Hi all,
Has anyone frozen the murine tissue samples (lung or spleen) in -80 and then later thawed these to go ahead with cell purification (PMNs etc) for RNA extraction? I am wondering if it works or it kills the cells? I am trying to optimize the stuff for RNA-sequencing and was wondering if on day 1, I can freeze the samples and next day go for cell isolation and RNA purification?
Any idea, inputs or protocol for doing so will be helpful.
Thanks!
Hi! Here we first do the homogenization of the tissue with appropriate buffer and then flash freeze it in liquid nitrogen before storing it at -80. Freezing the tissue prior to homogenization could lead to lower quality RNA (the longer it is freezed the lower will be the quality of your RNA).
Regards
- Agree with above
- Flash freeze in Liquid nitrogen before storage @ -80C to maximise your chances of purifying undegraded RNA
- You can freeze directly @ -80C but that will not freeze the innder core of tissue quick enough to ensure all of the RNA remains in tact
- Hence flash freezing first
freezing at -80C works well; RNA Later reagent is a great alternative- widely used for RNA preservation in tissue and other sample types
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