Samples homogenized in a phenol–guanidine reagent such as TRIzol or TriPure are commonly held at 4 °C overnight, and frozen for longer storage. The reagent inactivates RNases in lysed material, so a homogenate is far more stable than fresh tissue — but it is not a permanent storage medium, and exact limits differ between products. Always check your reagent’s manual; the points below are general good practice.
Safe pause points during a phenol–guanidine extraction
- After homogenization: the best place to pause. Short-term at 4 °C (overnight), or freeze at −20 °C/−80 °C for longer.
- After phase separation: do not leave the aqueous phase standing for long; move on to precipitation.
- After precipitation and washing: the RNA pellet can be kept under 75% ethanol at −20 °C or −80 °C until you dissolve it.
- Purified RNA: store in RNase-free buffer or water at −80 °C, in aliquots.
Tips that prevent losses
- Homogenize completely before storage; intact tissue chunks in reagent are not protected throughout.
- Use tubes with secure caps and keep phenol-containing samples in a fume-hood-appropriate workflow.
- Thaw frozen homogenates at room temperature, mix, and proceed directly — avoid repeated freeze–thaw.
- Expect some loss of yield or integrity with long holds; run a gel on a pilot sample after storage.
Phenol-free alternatives
If you would rather avoid phenol and chloroform, column-based kits such as the CYRN03 RNApure Total RNA Kit (a TriPure isolation reagent alternative) give comparable total RNA with a simpler workflow.
Common questions
Can I leave samples in TRIzol or TriPure at 4 °C?
Short-term, yes: homogenized samples in phenol–guanidine reagent are commonly held at 4 °C overnight. For longer storage, freeze them. Always check the storage limits in your reagent’s own manual.
How long can samples be stored in TRIzol at −80 °C?
Homogenates in phenol–guanidine reagent are routinely stored frozen for weeks to months. Follow the manufacturer’s stated limit for your reagent and avoid repeated freeze–thaw.
Should I homogenize before storing in TRIzol?
Yes. Store homogenized lysate, not intact tissue sitting in reagent; the reagent inactivates RNases only where it has contacted lysed cells.
Can I stop after phase separation and store the aqueous phase?
It is better to precipitate RNA from the aqueous phase promptly, or store the precipitated RNA pellet in 75% ethanol at −20 °C/−80 °C, than to leave the aqueous phase standing.
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