Do formalin fixed paraffin-embedded tissues (FFPEt) represent a suitable source of nucleic acids for bio-molecular retrospective studies in veterinary oncology?

Purpose Formalin-fixed, paraffin-embedded tissues (FFPEt) represent a unique source of archived biological material. Today, it has become desirable to utilize canine tumour FFPEt as a source of nucleic acids for retrospective–omic investigations useful to correlate the patient transcriptome with treatment response and clinical outcome. Aim of the present study was the optimization of a protocol for nucleic acids extraction from canine tumour FFPEt. Methods Archived canine mast cell tumour FFPEt, with different formalin fixation times, were used to compare (a) performances of three and five different commercial DNA and RNA extraction kits, respectively; (b) the effect of formalin fixation times (from 24 hrs up to 8 days); (c) samples stored in RNAlater solution with the corresponding FFPEt. DNA and RNA concentration and quality were evaluated by using the Nanodrop spectrophotometer and capillary electrophoresis (Bionalyser), respectively. Real Time RT-PCR assays for two housekeeping genes (GOLGA1 and CGI-119), Ki67 and c-KIT, together with PCR for c-KIT exons 8, 9, 11 and 17 were used as end points. Results Kits for nucleic acid extraction showed remarkable differences in terms of nucleic acid yield and purity. Altogether, DNA and RNA (mostly) were for the most part degraded, even at lower formalin fixation times. Compared to fresh tissue, FFPEt showed a lower amplification efficiency, when present. Conclusions Present data suggest that archival FFPEt is not suitable for retrospective bio-molecular investigations. Accordingly, more standardized sampling and fixation procedures are needed to guarantee the preservation of nucleic acids. Study supported by grant from RC IZSVE 10/09.