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HUVEC serum starving


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#1 am1

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Posted 14 November 2009 - 07:22 PM

Hi all,

I'm trying to use HUVEC for MAPK signaling assay.

Before I stimulate them with H2O2, I serum starved them in RPMI + 1% FBS for 16 hrs, but I'm losing large amount of cells.
I collected the lysis after stimulation and 2x wash with PBS, and I got around 50ug of protein per 90mm dish...
I was expecting a lot more...I think I lost too many cells during starvation and stimulation.

I heard HUVECs are hard to serum starve, I was wondering if anyone has any advice on this problem?

Thanks in advance for your help!

#2 EnglishBreakfast

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Posted 04 December 2013 - 03:13 PM

I've gotten my HUVECs from ATCC and I grow them in Vascular basal cell medium supplemented with the endothelial growth kit-BBE (bovine brain extract), in 2% FBS. I tried starving them with just the basal medium, no FBS, no kit but they basically die on my just after 4 hours. Even with FBS, they're not happy if they don't get the BBE kit.

 

WhenI starve them using 0.1% FBS, fully supplemented medium overnight, they look heathy and relatively fine the next day. Hope it helps.






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