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Low RNA content: sample type, sample age.
Sample preparation: Fixation process (duration, temperature) and delayed embedding can lead to sample degradation resulting to lower yields. Large tissue fragments can also lead to inhomogeneous tissue preservation and result in lower yields.
Sample storage: FFPE samples should be kept at 42-8°C or -20 to -80°C 70°C for longer storage period.
Input amount used for the RNA extraction.
Suboptimal preparation of the column.
Presence of Paraffin in the sample: We recommend to remove as much as possible surrounding paraffin in the sample before starting the extraction. If paraffin is still visible in the aqueous phase, heat again the sample to melt the paraffin and centrifuge.
Centrifugation speeds.
Sample loading on the column: it is important to slowly load the sample in the center of the matrix to not destroy or disturb the matrix. Do not touch the matrix with the pipet tip. This can damage the matrix and generated generate short-circuit current which can result in introduction of lysis components into the sample and reduce yields.