Directed In Vivo Angiogenesis Assay
Directed In Vivo Angiogenesis Assay Summary
Why Use Directed In Vivo Angiogenesis Assay?
DIVAA is the first in vivo system for the study of angiogenesis that provides quantitative and reproducible results. During the course of the assay, implant grade silicone cylinders closed at one end, called angioreactors, are filled with 20 μl of Cultrex Basement Membrane Extract (BME) premixed with or without angiogenic-modulating factors. These angioreactors are then implanted subcutaneously in the dorsal flank of nude mice. Accompanied with the onset of angiogenesis, vascular endothelial cells proceed to grow into the BME and form vessels in the angioreactor. As early as nine days post-implantation, there are enough cells to determine an effective dose response to angiogenic modulating factors. The sleek design of the angioreactor provides a standardized platform for reproducible and quantifiable in vivo angiogenesis assays. Compared to the plug assay, the angioreactor prevents assay errors due to absorption of the basement membrane extract by the mouse. In addition, the angioreactor uses only a fraction of the materials conserving both BME and test compounds used, and up to four angioreactors may be implanted in each mouse, allowing for greater statistical power. DIVAA has been used in a variety of investigations.
• DIVAA BME
• DIVAA FGF-2 Positive Control
• DIVAA Cell Sperse
• DIVAA FGF-2 (300 ng)/VEGF(100 ng)
• DIVAA Angioreactor
• DIVAA Wash Buffer
• DIVAA FITC-Lectin 200X
• 25X FITC-Lectin Diluent
• Heparin Solution
Note: The components for this kit may require different storage/shipping temperatures and may arrive in separate packaging.
For research use only. Not for diagnostic use.
Citations for Directed In Vivo Angiogenesis Assay
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
Citations: Showing 1 - 7
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Interleukin-1 Receptor Type 2 Acts with c-Fos to Enhance the Expression of Interleukin-6 and Vascular Endothelial Growth Factor A in Colon Cancer Cells and Induce Angiogenesis.
Authors: Mar A, Chu C, Lee H, Chien C, Cheng J, Yang S, Jiang J, Lee T
J Biol Chem, 2015;290(36):22212-24. 2015
A urokinase receptor-derived peptide inhibiting VEGF-dependent directional migration and vascular sprouting.
Authors: Bifulco K, Longanesi-Cattani I, Liguori E, Arra C, Rea D, Masucci M, De Rosa M, Pavone V, Stoppelli M, Carriero M
Mol Cancer Ther, 0;12(10):1981-93. 0
Syndecan-4 is a major syndecan in primary human endothelial cells in vitro, modulated by inflammatory stimuli and involved in wound healing.
Authors: Vuong T, Reine T, Sudworth A, Jenssen T, Kolset S
J Histochem Cytochem, 0;63(4):280-92. 0
Cryptosporidium parvum induces an endoplasmic stress response in the intestinal adenocarcinoma HCT-8 cell line.
Authors: Morada M, Pendyala L, Wu G, Merali S, Yarlett N
J Biol Chem, 0;288(42):30356-64. 0
High Aldehyde Dehydrogenase Activity Identifies a Subset of Human Mesenchymal Stromal Cells with Vascular Regenerative Potential.
Authors: Sherman S, Kuljanin M, Cooper T, Putman D, Lajoie G, Hess D
Stem Cells, 0;35(6):1542-1553. 0
Liver X receptor activation reduces angiogenesis by impairing lipid raft localization and signaling of vascular endothelial growth factor receptor-2.
Authors: Noghero A, Perino A, Seano G, Saglio E, Lo Sasso G, Veglio F, Primo L, Hirsch E, Bussolino F, Morello F
Arterioscler Thromb Vasc Biol, 0;32(9):2280-8. 0
Soluble carcinoembryonic antigen activates endothelial cells and tumor angiogenesis.
Authors: Bramswig K, Poettler M, Unseld M, Wrba F, Uhrin P, Zimmermann W, Zielinski C, Prager G
Cancer Res, 0;73(22):6584-96. 0
What is DIVAA™?
The Directed In Vivo Angiogenesis Assay (DIVAA™) is the first in vivo system for the study of angiogenesis that provides quantitative and reproducible results. Angioreactors (silicone cylinders closed at one end) containing 20 µL of basement membrane with/without angiogenic-modulating factors are implanted subcutaneously in the dorsal flank of nude mice. With the onset of angiogenesis, vascular endothelial cells proceed to grow into the basement membrane extract and form vessels in the angioreactor. As early as nine days post-implantation, there are enough cells to determine an effective dose response to angiogenic modulating factors.
What strains of mice has DIVAA™ been used with?
DIVAA™ has been used with nude mice, the recommended and qualified strain for the assay. The mouse strain C57Bl has reportedly been used successfully with the kit. However, optimization is required, and variability is commonly reported. If you require C57BI mice for your experiments, it is recommended to allow the implantation to go for up to 15 days.
Are materials supplied in the DIVAA™ kit sterile?
Yes, the materials supplied are tested following USP <71> sterility guidelines.
Why does DIVAA™ contain a FGF-2/VEGF mixture to promote angiogenesis?
While FGF-2 and VEGF have both been demonstrated to promote angiogenesis in DIVAA™, the FGF-2/VEGF mixture provides a synergistic effect allowing a drastic increase in response using less total growth factor.
What is the difference between the FITC-Lectin and FITC-Dextran in the DIVAA protocol?
FITC-Lectin specifically binds to endothelial cells, so it counts the total number of endothelial cells contained within the angioreactor. FITC-Dextran does not bind endothelial cells. Instead, it is dispersed within the blood of the mouse, being evenly distributed within the blood vessels, so it quantitates the total volume of blood contained within the vessels within the angioreactor. Both procedures have demonstrated equivalent results empirically.
Where are the DIVAA™ angioreactors implanted?
The angioreactors should be implanted on the host mouse dorsal lateral flanks. These areas exhibit sufficient vasculature to support initiation of angiogenesis, and they are also strategically positioned to prevent removal by the host mouse. Other areas may be used for implantation; however, this is outside of the product specification. Please consult the scientific literature for more information.
Can cells be placed within the angioreactor?
While some groups have had success with implanting angioreactors containing viable cells, this application has not been qualified by R&D Systems. Please consult the scientific literature for more information.
What strains of mice have been evaluated using the DIVAA kit?
The DIVAA kit has been evaluated using nude mice. Although C57Bl/6 mice can be used for this assay, we recommend the use of nude mice for optimal results.
Around the angioreactor in the mouse, a capsule forms with time. When harvesting the angioreactor, it is difficult to dissect the capsule tissue from the opening of the angioreactor. There is a plug of tissue growing into the chamber. Is this normal? Do you have any recommendations on how to handle chambers like this?
Growth of connective tissue into the angioreactor during angiogenesis is normal. The amount of connective tissue is dependent on the strain of mice, treatment, and response. The cell dissociation step may be increased up to 3 hours to improve dissociation/degradation, and any remaining connective tissue may be removed. If this does not solve the problem, it may be more appropriate to use the Optional Protocol for Dextran-FITC Detection (which measures capillary volume and discards cellular/connective tissue debris).
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