Three methods for examining the de novo proteome of microglia using BONCAT bioorthogonal labeling and FUNCAT click chemistry
Summary: Bioorthogonal labeling and click chemistry techniques allow the detailed examination of cellular physiology through tagging and visualizing newly synthesized proteins. Here, we describe three methods applying bioorthogonal non-canonical amino acid tagging and fluorescent non-canonical amino...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2023-09-01
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| Series: | STAR Protocols |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2666166723003854 |
| _version_ | 1797782655619563520 |
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| author | Alison Keolani Carlisle Jürgen Götz Liviu-Gabriel Bodea |
| author_facet | Alison Keolani Carlisle Jürgen Götz Liviu-Gabriel Bodea |
| author_sort | Alison Keolani Carlisle |
| collection | DOAJ |
| description | Summary: Bioorthogonal labeling and click chemistry techniques allow the detailed examination of cellular physiology through tagging and visualizing newly synthesized proteins. Here, we describe three methods applying bioorthogonal non-canonical amino acid tagging and fluorescent non-canonical amino acid tagging to quantify protein synthesis in microglia. We describe steps for cell seeding and labeling. We then detail microscopy, flow cytometry, and Western blotting techniques. These methods can be easily adapted for other cell types to explore cellular physiology in health and disease.For complete details on the use and execution of this protocol, please refer to Evans et al. (2021).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. |
| first_indexed | 2024-03-13T00:14:02Z |
| format | Article |
| id | doaj.art-ee6d62337e0e4c569509813152def57a |
| institution | Directory Open Access Journal |
| issn | 2666-1667 |
| language | English |
| last_indexed | 2024-03-13T00:14:02Z |
| publishDate | 2023-09-01 |
| publisher | Elsevier |
| record_format | Article |
| series | STAR Protocols |
| spelling | doaj.art-ee6d62337e0e4c569509813152def57a2023-07-12T04:20:41ZengElsevierSTAR Protocols2666-16672023-09-0143102418Three methods for examining the de novo proteome of microglia using BONCAT bioorthogonal labeling and FUNCAT click chemistryAlison Keolani Carlisle0Jürgen Götz1Liviu-Gabriel Bodea2Clem Jones Centre for Ageing and Dementia Research, Queensland Brain Institute, The University of Queensland, St Lucia, QLD 4072, AustraliaClem Jones Centre for Ageing and Dementia Research, Queensland Brain Institute, The University of Queensland, St Lucia, QLD 4072, AustraliaClem Jones Centre for Ageing and Dementia Research, Queensland Brain Institute, The University of Queensland, St Lucia, QLD 4072, Australia; Corresponding authorSummary: Bioorthogonal labeling and click chemistry techniques allow the detailed examination of cellular physiology through tagging and visualizing newly synthesized proteins. Here, we describe three methods applying bioorthogonal non-canonical amino acid tagging and fluorescent non-canonical amino acid tagging to quantify protein synthesis in microglia. We describe steps for cell seeding and labeling. We then detail microscopy, flow cytometry, and Western blotting techniques. These methods can be easily adapted for other cell types to explore cellular physiology in health and disease.For complete details on the use and execution of this protocol, please refer to Evans et al. (2021).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.http://www.sciencedirect.com/science/article/pii/S2666166723003854Cell CultureFlow Cytometry/Mass CytometryCell-based AssaysMicroscopyNeuroscienceProtein Biochemistry |
| spellingShingle | Alison Keolani Carlisle Jürgen Götz Liviu-Gabriel Bodea Three methods for examining the de novo proteome of microglia using BONCAT bioorthogonal labeling and FUNCAT click chemistry STAR Protocols Cell Culture Flow Cytometry/Mass Cytometry Cell-based Assays Microscopy Neuroscience Protein Biochemistry |
| title | Three methods for examining the de novo proteome of microglia using BONCAT bioorthogonal labeling and FUNCAT click chemistry |
| title_full | Three methods for examining the de novo proteome of microglia using BONCAT bioorthogonal labeling and FUNCAT click chemistry |
| title_fullStr | Three methods for examining the de novo proteome of microglia using BONCAT bioorthogonal labeling and FUNCAT click chemistry |
| title_full_unstemmed | Three methods for examining the de novo proteome of microglia using BONCAT bioorthogonal labeling and FUNCAT click chemistry |
| title_short | Three methods for examining the de novo proteome of microglia using BONCAT bioorthogonal labeling and FUNCAT click chemistry |
| title_sort | three methods for examining the de novo proteome of microglia using boncat bioorthogonal labeling and funcat click chemistry |
| topic | Cell Culture Flow Cytometry/Mass Cytometry Cell-based Assays Microscopy Neuroscience Protein Biochemistry |
| url | http://www.sciencedirect.com/science/article/pii/S2666166723003854 |
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