title: Comparison of Automated and Manual DNA Isolation Methods for DNA Methylation Analysis of Biopsy, Fresh Frozen, and Formalin-Fixed, Paraffin-Embedded Colorectal Cancer Samples.
abstract: Automated DNA isolation can decrease hands-on time in routine pathology. Our aim was to apply automated DNA isolation and perform DNA methylation analyses. DNA isolation was performed manually from fresh frozen (CRC = 10, normal = 10) specimens and colonic biopsies (CRC = 10, healthy = 10) with QIAamp DNA Mini Kit and from FFPE blocks (CRC = 10, normal = 10) with QIAamp DNA FFPET Kit. Automated DNA isolation was performed with MagNA Pure DNA and Viral NA SV kit on MagNA Pure 96 system. DNA methylation of MAL, SFRP1, and SFRP2 were analyzed with methylation-specific high-resolution melting analysis. Yield of automatically isolated samples was equal in fresh frozens and significantly lower compared to manually isolated biopsy and FFPE samples. OD260/280 of fresh frozen and biopsy samples were similar after both isolations, automated isolation resulted in lower purity in FFPE samples. Both protocols resulted in similar OD260/230 from fresh frozens, automated isolation method was superior in biopsies and manual protocol in FFPE samples. DNA methylation of biopsies, fresh frozen samples were highly similar after both methods, results of automatically and manually isolated FFPE samples were different. Automated DNA isolation from fresh frozen samples can be suitable for high-throughput laboratories.
authors.affiliation: 2nd Department of Internal Medicine, Semmelweis University, Budapest, Hungary Molecular Medicine Research Unit, Hungarian Academy of Sciences, Budapest, Hungary alexandra.kalmar@gmail.com.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Kalmár
authors.affiliation: 2nd Department of Internal Medicine, Semmelweis University, Budapest, Hungary Molecular Medicine Research Unit, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Bálint
authors.initials: B
authors.lastname: Péterfia
authors.affiliation: Molecular Medicine Research Unit, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Barnabás
authors.initials: B
authors.lastname: Wichmann
authors.affiliation: 2nd Department of Internal Medicine, Semmelweis University, Budapest, Hungary.
authors.forename: Árpád V
authors.initials: ÁV
authors.lastname: Patai
authors.affiliation: 2nd Department of Internal Medicine, Semmelweis University, Budapest, Hungary.
authors.forename: Barbara K
authors.initials: BK
authors.lastname: Barták
authors.affiliation: 2nd Department of Internal Medicine, Semmelweis University, Budapest, Hungary.
authors.forename: Zsófia B
authors.initials: ZB
authors.lastname: Nagy
authors.affiliation: 2nd Department of Internal Medicine, Semmelweis University, Budapest, Hungary.
authors.forename: István
authors.initials: I
authors.lastname: Furi
authors.affiliation: 2nd Department of Internal Medicine, Semmelweis University, Budapest, Hungary Molecular Medicine Research Unit, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Zsolt
authors.initials: Z
authors.lastname: Tulassay
authors.affiliation: 2nd Department of Internal Medicine, Semmelweis University, Budapest, Hungary Molecular Medicine Research Unit, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Béla
authors.initials: B
authors.lastname: Molnár
mesh_terms: D057205:Automation, Laboratory / Q000379:methods*; D001706:Biopsy; D015179:Colorectal Neoplasms / Q000175:diagnosis* / Q000473:pathology; D004247:DNA / Q000737:chemistry / Q000302:isolation & purification*; D019175:DNA Methylation; D006801:Humans; D010340:Pathology, Surgical / Q000379:methods; D013048:Specimen Handling / Q000379:methods*
chemical_list: D004247:DNA
keywords: DNA methylation; automated genomic DNA isolation; colorectal cancer; high-throughput screening