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Product Name Human Brain MicrovascularAngiogenesis KitCatalog Number EP003Product Format 6,12, and 24 wellStorage 37°CGENRAL INFORMATIONOur Human Brain Microvascular Angiogenesis Kit formed capillary endothelial cells, whichare connected by tight junctions with an extremely high electrical resistivity. This modelwill help overcoming the difficulty of delivering therapeutic agents to specific regions of thebrain presents a major challenge to treatment of most brain disorders. The endothelial cellsinitially form small islands within the culture matrix. They subsequently begin toproliferate and then enter a migratory phase during which they move through the matrix toform threadlike tubule structures with lumens. These gradually join up by (1-2 weeks) toform a network of anastomosing tubules which closely resembles the capillary bed found invivo.This model can be used to study the following disease, but not limited to thoseapplications:1) Late-stage neurological trypanosomiasis ( sleeping sickness)2) Progressive multifocal leukoencephalopathy (PML)3) Meningitis brain abscess epilepsy multiple sclerosis4) De vivo disease Alzheimer’s disease cerebral edema5) Prion and prion-like diseases6) HIV encephalitis7) Rabies8) Neuromyelitis opticaThe Human Brain Microvascular Angiogenesis Kit contains all of the materials necessary toperform multiple angiogenesis assays in a 24-well format. The kit is designed that thetesting materials, i.e. compounds, conditioned media, or tissue explants, can be added intothe system at any time, ranging from the onset of vasculogenesis to advanced angiogenesis.The resulting effect on tubule formation (tubular length, number of branches et al) can bemonitored throughout the whole process under inverted fluorescence microscope.
Reagents and Materials Provided:(1) 1 x vial of mixture of GFP-tagged Human Brain Microvascular ECs and RFP-taggedsupporting cells (-80°C or liquid N2)(2) 1 x 24-well Alpha Coat Solution coated plate (Room temperature, for 2 months)(3) 1 x 500ml of Endo-Growth Medium (4°C)Protocols: Day 11. Pre warm Endo-Growth Medium to 37ºC in a water bath2. Accurately pipette 24ml Endo-Growth Medium into a 50ml Falcon tube;3. Rapidly thaw the vial of cryopreserved cells in a 37ºC water bath;4. Transfer all cells gently into 24ml pre warmed Endo-growth medium;5. Mix well the cells gently using a serological pipette;6. Add 1.0ml of cell suspension to each well of the pre coated 24-well plate.7. Make sure the cells are evenly dispersed in the wells.8. Place the plate in an incubator (37ºC, 5% CO2 and humidified).Day 29. Take the plate from the incubator and examine cells under inverted fluorescencemicroscopy (GFP positive Human Brain Microvascular ECs should sparsely and evenlydistributed among RFP positive human supporting cells).10. Wash the cells one with 2 ml of PBS11. Add 2.0ml of fresh Endo-Growth medium (control) or Experimental media (Endo-Growthmedium, plus pro- or anti-angiogenic regents according to customer’s needs).12. Place the plate back into the incubator. Day 4, 6, 8, 10, 12, and 14…… 13. Replace themedium every 2 days until the end of the experiments.