could someone advice me on how to coat my tissue culture plates with fibronectin? the negative and positive controls? and do i have to culture my cells in serum free medium in this condition?
Thanx
Edited by youyou, 20 April 2009 - 06:16 AM.
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#1
youyou
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Posted 20 April 2009 - 06:14 AM
Hi everyone
could someone advice me on how to coat my tissue culture plates with fibronectin? the negative and positive controls? and do i have to culture my cells in serum free medium in this condition?
Thanx
could someone advice me on how to coat my tissue culture plates with fibronectin? the negative and positive controls? and do i have to culture my cells in serum free medium in this condition?
Thanx
#2
aimikins
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Posted 20 April 2009 - 02:40 PM
I can't answer the SFM, but as for coating I used to do that pretty often.
make fibronectin at 1mg/mL in PBS. it will be clumpy, so assume your 'real' concentration is a bit lower. this is OK, because mostly you are just using an excess amount and you'll get good coverage.
vortex the holy living daylights out of the tube.
pipet an appropriate volume into your well/dish/whatever.
minimum time is 1 hour at 37C. I like to leave it overnight or 1/2 day if it's a critical step. this is the method we used when studying skin keratinocytes at my last job.
make fibronectin at 1mg/mL in PBS. it will be clumpy, so assume your 'real' concentration is a bit lower. this is OK, because mostly you are just using an excess amount and you'll get good coverage.
vortex the holy living daylights out of the tube.
pipet an appropriate volume into your well/dish/whatever.
minimum time is 1 hour at 37C. I like to leave it overnight or 1/2 day if it's a critical step. this is the method we used when studying skin keratinocytes at my last job.
"it is a miracle that curiosity survives formal education" -A.E.
