Evaluation of pre-analytical factors affecting plasma DNA analysis

Havell Markus, Tania Contente-Cuomo, Maria Farooq, Winnie S. Liang, Mitesh J Borad, Shivan Sivakumar, Simon Gollins, Nhan Tran, Harshil D. Dhruv, Michael E. Berens, Alan H Bryce, Aleksandar D Sekulic, Antoni Ribas, Jeffrey M. Trent, Patricia M. Lorusso, Muhammed Murtaza

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

Pre-analytical factors can significantly affect circulating cell-free DNA (cfDNA) analysis. However, there are few robust methods to rapidly assess sample quality and the impact of pre-analytical processing. To address this gap and to evaluate effects of DNA extraction methods and blood collection tubes on cfDNA yield and fragment size, we developed a multiplexed droplet digital PCR (ddPCR) assay with 5 short and 4 long amplicons targeting single copy genomic loci. Using this assay, we compared 7 cfDNA extraction kits and found cfDNA yield and fragment size vary significantly. We also compared 3 blood collection protocols using plasma samples from 23 healthy volunteers (EDTA tubes processed within 1 hour and Cell-free DNA Blood Collection Tubes processed within 24 and 72 hours) and found no significant differences in cfDNA yield, fragment size and background noise between these protocols. In 219 clinical samples, cfDNA fragments were shorter in plasma samples processed immediately after venipuncture compared to archived samples, suggesting contribution of background DNA by lysed peripheral blood cells. In summary, we have described a multiplexed ddPCR assay to assess quality of cfDNA samples prior to downstream molecular analyses and we have evaluated potential sources of pre-analytical variation in cfDNA studies.

Original languageEnglish (US)
Article number7375
JournalScientific Reports
Volume8
Issue number1
DOIs
StatePublished - Dec 1 2018

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DNA
cells
sampling
droplets
blood
assays
blood cells
volunteers
genomics
loci
methodology

ASJC Scopus subject areas

  • General

Cite this

Markus, H., Contente-Cuomo, T., Farooq, M., Liang, W. S., Borad, M. J., Sivakumar, S., ... Murtaza, M. (2018). Evaluation of pre-analytical factors affecting plasma DNA analysis. Scientific Reports, 8(1), [7375]. https://doi.org/10.1038/s41598-018-25810-0

Evaluation of pre-analytical factors affecting plasma DNA analysis. / Markus, Havell; Contente-Cuomo, Tania; Farooq, Maria; Liang, Winnie S.; Borad, Mitesh J; Sivakumar, Shivan; Gollins, Simon; Tran, Nhan; Dhruv, Harshil D.; Berens, Michael E.; Bryce, Alan H; Sekulic, Aleksandar D; Ribas, Antoni; Trent, Jeffrey M.; Lorusso, Patricia M.; Murtaza, Muhammed.

In: Scientific Reports, Vol. 8, No. 1, 7375, 01.12.2018.

Research output: Contribution to journalArticle

Markus, H, Contente-Cuomo, T, Farooq, M, Liang, WS, Borad, MJ, Sivakumar, S, Gollins, S, Tran, N, Dhruv, HD, Berens, ME, Bryce, AH, Sekulic, AD, Ribas, A, Trent, JM, Lorusso, PM & Murtaza, M 2018, 'Evaluation of pre-analytical factors affecting plasma DNA analysis', Scientific Reports, vol. 8, no. 1, 7375. https://doi.org/10.1038/s41598-018-25810-0
Markus H, Contente-Cuomo T, Farooq M, Liang WS, Borad MJ, Sivakumar S et al. Evaluation of pre-analytical factors affecting plasma DNA analysis. Scientific Reports. 2018 Dec 1;8(1). 7375. https://doi.org/10.1038/s41598-018-25810-0
Markus, Havell ; Contente-Cuomo, Tania ; Farooq, Maria ; Liang, Winnie S. ; Borad, Mitesh J ; Sivakumar, Shivan ; Gollins, Simon ; Tran, Nhan ; Dhruv, Harshil D. ; Berens, Michael E. ; Bryce, Alan H ; Sekulic, Aleksandar D ; Ribas, Antoni ; Trent, Jeffrey M. ; Lorusso, Patricia M. ; Murtaza, Muhammed. / Evaluation of pre-analytical factors affecting plasma DNA analysis. In: Scientific Reports. 2018 ; Vol. 8, No. 1.
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