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Immortalized-Mouse-Cerebellar-Capillary-Endothelial-Cells-(cerebEND)-

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Immortalized-Mouse-Cerebellar-Capillary-Endothelial-Cells-(cerebEND)-

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Immortalized-Mouse-Cerebellar-Capillary-Endothelial-Cells-(cerebEND)-

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	Print Version
BioSafety Level	II
Organism	Neonatal mouse (WT 129 Sv strain)
Source Organ	Cerebellar cortex
Growth Properties	Adherent
Morphology	spindle shaped
Recommended Seeding Density	Recommended split ratio: no greater than 1:4
Markers	Claudin-5, Occuldin, Glut-1, VE-Cadherin
Applications	For Research Use Only
Immortalization Method	Transformation with oncoprotein of murine Polyomavirus, Polyoma middle T antigen (PymT)
Description	The mammalian cerebellum is indispensable in the coordination of voluntary movement (i.e. balance and locomotion), muscular tonus and equilibrium. The cerebellar blood brain barrier (BBB) helps maintain the homeostasis of the cerebellum microenvironment and its integrity is often connected to many disorders of the central nervous system (CNS). The Immortalized Mouse Cerebellar Capillary Endothelial Cell Line (cerebEND) provides a useful model for studies involving the differentiation and regulation of BBB as it shares principal features of the BBB in vivo, specifically in that 1) it retains endothelial markers VE-Cadherin and PECAM-1; 2) it displays tight junction associated markers such as occludin, claudin-3, -5, -12, 3) it expresses the BBB marker protein Glut-1 and 4) it shows high electrical resistance, representing barrier properties. This cell line is also responsive to glucocorticoid, estrogen-treatment and pro-inflammatory mediator such as TNFα, making this cell line valuable in elucidating cellular responses of endothelial cells to different stimuli. Compared to the Immortalized Mouse Cerebral Capillary Endothelial Cell Line (cEND), the cerebEND shows a higher fragility of the cerebellar BBB to inflammatory stimuli; together the two cell lines represent important model systems for these different brain regions.
Procedure Overview
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Propagation	Use of PriCoat? T25 Flasks (G299) or Applied Cell Extracellular Matrix (G422) is required for cell adhesion to the culture vessels. Grow cells in ECM-coated culture vessels with the following conditions. The base medium for this cell line is Prigrow III medium available from ABM (TM003). To make the completed growth medium, add the following components to the base medium: heat-inactivated fetal bovine serum (TM999) to final concentration of 10% and 1% Penicillin/Streptomycin (G255). Atmosphere: air: 95%, CO?: 5%; Temperature: 37.0°C. The Differentiation Medium is composed of 2% heat-inactivated fetal bovine serum (TM999) instead of 10%. Culturing cerebEND cells in serum-reduced medium will lead to an increase in TER as well as change in cell morphology (from spindle-shaped to cobble-stone like). TER can be further increased by the addition of 110nM hydrocortisone and/or 1μM insulin into the differentiation medium.
Preservation	1. Freeze Medium: Complete growth medium with 20% FBS and 10% DMSO. 2. Storage Temperature: Liquid nitrogen vapour phase.
Quality Control	1) Endothelial marker VE-Cadherin and PECAM-1 assessed by immunostainning; 2) Tight junction protein Claudin-5 assessed by immunostainning and claudin-1, -3 and -12 detected in mRNA levels; 2) BBB marker protein Glut-1 and tight junction-associated protein occludin measured by western blot analysis. The cell line's response to pro-inflammatory stimuli was measured by 1) TER, 2) western blot and 3) gene expression analysis using RT-PCR.
Disclaimer	1. The CoA for this product (provided upon request) verifies the cell-type specific gene expression via RT-PCR only. All test parameters provided in the CoA are conducted using abm's standardized culture system and procedures. The stated values may vary under the end-user's culture conditions. Please verify that the product is suitable for your studies by referencing published papers or ordering RNA (0.5 μg, Cat.# C207, $450.00) or cell lysate (100 μg, Cat.# C206, $600.00) to perform preliminary experiments, or alternatively use our Gene Expression Assay Service (Cat# C138). All sales are final. 2. We strongly recommend live cell shipments for ease of cell transfer and this option can be requested at the time of ordering. Please note that the end-user will need to evaluate the feasibility of live cell shipment by taking into account the final destination's temperature variation and its geographical location. In addition, we thoroughly test our cell lines for freeze-thaw recovery. If frozen cells were received and not recovered in your lab under the exact, specified conditions (using recommended culture vessel, media, additional supplements, and atmospheric conditions), a live cell replacement is possible at a cost (plus shipping). 3. All of abm's cell biology products are for research use ONLY and NOT for therapeutic/diagnostic applications. abm is not liable for any repercussions arising from the use of its cell biology product(s) in therapeutic/diagnostic application(s). Please contact a technical service representative for more information. 4. abm makes no warranties or representations as to the accuracy of the information on this site. Citations from literature and provided for informational purposes only. abm does not warrant that such information has been shown to be accurate.