Please find below a list of key points to consider to maximize your RNA extraction:
Remove as much paraffin as possible around the tissue.
If you have a tissue with a small surface area, you can consider reducing the amount of FFPE Lysis Buffer (down to 70μl). Please see How can I increase my final yield? for more information.
Check that the sample is well submerged in the Lysis Buffer.
The Removal Solution is light sensitive and provided in a brown bottle. Please follow carefully the storage and handling recommendation described in the Userguide.
Ensure Spin Column is prepared appropriately. Please vortex well and spin down prior to removing Storage Buffer, and ensure no air bubbles are present. Please see the following FAQ Do you have any specific recommendations for spin column preparation? for additional tips on column preparation.
Prior to DNase digestion, check for the presence of paraffin in the upper phase when letting the sample cools down. If you observe a jelly layer on top of the aqueous phase, this means that you have some paraffin left in your sample. In this case, please re-incubate the sample at 60°C and centrifuge again.
Increase the centrifugation speed to maximize the phase separation.
When centrifuging the final eluate, do not increase the speed. If >1,000 × g is used, this can lead to impurities in your final sample.