SLOW cell growth - (Dec/03/2007 )
I have plated early passage smooth muscle cells from a young and old monkey aorta (thoracic and abdominal) in plastic flasks. The old cells are growing slowly, while the young show little to no growth. I have had to wait in excess of one month for a flask, seeded at roughly 35% confluency, to reach 100%. I have not yet tried plating the cells on another substance, i.e. collagen, but I find it hard to imagine that it will completely remedy this problem. I am using DMEM supplemented with 10% FBS. Any thoughts?
You may need specialized medium, coating etc. Check & see if they have anything that can be helpful. www.lifelinecelltech.com.
How are you isolating your cells ?
If you are using explants cultures then your cell should "walk off" your smooth muscle explants within 7-10 days. We have in our group tried to culture SMC's from human vein. From young donors, cells grow very well (i.e. <30 years of age). Older tissue results in very poor cell growth i.e. not enough for experimentation.
Enzyme dissociation of tissue gives rise to too many contaminating cells.....so we do not use this method.
You do not need to use collagen, Poly-L-Lysine, Fibronectin etc etc etc to aid cell attachment.
What you do need to do is use QUALITY SERUM......... my question to you is WHICH IS THE BEST QUALITY SERUM IN THE WORLD ?
What plastic are you using... there is MASSIVE differences in the TC plasticware.....+ charges/negative charges etc....you really need to optimise what platics to use.
Primary SMC's once isolated should be CHARACTERISED and then MYCOPLASMA TESTED...if you want to publish your findings.
I do hope this is useful.