Cited by 763Angiogenesis and VEGF in CancerCell Adhesion Molecules ResearchCellular Mechanics and Interactions

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In vitro angiogenesis: endothelial cell tube formation on gelled basement membrane extract

Arnaoutova, Irina, Kleinman, Hynda K·2010·Nature Protocols

DOI 10.1038/nprot.2010.6

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Abstract

A protocol is presented here for a rapid, quantitative and reliable in vitro angiogenesis assay that can be adapted for high throughput use. Endothelial cells are plated on a gelled basement matrix, their natural substrate, and form capillary-like structures with a lumen. The assay can be used to identify inhibitors or stimulators of angiogenesis, as well as genes and signaling pathways involved in angiogenesis. It has also been used to identify endothelial progenitor cells. This assay involves endothelial cell adhesion, migration, protease activity and tubule formation. This tube formation assay is preferred, as other in vitro assays for angiogenesis, such as cell adhesion, migration and invasion, measure limited steps in the angiogenesis process. The tube formation assay on basement membrane can be completed in a day because transformed endothelial cells form tubes within 3 h, whereas non-transformed endothelial cells form tubes within 6 h.

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“The capacity to assemble into capillary-like tubules is a hallmark phenotype of endothelial cells, 59 and the cells derived via FGFR c-isoform activation demonstrated a robust 2D network formation capacity (measured by numbers of nodes, segments, and meshes in a tube formation assay) (Figures 5F…”
Background
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“Tube formation assay The endothelial tube formation assay was performed as described by Arnaoutova and co-workers (Arnaoutova and Kleinman 2010).”
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“Endogenous endothelial (CD31 + ) cells are also present at the migrating cell front within the matrix between channels, where both individual cells and more chord-like structures are observed [29, 30].”
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Influential
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“In vitro tube formation assay In vitro tube formation assays were performed according to the protocol described by Arnaoutova and Kleinman (2010). Briefly, Cultrex basement membrane extract (BME) with reduced growth factors”
Netrin-4 promotes mural cell adhesion and recruitment to endothelial cells
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“Experiments were performed a minimum of three times.(25,33,34)”

Authors: Arnaoutova, I., & Kleinman, H. K.
Affiliations: Trevigen (US)
National Institutes of Health (US)
Corresponding author: Arnaoutova, Irina
Kleinman, Hynda K
ORCID: Kleinman, Hynda K
Venue: Nature Protocols
Publisher: Springer Science and Business Media LLC
Published: 2010-03-11
Volume: 5
Issue: 4
Pages: 628-635
ISSN: 1750-2799 (ISSN-L)
1754-2189
DOI: 10.1038/nprot.2010.6
OpenAlex: W2015186671
Cited by: 760
References: 35
Citations by year: 2026: 29 · 2025: 60 · 2024: 55 · 2023: 54 · 2022: 60 · 2021: 62 · 2020: 57 · 2019: 56 · 2018: 59 · 2017: 66 · 2016: 54 · 2015: 31 · 2014: 41 · 2013: 33 · 2012: 26
FWCI: 9.89
Type: journal article
Language: en
Open access: closed
Licence: www.springer.com/tdm (tdm)
Locations: Nature Protocols (publishedVersion)
PubMed (publishedVersion)
Topics: Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Angiogenesis and VEGF in Cancer
Health Sciences › Medicine › Immunology and Allergy › Cell Adhesion Molecules Research
Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular Mechanics and Interactions
Keywords: in vitro angiogenesis assay, endothelial tube formation, basement membrane extract, capillary-like structures, endothelial cell adhesion, endothelial cell migration, proteolytic activity, endothelial progenitor cells, angiogenesis inhibitors, high-throughput screening
MeSH terms: Basement Membrane, Capillaries, Cells, Cultured, Cytological Techniques, Gels, Humans, Neovascularization, Physiologic, Endothelial Cells, In Vitro Techniques

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In vitro angiogenesis: endothelial cell tube formation on gelled basement membrane extract

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