In Vitro Angiogenesis Assay Kit

Protocol tips

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	- Cells are treated with EBM-2 and observed microscopically and photographed after 4 h incubation.

Protocol tips
- Cells are treated with EBM-2 and observed microscopically and photographed after 4 h incubation.

Publication protocol

To evaluate the effectiveness of PAM for preventing angiogenesis, a tube formation assay was performed, as described previously31. In brief, HRECs (Cell Systems; Kirkland, WA, USA) were cultured with endothelial basal medium (EBM)-2 (Lonza, Tokyo, Japan) in an incubator with 5% CO2 in air. Extracellular matrix gels were prepared using a Chemicon in vitro angiogenesis assay kit (EMD Millipore, Billerica, MA). Gels were solidified in wells of a 96-well microplate, and 1.0 × 104 HRECs were added to the surface of each gel in EBM-2-based PAM or control media. After 4 h of incubation, the images of formed tubes were taken using a phase contrast microscope (FSX-100, Olympus, Tokyo, Japan). The numbers of branching points per area were calculated using ImageJ as previously described


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