Tissue homogenization buffer to keep viral RNA intact
Posted 17 November 2010 - 06:35 AM
I would like to prepare some spleen samples for later RNA isolation. In the meantime I was planning to store all homogenates at -20 C. Can I use just PBS (RNAse free) as homogenization buffer (Homogenizer: Mikro-Dismembrator S, Sartorius)and keep RNA intact? After I collect enough samples, I intend to extract viral RNA from supernatant of centifuged homogenates by using QIAamp viral RNA mini kit for isolation of viral RNA from cell-free body fluids (only available in my lab at this time). I will treat that supernatant as it is a body fluid, but I'm wondering if this protocol may work?
Please help me with some advice!
Posted 18 November 2010 - 04:17 PM
well, if i were you i would homogenize my tissue samples and centrifuge them once received, and then store the supernatant at -80C.
PBS looks a fine choice as a storage buffer, but as a homogenizer am not sure ... sorry for this
see the attached link : Top Ten Ways To Improve Your RNA Analysis Experiment - invitrogen
QIAamp viral RNA mini kit works fine too ...
read the kit insert carefully concerning tissue homogenates, if it includes any specific tips on this ...
for example : homogenizing buffer / storage buffer choice ...
and to check whether to treat your samples as body fluids ( supernatant ) or if there is any mentioning about tissue homogenates specifically.
All The Good Luck.
Posted 18 November 2010 - 04:49 PM
Posted 02 December 2010 - 09:53 PM
Posted 13 December 2010 - 05:57 AM
Thank you for the information!
Can I mix that LyseNow reagent from Metammune with PBS and use it for tissue homogenization, or even better, can I prepare a hommade equivalent? Thank you in advance!
Posted 17 December 2010 - 02:24 AM
4M guanidine thiocyanate
25mM sodium citrate (pH7)
dissolved in PBS
Posted 17 December 2010 - 05:20 AM