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

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 li...

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Detalles Bibliográficos
Autores principales: Sanchini, Caterina, Rosito, Maria, Comincini, Alessandro, De Panfilis, Simone, Bartolini, Francesca, Di Angelantonio, Silvia
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2023
Materias:
Protocol
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10448205/
https://www.ncbi.nlm.nih.gov/pubmed/37573502
http://dx.doi.org/10.1016/j.xpro.2023.102499
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author Sanchini, Caterina
Rosito, Maria
Comincini, Alessandro
De Panfilis, Simone
Bartolini, Francesca
Di Angelantonio, Silvia
author_facet Sanchini, Caterina
Rosito, Maria
Comincini, Alessandro
De Panfilis, Simone
Bartolini, Francesca
Di Angelantonio, Silvia
author_sort Sanchini, Caterina
collection PubMed
description 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).(1)
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spelling pubmed-104482052023-08-25 Protocol for observing microtubules and microtubule ends in both fixed and live primary microglia cells Sanchini, Caterina Rosito, Maria Comincini, Alessandro De Panfilis, Simone Bartolini, Francesca Di Angelantonio, Silvia STAR Protoc Protocol 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).(1) Elsevier 2023-08-12 /pmc/articles/PMC10448205/ /pubmed/37573502 http://dx.doi.org/10.1016/j.xpro.2023.102499 Text en © 2023 The Author(s) https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Protocol
Sanchini, Caterina
Rosito, Maria
Comincini, Alessandro
De Panfilis, Simone
Bartolini, Francesca
Di Angelantonio, Silvia
Protocol for observing microtubules and microtubule ends in both fixed and live primary microglia cells
title Protocol for observing microtubules and microtubule ends in both fixed and live primary microglia cells
title_full Protocol for observing microtubules and microtubule ends in both fixed and live primary microglia cells
title_fullStr Protocol for observing microtubules and microtubule ends in both fixed and live primary microglia cells
title_full_unstemmed Protocol for observing microtubules and microtubule ends in both fixed and live primary microglia cells
title_short Protocol for observing microtubules and microtubule ends in both fixed and live primary microglia cells
title_sort protocol for observing microtubules and microtubule ends in both fixed and live primary microglia cells
topic Protocol
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10448205/
https://www.ncbi.nlm.nih.gov/pubmed/37573502
http://dx.doi.org/10.1016/j.xpro.2023.102499
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