Matrigel cytometry: a novel method for quantifying angiogenesis in vivo.
Many of the current in vivo methods to evaluate angiogenesis are poorly quantifiable. Recently, the Matrigel plug assay has become the method of choice in many studies involving in vivo testing for angiogenesis. When known angiogenic factors are mixed with Matrigel and injected subcutaneously into mice, endothelial cells migrate into the gel plug. These endothelial cells form vessel-like structures, a process that mimics the formation of capillary networks. Here, we present a modification of the traditional Matrigel assay with improved method to quantify the amount of endothelial cells that incorporate into the plug. The removed plugs were subjected to a mild protease treatment, yielding intact cells. The liberated cells were then stained using an endothelial cell-specific markers, and counted by flow cytometry. This novel combination of FACS analysis with the traditional Matrigel assay improves the ability to quantify in vivo angiogenesis, and for the first time enables to determine the number of migrating and proliferating endothelial cells which reflects the angiogenesis rate.
Adini
A
A
Vascular Biology Program, Department of Surgery, Children's Hospital, and the Department of Ophthalmology, both at Harvard Medical School, Boston, Massachusetts, USA. avner.adini@childrens.harvard.edu
Fainaru
O
O
Udagawa
T
T
Connor
K M
KM
Folkman
J
J
D'Amato
R J
RJ
D043925:Angiogenesis Inducing Agents / Q000494:pharmacology; D000818:Animals; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D003094:Collagen / Q000494:pharmacology*; D004338:Drug Combinations; D042783:Endothelial Cells / Q000187:drug effects* / Q000502:physiology; D004730:Endothelium, Vascular / Q000187:drug effects; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D007797:Laminin / Q000494:pharmacology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D011509:Proteoglycans / Q000494:pharmacology*; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
D043925:Angiogenesis Inducing Agents; D004338:Drug Combinations; D007797:Laminin; D011509:Proteoglycans; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D016222:Fibroblast Growth Factor 2; C059485:matrigel; D003094:Collagen