Protocol abstract
Nature Protocols 2, - 2918 - 2923 (2007)
Published online: 8 November 2007 | doi:10.1038/nprot.2007.412
Subject Categories: Cell and developmental biology | Imaging | Model organisms | Pharmacology and toxicology
The zebrafish/tumor xenograft angiogenesis assay
Stefania Nicoli1 & Marco Presta1
Abstract
Here we describe a method to study tumor angiogenesis in zebrafish (Danio rerio) based on the injection of proangiogenic mammalian tumor cells into the perivitelline space of zebrafish embryos at 48 h post-fertilization. Within 24–48 h, proangiogenic tumor grafts induce a neovascular response originating from the developing subintestinal vessels. This can be observed at macroscopic and microscopic levels after whole-mount alkaline phosphatase staining of wild-type zebrafish embryos, or by fluorescence microscopy in transgenic VEGFR2:G-RCFP embryos in which endothelial cells express the green fluorescent protein under the control of the VEGFR2/KDR promoter. Angiogenesis inhibitors added to the injected cell suspension or to the fish water prevent tumor-induced neovascularization. The assay is rapid and inexpensive, representing a novel tool for investigating tumor angiogenesis and for antiangiogenic drug discovery. Also, gene inactivation by antisense morpholino oligonucleotides injection in zebrafish embryos may allow the identification of genes involved in tumor angiogenesis.
- General Pathology and Immunology, Department of Biomedical Sciences and Biotechnology, University of Brescia Medical School, Viale Europa 11, 25123 Brescia, Italy.
Correspondence to: Marco Presta1 e-mail: presta@med.unibs.it
nature-products
A-Z product listing
- 1-Phenyl-2-thiourea(Sigma-Aldrich)
- 4',6-diamidino-2-phenylindole(Sigma-Aldrich)
- Air incubator(International PBI S.p.A)
- Borosilicate needles(Harvard Apparatus)
- digital camera(Leica)
- Epifluorescence Axiovert 200M microscope(Zeiss)
