The emerging interest in circulating tumor DNA (ctDNA) analyses for clinical trials has necessitated the development of a high-throughput method for fast, reproducible, and efficient isolation of ctDNA. Currently, the majority of ctDNA studies use the manual QIAamp (QA) platform to isolate DNA from blood. The purpose of this study was to compare two competing automated DNA isolation platforms [Maxwell (MX) and QIAsymphony (QS)] to the current ‘gold standard’ QA to facilitate high-throughput processing of samples in prospective trials. We obtained blood samples from healthy blood donors and metastatic cancer patients for plasma isolation. Total cell-free DNA (cfDNA) quantity was assessed by TERT quantitative PCR. Recovery efficiency was investigated by quantitative PCR analysis of spiked-in synthetic plant DNA. In addition, a b-actin fragmentation assay was performed to determine the amount of contamination by genomic DNA from lysed leukocytes. ctDNA quality was assessed by digital PCR for somatic variant detection. cfDNA quantity and recovery efficiency were lowest using the MX platform, whereas QA and QS showed a comparable performance. All platforms preferentially isolated small (136 bp) DNA fragments over large (420 and 2000 bp) DNA fragments. Detection of the number variant and wild-type molecules was most comparable between QA and QS. However, there was no significant difference in variant allele frequency comparing QS and MX to QA. In summary, we show that the QS platform has comparable performance to QA, the ‘gold standard’, and outperformed the MX platform depending on the readout used. We conclude that the QS can replace the more laborious QA platform, especially when high-throughput cfDNA isolation is needed.

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Keywords automation, cell-free DNA, circulating tumor DNA, isolation
Persistent URL dx.doi.org/10.1002/1878-0261.12415, hdl.handle.net/1765/115214
Journal Molecular Oncology
Citation
van Dessel, L, Vitale, S.R, Peeters, A.M.A, Wilting, S.M, van der Vlugt-Daane, M, Oomen - de Hoop, E, … Lolkema, M.P. (2018). High-throughput isolation of circulating tumor DNA. Molecular Oncology, 13(2), 392–402. doi:10.1002/1878-0261.12415