ECM630 | Fibrin In Vitro Angiogenesis Assay

ECM630
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      Overview

      Replacement Information

      Key Spec Table

      Key Applications
      ACT
      Description
      Catalogue NumberECM630
      Brand Family Chemicon®
      Trade Name
      • Chemicon
      DescriptionFibrin In Vitro Angiogenesis Assay
      OverviewIntroduction

      Angiogenesis is the process of generating new capillary blood vessels. It is a fundamental component of a number of normal (reproduction and wound healing) and pathological processes (diabetic retinopathy, rheumatoid arthritis, tumor growth and metastasis)1.

      The CHEMICON Fibrin Gel In Vitro Angiogenesis Assay Kit provides a convenient system for evaluation of tube formation by endothelial cells in 96-well or other formats. When cultured on top or within a fibrin gel, endothelial cells rapidly align and form interconnecting networks that can display patent lumina2,3. Tube formation is a multi-step process involving cell adhesion, migration, differentiation and growth4. The formation of intercellular connections and lumina within endothelial cell networks in fibrin gels is dependent upon the actions of VE-cadherin, avb3 and a5b1 integrins, the cdc42 and Rac1 GTPases, and membrane-type matrix metalloproteinases (MT-MMPs)2,3,5,6. Angiogenesis within fibrin gels in vitro is regarded as an accurate model for wound healing and tumor angiogenesis, as tumor cell-derived vascular endothelial growth factor/vascular permeability factor promotes leakage of fibrinogen from the tumor vasculature and formation of a fibrin-rich proangiogenic provisional matrix7.

      Fibrin gel formation is initiated by enzymatic cleavage of fibrinogen, a heterotrimer, by thrombin8. The resulting cleaved fibrin molecules form regular, multimolecular arrays that are highly translucent. The concentrations and formulations of the fibrinogen and thrombin in this kit are optimized for maximal tube-formation by HUVEC and easy visualization of these tubes.
      Materials Required but Not Delivered1. 96-well or other sized Tissue Culture plate

      2. Microcentrifuge Tubes, sterile

      3. 37°C Tissue Culture Incubator

      1. Inverted Light Microscope

      2. HUVEC cells or other experimental cell line (any species), and appropriate growth medium and subculturing reagents. Early passage cells are preferred, as they are less prone to cell death in this assay.

      3. Pipet capable of delivering 100-200 μL, preferably a multichannel pipet with 96-well plate

      4. Basal medium - EBM (Cambrex/BioWhittaker) or MCDB131 (Sigma) supplemented with 1% BSA and antibiotic.

      5. PMA (optional) to add to basal media and angiogenic supplement for positive control.
      References
      Product Information
      Components
      • Fibrinogen Solution: (Part No. 90244) One 10 mL bottle.
      • Thrombin solution: (Part No. 90246) One 7.5 mL bottle.
      • 100x Positive Control Angiogenic Supplement: (Part No. 90247) One 0.5 mL vial. (Contains1.0 mg/ml insulin from bovine pancreas, 0.55 mg/ml human transferrin (substantially iron-free), and 0.5 μg/ml sodium selenite in EBSS without phenol red).
      Applications
      ApplicationThe Fibrin Gel In Vitro Angiogenesis Assay Kit represents a simple model of angiogenesis in which the induction or inhibition of tube formation by exogenous signals can be easily monitored.
      Key Applications
      • Activity Assay
      Application NotesApplication

      The CHEMICON Fibrin Gel In Vitro Angiogenesis Assay Kit represents a simple model of angiogenesis in which the induction or inhibition of tube formation by exogenous signals can be easily monitored. Fibrin gels are easily and quickly formed in culture dishes by mixing Fibrinogen and Thrombin Solutions. For assaying inhibitors or stimulators of tube formation, simply premix the endothelial cell suspension with different concentrations of the inhibitor or stimulator to be tested, before adding the cells to the top of the fibrin gel. Addition of a second layer of fibrin gel on the day after plating the cells is optional, but will promote optimal survival and a higher degree of network and lumen formation. The assay can be used to monitor the extent of tube assembly in various endothelial cells, e.g. human umbilical vein cells (HUVEC) or bovine capillary endothelial (BCE) cells.
      Biological Information
      Species Reactivity
      • Vertebrates
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Usage Statement
      • Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
      Storage and Shipping Information
      Storage ConditionsStore unopened kit materials at -20°C or -70°C for up to their expiration date. Once Fibrinogen Solution has been thawed, store at room temperature for up to two weeks. Thawed Thrombin Solution should be stored at 4ºC for up to one month. Thawed 100x Positive Control Angiogenic Supplement can be stored at 4ºC for up to one year.
      Packaging Information
      Material Size1 kit
      Transport Information
      Supplemental Information
      Specifications

      Documentation

      SDS

      Title

      Safety Data Sheet (SDS) 

      References

      Reference overviewPub Med ID
      Mesenchymal stem cells secrete multiple cytokines that promote angiogenesis and have contrasting effects on chemotaxis and apoptosis.
      Robert A Boomsma,David L Geenen
      PloS one  7  2012

      Show Abstract
      22558198 22558198
      High-throughput production of gene replacement mutants in Neurospora crassa.
      Gyungsoon Park,Hildur V Colot,Patrick D Collopy,Svetlana Krystofova,Christopher Crew,Carol Ringelberg,Liubov Litvinkova,Lorena Altamirano,Liande Li,Susan Curilla,Wei Wang,Norma Gorrochotegui-Escalante,Jay C Dunlap,Katherine A Borkovich
      Methods in molecular biology (Clifton, N.J.)  722  2011

      Show Abstract
      21590421 21590421
      Direct interactions of intraflagellar transport complex B proteins IFT88, IFT52, and IFT46.
      Ben F Lucker,Mark S Miller,Slawomir A Dziedzic,Philip T Blackmarr,Douglas G Cole
      The Journal of biological chemistry  285  2010

      Show Abstract Full Text Article
      20435895 20435895
      Coupling aerobic biodegradation of methanol vapors with heterologous protein expression of endochitinase Ech42 from Trichoderma atroviride in Pichia pastoris.
      Sonia Arriaga,Julia A Acosta-Munguía,Ana S Pérez-Martínez,Antonio De León-Rodríguez,Ana P Barba de la Rosa
      Bioresource technology  101  2010

      Show Abstract
      20709543 20709543
      The anti-viral protein of trichosanthin penetrates into human immunodeficiency virus type 1.
      Wenlong Zhao,Du Feng,Shan Sun,Ting Han,Senfang Sui
      Acta biochimica et biophysica Sinica  42  2010

      Show Abstract
      20119629 20119629
      The transcriptionally active amyloid precursor protein (APP) intracellular domain is preferentially produced from the 695 isoform of APP in a {beta}-secretase-dependent pathway.
      Nikolai D Belyaev,Katherine A B Kellett,Caroline Beckett,Natalia Z Makova,Timothy J Revett,Natalia N Nalivaeva,Nigel M Hooper,Anthony J Turner
      The Journal of biological chemistry  285  2010

      Show Abstract Full Text Article
      20961856 20961856
      A novel sorting strategy of trichosanthin for hijacking human immunodeficiency virus type 1.
      Wen-Long Zhao,Fan Zhang,Du Feng,Ju Wu,Shan Chen,Sen-Fang Sui
      Biochemical and biophysical research communications  384  2009

      Show Abstract
      19409877 19409877
      Subcellular localization and expression of bamboo mosaic virus satellite RNA-encoded protein.
      Palani, PV; Chiu, M; Chen, W; Wang, CC; Lin, CC; Hsu, CC; Cheng, CP; Chen, CM; Hsu, YH; Lin, NS
      The Journal of general virology  90  507-18  2009

      Show Abstract Full Text Article
      19141462 19141462
      Recombinant norovirus-specific scFv inhibit virus-like particle binding to cellular ligands.
      Khalil Ettayebi,Michele E Hardy
      Virology journal  5  2008

      Show Abstract Full Text Article
      18237416 18237416
      Association of hepatitis C virus replication complexes with microtubules and actin filaments is dependent on the interaction of NS3 and NS5A.
      Chao-Kuen Lai,King-Song Jeng,Keigo Machida,Michael M C Lai
      Journal of virology  82  2008

      Show Abstract Full Text Article
      18562541 18562541

      Brochure

      Title
      Advancing cancer research: From hallmarks & biomarkers to tumor microenvironment progression
      EpiGRO™, EndoGRO™ and FibroGRO™ Reagents

      Data Sheet

      Title
      Fibrin In Vitro Angiogenesis Assay - Data Sheet
      Fibrin In Vitro Angiogenesis Assay - Data Sheet

      Posters

      Title
      Poster: Tumor Metastasis

      FAQ

      QuestionAnswer
      Can ECM630 be used for vascular endothelial cells other than HUVEC?There is currently literature showing that human dermal micorvascular EC, bovine pulmonary artery EC, and bovine aortic EC can undergo differentiation in fibrin gels as in ECM630.
      How many cells per well have been tested in the kit?In-house usage is 50,000 cells/well of a 24 well plate for the standard assay. Higher concentrations tested yield a monolayer with less tube formation.

      Related Products & Applications

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      Categories

      Life Science Research > Cell Analysis > Cell-based Assays > Angiogenesis & Endothelial Transmigration Assays