April 19, 2022

What do I need to know when extracting RNA from FFPE samples?

By: Jacqueline Steenhuis

Avoid methods based on organic solvents (Trizol)
Avoid harsh chaotropic salts (i.e. guanidinium)
Avoid detergents that impact downstream quantitation by UV and/or Qubit (e.g. Triton X-100)
Do not rely on RIN to quantitate the integrity of an FFPE-derived sample, use DV200 instead.
Use a kit or method that removes chemical modifications from formalin. Do not raise the temperature to 80?C or above. Even short times at this temperature will significantly lower integrity.
Be wary of Qubit and Nanodrop concentrations because of the possibility of contamination by organic molecules or DNA.
Use qPCR to quantitate your RNA, and always look carefully at melt curves to determine whether nonspecific amplification may have occurred.