Mouse Brain Microvascular Endothelial Cells

Description

Catalog Number M1011
Product Name Mouse brain microvascular endothelial cells
Storage 37°C C0₂ incubator or Liquid Nitrogen
Product Format Proliferating culture
Cells Number >90% confluent in T25 flask
*Caution: Although primary cells are tested pathogen-free, investigators should handle these cells with caution and treat all animal cells as
potential pathogens, since no test procedure can completely guarantee the absence of infectious agents. Proper precautions must be taken to
avoid exposure. Always wear proper protective equipment (Gloves, safety glasses, etc.) when handling these materials. We recommend
following the universal procedures for handling products of animal origin as the minimum precaution against contamination.
GENERAL INFORMATION
Mouse brain microvascular endothelial cells were isolated from E18 embryonic brain cortex
microvessels of Balb/C mouse and transformed by transfected with SV40 middle T antigen. The cells are
shipped in proliferating culture or frozen vial with a confluence of > 90% (the cells are provided at
passage 4-6). Cells are tested negative for common experimental animal pathogens, and mycoplasma in
vitro. ENDO-Growth medium (EGM-2102) containing 5% serum and growth supplement is
recommended for the expansion of these can be propagated to passage 6 and beyond without losing
their morphologic and phenotypic characteristics. When you receive the cells, leave the flask in 37°C C02
incubator for 1 hour first and then replace the transport medium with fresh ENDO-Growth medium
(EGM-2102). Let the cells grow for 24 hours before subculture.
CELL CHARACTERIZATION
PECAM1 >95% positive by immunofluorescence
VE-Cadherin >95% positive by immunofluorescence
Mouse brain microvascular endothelial cells Negative for mycoplasma
PRODUCT USE AND SHIPPING STATUS
Product Use Mouse brain microvascular endothelial cells are
for research only
Shipping Status Proliferating culture inT25 flask

T25 flask
*Coating T25 flasks. Add 2 ml AlphaBioCoat (AC001) into 3- T25 flasks and ensure entire interior
surface is coated with the solution. After 30 minutes, dispose of AlphaBioCoat (AC001) by
aspiration. Gently rinse and aspirate the flask with Phosphate Buffer Solution (1XPBS-001). The
flask is now ready for use (no need for overnight incubation when coated with AC001). Add
fresh media to flask, if color changes from pink to yellow, discard the media, and add fresh
media to each flask.
1. Inspect to make sure Flask is at 90% confluence, if not remove transport media, and add 5ml of
fresh media to the flask. Place flask in 37°C incubator until cells are at 90% confluence. Change
media every 2 days.
2. If flask is at 90% confluence, aspirate transport media from flask.
3. Rinse T25 flask containing cells with 5 ml 1XPBS (1XPBS-001).
4. Gently aspirate out the PBS after rinsing, and discard.
5. Add 2ml of RT trypsin/ EDTA to T25 flask containing cells (ensure entire interior surface is
covered.
6. Place T25 flask containing cells into 37°C incubator for 1 or 2 minutes (cells will normally come
off of the surface within 1 or 2 minutes).
7. Suspend the cells with 15ml of ENDO-Growth medium (EGM-2102) and transfer equally into 3
pre-coated T25 flasks (the cells are now at a subculture ratio of 1:3.
8. There is no need to spin cells during subculture.
9. Proliferating cell culture: ENDO-Growth medium (EGM-2102) should be changed every 2 days.
The cells normally become confluent within 7 days (when split at a 1:3 ratio)
10. Use ENDO- Basal media (EBM-002) containing 0.5% FBS to induce quiescent cells (after 18-24
hours).