How do other people freeze down and thaw huvec.
Primary huvec cell culture.
The following day non adherent cells are removed by changing the culture medium.
Product overview lonza s cryopreserved huvecs are shipped as frozen primaries.
Human umbilical vein endothelial cells huvec provide a classic model system to study many aspects of endothelial function and disease such as normal abnormal and tumor associated angiogenesis oxidative stress hypoxia and inflammation related pathways in endothelia under normal and pathological conditions cardiovascular related complications associated with various diseases mode of.
Aspirate the medium from a flask of primary huvec isolated from one or more umbilical cords at or near confluence see refs.
Many of the endothelial processes are commonly studied in vitro using human umbilical vein endothelial cells huvec or large vessel and microvascular endothelial cells isolated from various body tissues.
Keep the medium to surface area ratio at 1ml per 5 cm 2.
Wash the monolayer with 5 ml of hbss volumes are given for a 75 cm 2 flask.
All cells test negative for mycoplasma bacteria yeast and fungi.
Passage normal huvec cells when culture has reached approximately 80 confluence.
Warm both the trypsin edta for primary cells atcc pcs 999 003 and the trypsin neutralizing solution atcc pcs 999 004 to room temperature prior to dissociation.
Use 1 2 as much for 25 cm 2 flasks.
Pipette 20 ml of endothelial cell growth medium to a t 75 flask.
Primary human umbilical vein endothelial cells huvec are isolated from human umbilical vein and cryopreserved at the end of the primary culture the cells are.
Performance testedat least 16 population doublings when cultured in medium 200 supplemented with low serum growth supplement lsgs guarant.
I then spun down and resuspended and placed in a coated t 25 which i could culture fresh primary huvec on but no cells attached.
Add 2 ml of the cell suspension in each fibronectin coated petri dish cultures are placed in a humidified 95 air and 5 co2 atmosphere at 37 c.
Take the endothelial cell growth medium from the refrigerator decontaminate the bottle with 70 alcohol in a sterile hood.
To have good huvec monolayer a critical component is ecm.
Lonza offers a comprehensive range of normal human primary endothelial cells and supporting media for cardiac aortic coronary iliac artery pulmonary artery cardiac microvascular.
Preparing cell culture flasks for culturing huvec.
Coating cell culture dishes both flasks for expansion and or experimental dishes plates wells with 0 1 gelatin is fine.
