title: Nucleic acids from long-term preserved FFPE tissues are suitable for downstream analyses.
abstract: Tissues used for clinical diagnostics are mostly formalin-fixed and paraffin-embedded (FFPE) which provides many advantages. However, the quality of the obtained nucleic acids (NA) is reduced and this turns out to be a challenge for further molecular analyses. Although the spectrum of analyses of NA extracted from FFPE tissue has increased, the standard operating procedures for NA isolation from old tissue blocks still need to be improved. Here, we compared the efficiency of different NA extraction methods, using FFPE tissues of variable age and origin, with respect to downstream analyses. Our study showed that the phenol-chloroform isoamyl alcohol (PCI) and the commercial Qiagen protocol yielded samples with highest purity. The PCI protocol delivered the longest amplicons even from samples from the 1970s. We developed a short (1 h) tissue lysis procedure that turned out to be highly time- and cost-effective when DNA quality was tested using single and multiplex PCR. Compared to a 1-day lysis-protocol, the amplicons were only 100 bp shorter. In addition, single-copy genes used in daily routine were successfully amplified from long-term stored FFPE samples following 1-h tissue-lysis. The RNA integrity numbers (RIN) determined on RNA isolated from FFPE tissues indicated degraded RNA; however, all RINs were above the generally agreed threshold of 1.4. We showed that, depending on the purpose of the analysis, NA retrieved from FFPE tissues older than 40 years may be successfully used for molecular analysis.
authors.affiliation: Institute of Pathology, Helmholtz Zentrum München, German Research Center for Environmental Health, Ingolstaedter Landstrasse 1, 85764 Neuherberg, Germany.
authors.forename: Natalie
authors.initials: N
authors.lastname: Ludyga
authors.forename: Barbara
authors.initials: B
authors.lastname: Grünwald
authors.forename: Omid
authors.initials: O
authors.lastname: Azimzadeh
authors.forename: Sonja
authors.initials: S
authors.lastname: Englert
authors.forename: Heinz
authors.initials: H
authors.lastname: Höfler
authors.forename: Soile
authors.initials: S
authors.lastname: Tapio
authors.forename: Michaela
authors.initials: M
authors.lastname: Aubele
mesh_terms: D005404:Fixatives; D005557:Formaldehyde; D006801:Humans; D009696:Nucleic Acids / Q000032:analysis / Q000302:isolation & purification*; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction / Q000379:methods*; D013997:Time Factors; D016707:Tissue Fixation
chemical_list: D005404:Fixatives; D009696:Nucleic Acids; D005557:Formaldehyde