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Isolation of Cell-Free DNA from Maternal Plasma

Protocol
First Online:
Part of the Methods in Molecular Biology book series (MIMB, volume 1885)

Abstract

Noninvasive prenatal genetic tests analyzing the cell-free fetal DNA in the circulation of expectant mothers are now performed routinely in clinical diagnostic laboratories. Leveraging the power of next generation sequencing (NGS), these tests can detect variation in chromosomal copy number or microdeletions early in gestation. All methods begin with blood collection followed by transport to the diagnostic lab, plasma separation, and purification of ccfDNA from the plasma to prepare it for molecular analysis. Preservation of ccfDNA in blood samples and highly efficient purification from plasma are paramount since the quality and quantity of target nucleic acids determine the sensitivity and therefore success of these assays. Maximizing quality and quantity and minimizing variation in extraction yield pose significant challenges for diagnostic labs, many of which use manual isolation methods for plasma volumes greater than 5 mL. One way to reduce variability is to automate the extraction processes and, to the extent possible, minimize hand-on operations. This chapter details two procedures for isolating ccfDNA from 10 mL plasma by manual and automated means using the QIAamp Circulating Nucleic Acid Kit and the QIAsymphony Circulating DNA Kit. The ccfDNA recovered is suitable for downstream processing in noninvasive prenatal tests for aneuploidy detection.

Key words

Circulating cell-free DNA Automation Fetal Noninvasive prenatal testing QIAsymphony QIAamp Blood sample stabilization Circulating DNA extraction SNP-based aneuploidy detection 
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Notes

Disclaimer

The scalable ccfDNA isolation methods presented here are for performance evaluations only.

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© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Natera Inc.San CarlosUSA

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