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Isolation of Endothelial Cells from Mice
Published: Apr 20, 2012 DOI: 10.21769/BioProtoc.205 Views: 22314
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Abstract
Endothelial cells line the entire circulatory system, from the heart to the smallest capillary. In this protocol, attaining and maintaining vitality of the organ is critical. Minimizing the processing time between removing the organ and beginning the digestions will improve yields as cells are more viable and are not under hypoxic conditions for long. For this protocol to be most effective, mice should be 5-6 weeks of age. Older mice result in lower yields of endothelial cells. In our hands, as few as 5 mice can result in good yields.
Materials and Reagents
- Endothelial cells (EC)
- Endothelial cell growth supplement (ECGS) (Sigma-Aldrich, catalog number: E2759 )
- Collagenase I (Life Technologies, InvitrogenTM, catalog number: 17100-017 )
- Anti-CD31 antibody (BD Biosciences, PharmingenTM, catalog number: 550274 )
- BSA
- DMEM
- Fetal calf serum (FCS)
- Fetal bovine serum (FBS)
- EDTA
- Phosphate buffered saline (PBS)
- Antibiotic
- Heparin
- F12 medium
- Isoflourane
- 70% ethanol
- Dynabeads
- M199
- Gelatin
- Pre-warmed digestion solution
- Digestion solution (see Recipes)
- Culture media (see Recipes)
Equipment
- Centrifuges
- Sterile culture hood
- Magnet
Procedure
Category
Cell Biology > Cell isolation and culture > Cell isolation
Cell Biology > Tissue analysis > Tissue isolation
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