In Vivo Angiogenesis Assay for HTS
Principal Investigator/Assay Provider:
Thanh N. Doan, PhD, Zygogen, LLC
Identification of novel anti-angiogenic compounds
Pathological angiogenesis contributes to over 70 diseases including cancer, age-related macular degeneration, diabetic blindness, and rheumatoid arthritis. The Angiogenesis Foundation estimates that at least 184 million patients in Western nations could benefit from anti-angiogenic therapy. To provide an in vivo system for screening for anti-angiogenic compounds, Zygogen, LLC, has developed a transgenic zebrafish line that expresses a green fluorescent reporter specifically in blood vessels [TG(VEGR2:GRCFP)] using Zygogen’s Z-TagSM technology. Zygogen also developed automated methods for imaging and quantifying angiogenic blood vessel growth in the zebrafish trunk and methods for automated compound screening. Zebrafish offer the high physiological relevance of an in vivo system with the benefits of a model organism, including short generation time, high fecundity, and easy manipulation. Zebrafish also allow for simultaneous assessment of drug toxicity and efficacy during screening.
For screening, 1 day post fertilization (dpf) zebrafish are arrayed into 384-well plates (1 embryo/well) and treated with 30 M of each test compound. At 2dpf, fluorescent images of each live embryo are captured. The trunk of each zebrafish is manually selected using MetaMorph imaging software, and the intersegmental vessels and their branching arteries are quantified using automated algorithms developed by Zygogen.