Abstract
Urinary cell-free DNA (cfDNA) is an attractive body fluid for liquid biopsy. In this study, we compared the efficiencies of four commercial kits for urinary cell-free DNA (cfDNA) isolation and of various sample storage conditions. Urinary cfDNA was isolated from 10 healthy individuals using four commercial kits: QIAamp Circulating Nucleic Acid Kit (QC; Qiagen), MagMAX™ Cell-Free DNA Isolation Kit (MM; Applied Biosystems), Urine Cell-Free Circulating DNA Purification Midi Kit (NU; Norgen Biotek), and Quick-DNA™ Urine Kit (ZQ; Zymo Research). To assess the isolation efficiency, an Agilent 2100 Bioanalyzer with High Sensitivity DNA chips was used, and cfDNA yield was defined as the amount of cfDNA obtained from 1 mL of urine. MM and QC provided the highest cfDNA yield in the 50–300 bp range, and MM and NU gave the highest cfDNA yield in the 50–100 bp range. In particular, the NU kit was efficient for isolation of more fragmented cfDNA in the range of 50–100 bp with the lowest cellular genomic DNA contamination. ZQ had the best cost-efficiency for isolating the same amount of urinary cfDNA. Samples stored at −70 °C with the addition of 10 mM EDTA resulted in the highest cfDNA yield 3 months after sample collection.
Highlights
Liquid biopsy has emerged as an important non-invasive tool for cancer diagnostics
Liquid biopsy utilizes genetic material from body fluids; this method can overcome the limitations associated with traditional tissue biopsy such as invasiveness and the difficulty of repeated sampling [1,2,3]
Many researchers have investigated the optimal conditions for each step, encompassing sample collection, handling, and storage to maximize the recovery of cell-free DNA (cfDNA) [7,8,9,10]
Summary
Liquid biopsy has emerged as an important non-invasive tool for cancer diagnostics. Liquid biopsy utilizes genetic material from body fluids; this method can overcome the limitations associated with traditional tissue biopsy such as invasiveness and the difficulty of repeated sampling [1,2,3].Cell-free DNA (cfDNA) is one of the most important sources for liquid biopsy. Liquid biopsy has emerged as an important non-invasive tool for cancer diagnostics. Liquid biopsy utilizes genetic material from body fluids; this method can overcome the limitations associated with traditional tissue biopsy such as invasiveness and the difficulty of repeated sampling [1,2,3]. Cell-free DNA (cfDNA) is one of the most important sources for liquid biopsy. Many researchers have examined genetic alteration of cfDNA using various analytical tools such as real-time PCR, droplet digital PCR (ddPCR), and next-generation sequencing (NGS) [4,5,6]. To achieve proper results with various genetic tests, it is important to establish a method to extract a high yield of cfDNA. Many researchers have investigated the optimal conditions for each step, encompassing sample collection, handling, and storage to maximize the recovery of cfDNA [7,8,9,10]
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