Fast in vitro protocol for the visualization and quantitative high-throughput analysis of sprouting angiogenesis by confocal microscopy
Summary: We describe an optimized, cost-effective, reproducible, and robust protocol to study sprouting angiogenesis in glass-bottom 96-well plates by confocal microscopy, ideal for screening of drug or shRNA libraries. Effective and stable knockdown of gene expression in primary endothelial cells i...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2021-09-01
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| Series: | STAR Protocols |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S266616672100397X |
| _version_ | 1818717495893164032 |
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| author | Lanette Kempers Ivo van der Bijl Anne-Marieke D. van Stalborch Bas Ponsioen Coert Margadant |
| author_facet | Lanette Kempers Ivo van der Bijl Anne-Marieke D. van Stalborch Bas Ponsioen Coert Margadant |
| author_sort | Lanette Kempers |
| collection | DOAJ |
| description | Summary: We describe an optimized, cost-effective, reproducible, and robust protocol to study sprouting angiogenesis in glass-bottom 96-well plates by confocal microscopy, ideal for screening of drug or shRNA libraries. Effective and stable knockdown of gene expression in primary endothelial cells is achieved by lentiviral transduction. Dynamic behavior of individual cells and fluorescent proteins is analyzed by time-lapse imaging, while competitive advantages in tip cell formation are assessed using mixtures of differentially labeled cell populations. Finally, we present a macro for high-throughput analysis.For complete information on the use and execution of this protocol, please refer to van der Bijl et al. (2020) and Kempers et al. (2021). |
| first_indexed | 2024-12-17T19:36:04Z |
| format | Article |
| id | doaj.art-56ea7521564549ae946a0a00a55e8343 |
| institution | Directory Open Access Journal |
| issn | 2666-1667 |
| language | English |
| last_indexed | 2024-12-17T19:36:04Z |
| publishDate | 2021-09-01 |
| publisher | Elsevier |
| record_format | Article |
| series | STAR Protocols |
| spelling | doaj.art-56ea7521564549ae946a0a00a55e83432022-12-21T21:35:09ZengElsevierSTAR Protocols2666-16672021-09-0123100690Fast in vitro protocol for the visualization and quantitative high-throughput analysis of sprouting angiogenesis by confocal microscopyLanette Kempers0Ivo van der Bijl1Anne-Marieke D. van Stalborch2Bas Ponsioen3Coert Margadant4Sanquin Research and Landsteiner laboratory, Amsterdam University Medical Center, 1066 CX Amsterdam, The NetherlandsSanquin Research and Landsteiner laboratory, Amsterdam University Medical Center, 1066 CX Amsterdam, The NetherlandsSanquin Research and Landsteiner laboratory, Amsterdam University Medical Center, 1066 CX Amsterdam, The NetherlandsMolecular Cancer Research, Center for Molecular Medicine, University Medical Center Utrecht, Utrecht University, 3584 CX Utrecht, The NetherlandsAngiogenesis Laboratory, Department of Medical Oncology, Cancer Center Amsterdam, Amsterdam University Medical Center, 1081 HV Amsterdam, The Netherlands; Corresponding authorSummary: We describe an optimized, cost-effective, reproducible, and robust protocol to study sprouting angiogenesis in glass-bottom 96-well plates by confocal microscopy, ideal for screening of drug or shRNA libraries. Effective and stable knockdown of gene expression in primary endothelial cells is achieved by lentiviral transduction. Dynamic behavior of individual cells and fluorescent proteins is analyzed by time-lapse imaging, while competitive advantages in tip cell formation are assessed using mixtures of differentially labeled cell populations. Finally, we present a macro for high-throughput analysis.For complete information on the use and execution of this protocol, please refer to van der Bijl et al. (2020) and Kempers et al. (2021).http://www.sciencedirect.com/science/article/pii/S266616672100397XCell BiologyCell cultureCell-based AssaysDevelopmental biologyHigh Throughput ScreeningMicroscopy |
| spellingShingle | Lanette Kempers Ivo van der Bijl Anne-Marieke D. van Stalborch Bas Ponsioen Coert Margadant Fast in vitro protocol for the visualization and quantitative high-throughput analysis of sprouting angiogenesis by confocal microscopy STAR Protocols Cell Biology Cell culture Cell-based Assays Developmental biology High Throughput Screening Microscopy |
| title | Fast in vitro protocol for the visualization and quantitative high-throughput analysis of sprouting angiogenesis by confocal microscopy |
| title_full | Fast in vitro protocol for the visualization and quantitative high-throughput analysis of sprouting angiogenesis by confocal microscopy |
| title_fullStr | Fast in vitro protocol for the visualization and quantitative high-throughput analysis of sprouting angiogenesis by confocal microscopy |
| title_full_unstemmed | Fast in vitro protocol for the visualization and quantitative high-throughput analysis of sprouting angiogenesis by confocal microscopy |
| title_short | Fast in vitro protocol for the visualization and quantitative high-throughput analysis of sprouting angiogenesis by confocal microscopy |
| title_sort | fast in vitro protocol for the visualization and quantitative high throughput analysis of sprouting angiogenesis by confocal microscopy |
| topic | Cell Biology Cell culture Cell-based Assays Developmental biology High Throughput Screening Microscopy |
| url | http://www.sciencedirect.com/science/article/pii/S266616672100397X |
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