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