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Available methods, techniques and models
Isolation
of endothelial cells from tissues
We have developed techniques for isolation
of endothelial cells from tissues by immunomagnetic separation and
fluorescence-activated cell sorting (FACS). EC isolated from tumors and
corresponding normal tissues are a highly valuable resource for detailed
molecular profiling of the properties of these phenotypically different
cells.
Molecular profiling of angiogenesis
This method has been developed to obtain the angiogenesis expression
profile of >50
known angiogenesis factors and their receptors in
tumor cells and tumor vasculature
in vivo.
This real-time PCR based technique
using species specific primes,
can be used to determine the interactions between host- and tumor cells and
allows the analysis of angiostatic treatment on tumor cells and the tumor
vasculature
(Thijssen et al, Exp.Cell Res. 2004).
Endothelial cell migration
Migration and motility of endothelial cells is a fundamental necessity
for the process of angiogenesis. We developed a high-throughput wound
assay endothelial cell migration platform (Peira Scientific Instruments,
Belgium) making use of a 96-well pin tool and a computer assisted
automated digital analysis system.
Images are segmented and scratches are
detected and quantified using sophisticated software algorithms.
Endothelial cell sprout formation
Endothelial cells of choice (freshly isolated or cell lines) are grown
in spheroids of predefined size by hanging drop technology. Once
spheroids are formed, which can be in as early as 4 hours, they are
transferred into 3-dimensional collagen gels. Sprouting will be optimal
after 16-20 hours. An ImageJ based method developed by drs Ballini and Nowak-Sliwinska
(Lausanne, Switzerland) is available for quantification of sprouting.
 
In vivo angiogenesis. Chicken chorioallantoic membrane assay.
Angiogenesis is studied in vivo using this model. A little hatch is made
in the shell on day 3 of development. The chorioallantoic membrane is
visible and development of vasculature can be studied during embryo
development, but also after transplantation of (human) tumor tissues.
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