Jump to content

  • Log in with Facebook Log in with Twitter Log in with Windows Live Log In with Google      Sign In   
  • Create Account

Submit your paper to J Biol Methods today!
Photo
- - - - -

Extracting precious tissue from too large of volumes of RNAlater


  • Please log in to reply
5 replies to this topic

#1 dbratt

dbratt

    member

  • Members
  • Pip
  • 2 posts
0
Neutral

Posted 01 December 2009 - 01:24 PM

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!

#2 mdfenko

mdfenko

    an elder

  • Active Members
  • PipPipPipPipPipPipPipPipPipPip
  • 2,735 posts
125
Excellent

Posted 02 December 2009 - 08:51 AM

have you tried diluting the sample so that the solution is less dense?
talent does what it can
genius does what it must
i do what i get paid to do

#3 gfischer

gfischer

    Veteran

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 195 posts
9
Neutral

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?
Above all things, if kindness is your king,
then heaven will be yours, before you meet your end

#4 phage434

phage434

    Veteran

  • Global Moderators
  • PipPipPipPipPipPipPipPipPipPip
  • 2,410 posts
234
Excellent

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 gogreen

gogreen

    Somewhere I belong

  • Active Members
  • PipPipPipPipPip
  • 90 posts
0
Neutral

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 :lol:

Hope this helps

#6 dbratt

dbratt

    member

  • Members
  • Pip
  • 2 posts
0
Neutral

Posted 08 December 2009 - 12:28 PM

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!




Home - About - Terms of Service - Privacy - Contact Us

©1999-2013 Protocol Online, All rights reserved.