Evaluation of commercial kits for purification of circulating free DNA

  • Russell J. Diefenbach
    Affiliations
    Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Macquarie University, Sydney, NSW 2109, Australia

    The Poche Centre, Melanoma Institute Australia, NSW 2065, Australia
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  • Jenny H. Lee
    Affiliations
    Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Macquarie University, Sydney, NSW 2109, Australia

    The Poche Centre, Melanoma Institute Australia, NSW 2065, Australia
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  • Richard F. Kefford
    Affiliations
    Department of Clinical Medicine, Faculty of Medicine and Health Sciences, Macquarie University, Sydney, NSW 2109, Australia

    The Poche Centre, Melanoma Institute Australia, NSW 2065, Australia

    Department of Medical Oncology, Crown Princess Mary Cancer Centre, Westmead and Blacktown Hospitals, Sydney, NSW 2145, Australia
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  • Helen Rizos
    Correspondence
    Corresponding author at: Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Macquarie University, Sydney, NSW 2109, Australia.
    Affiliations
    Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Macquarie University, Sydney, NSW 2109, Australia

    The Poche Centre, Melanoma Institute Australia, NSW 2065, Australia
    Search for articles by this author

      Highlights

      • Spin column-based kits outperform magnetic bead-based commercial cfDNA kits.
      • The Qiagen spin column-based cfDNA kit remains the gold standard.
      • The Qiagen magnetic bead-based cfDNA kit is an alternative when factoring in price.

      Abstract

      Analysis of liquid biopsies and the identification of non-invasive biomarkers for the diagnosis and prognosis of solid tumors has grown exponentially over the last few years. This has led to an increasing number of commercial kits optimised for the purification of circulating free (cf) DNA and RNA/miRNA from biofluids such as plasma, serum and urine. To optimise and standardise current practices we sought to evaluate the performance of spin column-based and magnetic bead-based commercial kits. The following commercial cfDNA purification kits were analysed in this study: QIAamp circulating nucleic acid kit (Qiagen, Germany); Plasma/serum cell-free circulating DNA Purification midi kit (Norgen Biotek, Canada); QIAamp minElute ccfDNA mini kit (Qiagen); Maxwell RSC ccfDNA plasma kit (Promega, USA); MagMAX cell-free DNA isolation kit (Applied Biosystems, USA); and NextPrep-Mag cfDNA isolation kit (Bioo Scientific, USA). Extracted DNA from the plasma of healthy individuals, either nonspiked or spiked with DNA fragments or cfDNA, was evaluated for recovery using either a BioRad Experion or ddPCR analysis. This study represents the first to use a comprehensive size distribution of spiked-in DNA fragments to evaluate commercial cfDNA kits. The commonly used spin column-based Qiagen QIAamp circulating nucleic acid kit was found to be the most consistent performing kit across the two evaluation assays employed. The Qiagen QIAamp minElute ccfDNA mini kit represented the best performing magnetic bead-based kit and provides an alternative based on lower cost/sample with a simpler workflow than spin column-based kits.

      Keywords

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