after culturing for many days brain microvessels endothelial cells can survive ?
Hi,
Usually companies guarantie that cells with population doubling 15 have still the same phenotype
I use only HUVECs and HAECs and always use cells with cumulative population doubling 9-10 (for my method of passaging it is passage 5).
Never worked with brain microvessels, but would expect, that they would behave like another endothelial cells so take care about the confluency (never exceed 90% - once they become confluent they change phenotype and decrease population doubling speed) and do not work with high passages unless you test that they are still endothelial cells.
I would recommend to work always with cells from the same passage and cumulative population doubling.
Hope this helps
About the days before experiment I never use first passage after defrosting cells.
Have frozen stocks (P3 HUVEC/HAEC) 1mil cells, defrost in petri dish 60cm2 with 10ml media - it takes cca 3 days for cells to get 90% confluent.
Passage the cells again 1mil/petri dish - it takes cca 2 days to become 90% confluent and ready for experiment - I always use thesse cells.
So it takes you 5 days before experiment if you have frozen stocks to get the cells ready.
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