title: Utilizing mRNA extracted from small, archival formalin-fixed paraffin-embedded prostate samples for translational research: assessment of the effect of increasing sample age and storage temperature.
abstract: BACKGROUND
  Translational prostate cancer research is hampered by long intervals from diagnosis to patient progression and difficulty in obtaining cancer tissue for investigation. As such, it is imperative to utilise aging formalin-fixed paraffin-embedded (FFPE) tissue samples from the pathology archive with linked patient outcome data to allow current day research. This study aimed to assess the adequacy and quantity of mRNA extracted from formalin fixed paraffin embedded (FFPE) prostate tissue, including prostate biopsies, up to 15 years old. The decay of mRNA over time and under differing storage conditions was also assessed.
  
  
  METHODS
  Archived FFPE benign prostatic tissue up to 15 years old from transurethral resection of the prostate (TURP) and transrectal ultrasound guided (TRUS) biopsies as well as fresh tissue obtained from patients undergoing TURP for benign bladder outlet obstruction were used. Following mRNA extraction beta-actin real-time PCR was carried out using a set of 4 different primer/probes to assess mRNA quality and quantity.
  
  
  RESULTS
  There was no difference in mRNA quantity/quality extracted from "fresh" FFPE tissue from the same patient over a 4-month period following surgery. The temperature of block storage did not alter quality/quantity of the mRNA (P > 0.05, unpaired t test). Fresh tissue had a higher quality/quantity, indicated by a lower C                            (                               T                            )  value, than FFPE samples from the same patient (P ≤ 0.03, one-way ANOVA). Despite being up to 15 years old, all archived FFPE TURP and TRUS biopsy samples had "high" or "very high" levels of expression making them suitable for further analysis. However, the quality of the mRNA in archived FFPE samples did significantly decline with increasing sample age.
  
  
  CONCLUSIONS
  It is possible to extract mRNA of sufficient standard for further transcriptomic analysis from minute FFPE samples up to 15 years old. This work adds to the literature suggesting that exploitation of retrospective prostate tissue collections with robust associated clinical data is possible.
authors: Division of Pathology, Institute of Genetics and Molecular Medicine, University of Edinburgh, Western General Hospital, Crewe Road South, Edinburgh EH4 2XU, UK. grant.stewart@ed.ac.uk
authors: Grant D
authors: GD
authors: Stewart
authors: Jan
authors: J
authors: Baird
authors: Frances
authors: F
authors: Rae
authors: Jyoti
authors: J
authors: Nanda
authors: Antony C P
authors: AC
authors: Riddick
authors: David J
authors: DJ
authors: Harrison
mesh_terms: D000704:Analysis of Variance; D001706:Biopsy; D005557:Formaldehyde; D006801:Humans; D008297:Male; D016612:Paraffin Embedding; D011467:Prostate / Q000737:chemistry*; D011470:Prostatic Hyperplasia / Q000473:pathology* / Q000601:surgery; D012333:RNA, Messenger / Q000032:analysis*; D060888:Real-Time Polymerase Chain Reaction; D013048:Specimen Handling; D013696:Temperature; D013997:Time Factors
chemical_list: D012333:RNA, Messenger; D005557:Formaldehyde