The vasculature is the first organ that starts to function as an integrated system during embryonic development. Correct vascular development and normal daily function are critical to human health because the circulation functionally supports the demand from growing all other body tissues and organs.
Our 384-well angiogenesis assay is high throughput quantitative platform that can be used to identify chemicals and drugs capable of unintentionally altering VEGFR signaling.
Time course of mCherry stably transfected ECFCs after exposure to angiogenesis inducer with and without VEGFR inhibitor (Vatalanib).
Assay Principle
We use mCherry stably transfected Endothelial Colony Forming Cells for fluorescent-based analysis of tube formation. During the course of the assay, ECFCs are seeded in 384 well plates, then treated with various concentrations of compounds of interest, and then angiogenesis inducer is added. The cells self-organize in tubular networks. In the presence of inhibitors of angiogenesis, tube formation is disrupted. We use high content imager to image whole wells in 384-well plates.
Adjusted and analyzed images of tube formation in control (A, B) and in Vatalanib (VEGFR inhibitor)-treated wells (C, D).
Image Analysis
The images are analyzed by ImageJ and commercial software delivering 20 parameters of tube formation. Heatmaps and dose-response curves are plotted. Cell viability is assessed at the same time using live-dead nuclear stains.
