I am trying to recover very precious FACS samples (approx 8000 cells) that was stored in 2 mL of RNAlater. I am unable to pellet the tissue. I am really hoping there is a way to save the samples! Your advice is very much appreciated!
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Extracting precious tissue from too large of volumes of RNAlater
Started by dbratt, Dec 01 2009 01:24 PM
5 replies to this topic
#2
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an elder
Posted 02 December 2009 - 08:51 AM
have you tried diluting the sample so that the solution is less dense?
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#3
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Posted 02 December 2009 - 11:00 AM
With only about 8000 cells, you may be sedimenting the cells, but there isn't enough there to form a visible pellet. What do you want to use the cells for now?
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#4
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Posted 02 December 2009 - 11:55 AM
This sounds like a job for a microcon filter (Millipore). Although these are mostly for dialysis replacement protein purification, I think there is one with a 0.2 micron cutoff, which would be just the thing for this problem. You invert and spin the filtered product out.
#5
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Somewhere I belong
Posted 02 December 2009 - 02:16 PM
hi dbratt
Cells normally dont pellet effectively in RNAlater. The best way is to just add equal volume of PBS to the solution and spin it...I normally do a 9k rpm for 3 minutes...
Its better to take a maximum of 1 ml per tube during centrifugation, else you end up with some cells over the wall which would be disturbed when pipetting the solution...Just to make sure, keep the supernatent and spin once more
Hope this helps
Cells normally dont pellet effectively in RNAlater. The best way is to just add equal volume of PBS to the solution and spin it...I normally do a 9k rpm for 3 minutes...
Its better to take a maximum of 1 ml per tube during centrifugation, else you end up with some cells over the wall which would be disturbed when pipetting the solution...Just to make sure, keep the supernatent and spin once more
Hope this helps
#6
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Posted 08 December 2009 - 12:28 PM
gfischer, on Dec 2 2009, 11:00 AM, said:
With only about 8000 cells, you may be sedimenting the cells, but there isn't enough there to form a visible pellet. What do you want to use the cells for now?
HI Gfischer.....
the ultimate goal is to use for microarray!
