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DNA isolated from formalin-fixed paraffin-embedded healthy tissue after 30 years of storage can be used for forensic studies

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Abstract

Tissue formalin fixation and paraffin embedding (FFPE) is a standard method for long-term preservation and morphological and molecular analysis. The aim of this study was to analyze the effect of storage time on the integrity of DNA isolated from three different healthy FFPE tissues. DNA was isolated from FFPE heart, liver and brain tissues obtained from autopsy and archived from 1988 to 2017 using two different methods of DNA isolation: phenol–chloroform-isoamyl alcohol (PCI) and PureLink Genomic DNA Kit. The quantification and purity of DNA was measured spectrophotometrically at 260 nm and 280 nm. The quality of isolated DNA was evaluated by PCR amplification of GPD1 (150 bp), ACTB (262 bp) and RPL4 (407 bp) genes. The histomorphological characteristics of FFPE tissues were not significantly changed during 30 years of storage. Higher yield (272.9 ± 10.3 µg) and purity (A260/280 = 2.05) of DNA was obtained using the PCI method for DNA isolation from FFPE liver tissue. The PCI extraction method showed reproducible and consistent results in PCR amplification of all of three examined genes. The GPD1 gene can be amplified up to 30 years, the ACTB gene in the same samples up to 26 years and the RPL4 gene up to 6 years of storage in FFPE blocks. Although the best yield and purity of isolated DNA (using both isolation methods) was obtained from FFPE liver tissue, the DNA with the most preserved integrity was obtained from brain tissue archived up to 30 years. This is the first report using long-term archived healthy FFPE tissues (up to 30 years) that shows that the DNA isolated from these tissues is of preserved integrity and can be used in molecular autopsy.

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Funding

This study was supported by grant of Faculty of Medical Sciences, University of Kragujevac, Serbia (grand number JP: 05/13).

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Authors and Affiliations

  1. Faculty of Medical Sciences, Department of Forensic Medicine, University of Kragujevac, Kragujevac, Serbia

    Katarina Vitošević, Miloš Todorović & Živana Slović

  2. Institute of Forensic Medicine, Faculty of Medicine, University of Belgrade, Belgrade, Serbia

    Tatjana Varljen

  3. Institute of Pathology and Forensic Medicine, Military Medical Academy, Belgrade, Serbia

    Stevan Matić

  4. Faculty of Medical Sciences, Department of Genetics, University of Kragujevac, Kragujevac, Serbia

    Danijela Todorović

Authors
  1. Katarina Vitošević
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  2. Miloš Todorović
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  4. Tatjana Varljen
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  5. Stevan Matić
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Correspondence to Danijela Todorović.

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The authors declare that they have no conflict of interest.

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Institutional ethical approval was obtained in compliance with the Declaration of Helsinki. Authorization to use human biological samples for research was obtained. The Ethics Committee of the University of Kragujevac, Faculty of Medical Sciences, the Ethic Committee of Clinical Center of Kragujevac (no.01–2798), Appeal Public Prosecutor's Office from Kragujevac (A number 79/13) and Higher Court in Kragujevac (SU-VIII-110/13) agreed to these investigations.

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Vitošević, K., Todorović, M., Slović, Ž. et al. DNA isolated from formalin-fixed paraffin-embedded healthy tissue after 30 years of storage can be used for forensic studies. Forensic Sci Med Pathol 17, 47–57 (2021). https://doi.org/10.1007/s12024-020-00327-z

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