title: DNA extraction and molecular analysis of non-tumoral liver, spleen, and brain from autopsy samples: the effect of formalin fixation and paraffin embedding.
abstract: The use of molecular biology in combination with morphological analysis is increasing because of the treatments by target therapies. However, to improve the methods for obtaining DNA for molecular analyses from formalin-fixed, paraffin-embedded (FFPE) tissue is a challenge. The aim of this study was to evaluate the DNA extracted from FFPE tissue blocks (non-tumoral liver, spleen, and brain), obtained from autopsy, 8-24 h post mortem, using three methods of DNA extraction. PCR of the β-actin (136 pb) and human amelogenin (AMEL 212-218 bp/106-112 bp) genes, as well as short tandem repeat (STR) (100-400 bp fragments), reported in forensic scientific analysis, was performed to evaluate the effectiveness of the methods of DNA extraction. We used 28 archived (1 and 5 years) and 12 recent autopsy cases. The commercial kit showed reproducible and consistent results in the PCR amplification of the β-actin and AMEL genes and in analysis by STR used in forensic analysis. This is the first report using non-tumoral samples from FFPE autopsy tissues, comparing the three most common methods of DNA extraction and using the STR previously described in forensics. Our study has clarified the challenges for pathologists in applying the molecular biology approach in combination with methods suited for morphology, which must be improved. The data provided here should be used in other molecular studies in FFPE samples.
authors.affiliation: Department of Pathology - Escola Paulista de Medicina, Universidade Federal de São Paulo - EPM/UNIFESP, São Paulo, SP, Brazil.
authors.forename: Karina Silva
authors.initials: KS
authors.lastname: Funabashi
authors.forename: Denise
authors.initials: D
authors.lastname: Barcelos
authors.forename: Iria
authors.initials: I
authors.lastname: Visoná
authors.forename: Marcelo Souza
authors.initials: MS
authors.lastname: e Silva
authors.forename: Maria Luiza Almeida Prado Oliveira
authors.initials: ML
authors.lastname: e Sousa
authors.forename: Marcello Fabiano
authors.initials: MF
authors.lastname: de Franco
authors.forename: Edna Sadayo Miazato
authors.initials: ES
authors.lastname: Iwamura
mesh_terms: D000199:Actins / Q000235:genetics; D053523:Amelogenin / Q000235:genetics; D001344:Autopsy; D001923:Brain Chemistry; D004247:DNA / Q000302:isolation & purification*; D005260:Female; D005404:Fixatives / Q000009:adverse effects; D049429:Forensic Pathology; D005557:Formaldehyde / Q000009:adverse effects; D005821:Genetic Techniques; D006801:Humans; D008099:Liver / Q000737:chemistry*; D008297:Male; D018895:Microsatellite Repeats; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction; D011933:Reagent Kits, Diagnostic; D015203:Reproducibility of Results; D012732:Sex Determination Analysis; D013154:Spleen / Q000737:chemistry*; D013997:Time Factors; D016707:Tissue Fixation / Q000379:methods*
chemical_list: C507199:AMELX protein, human; D000199:Actins; D053523:Amelogenin; D005404:Fixatives; D011933:Reagent Kits, Diagnostic; D005557:Formaldehyde; D004247:DNA