Table 2.

Ex vivo angiogenesis assays.

Assay	Setting	Advantage	References
Rat aortic ring assay	Thoracic aorta is dissected, cleaned and cut into rings. Upon serum-starvation, rings are embedded in extracellular matrix components in the presence or absence of the test compound. Exponential vessel outgrowth from the explant of the tubule structures is observed within 10 days	Many rings available from few animals. Supporting cells are included in the formation of vessels. Visible lumenized tubule structures develop over a time course similar to that in vivo	Nicosia, 2009
Mouse aortic ring assay		Cost-efficient transgenic mouse technologies and gene manipulation available. Implementable for high-quality imaging and high-throughput screening	Baker et al., 2011
Miniature ring-supported gel assay	Isolated aortae segments are placed in low volume three-dimensional collagen gel supports, which are casted by a nylon mesh ring that improves the stability of the setting	Optimized system allows better specimen handling, staining, imaging, and a more economical use of extracellular matrix reagents	Reed et al., 2011
Human arterial ring assay	Human umbilical arteries are isolated from umbilical cords, sectioned into rings, and then embedded in extracellular matrix. Tubular structures are quantified by image analysis	Provides a three-dimensional system for identification of genes and drugs that regulate human angiogenesis	Seano et al., 2013
Retinal explant assay	Explanted retina is cut and placed, over a three-dimensional gel with the photoreceptor layer facing upward. Endothelial cell sprouting is observed from day 3 and peaks at day 7	Allows the study of tip endothelial cell angiogenic responses and acute responses of retinal blood vessels at the sprouting front	Rezzola et al., 2014
Fat-tissue microfragment assay	Human subcutaneous fat tissue is fragmented and embedded in fibrin. Blood vessel growth and elongation is examined after 15 days by microscopy	Uses intact human fat tissue with quiescent vessels from which other spontaneously derive. Assay could help predict response toward a treatment	Greenway et al., 2007
Choroid sprouting assay	The choroid, a vascular bed beneath the retinal pigment epithelium, is separated from the retina, segmented, and placed over a matrix. Outgrowth of vascular sprouts can be observed within 2-6 days.	Vascular sprouts consist of endothelial cells, pericytes and macrophages. Robust, reproducible and representative model of microvascular angiogenesis Semi-automated software for quantification of sprouting area is available	Shao et al., 2013