Abstract 5701: Innovative approach for isolation of circulating cell-free DNA from liquid biopsies

• Published in: Cancer research (Chicago, Ill.) Vol. 77; no. 13_Supplement; p. 5701
• Main Authors: Malykh, Andrei, Evtushenko, Vladimir
• Format: Journal Article
• Language: English
• Published: 01.07.2017
• ISSN: 0008-5472; 1538-7445
• DOI: 10.1158/1538-7445.AM2017-5701

Abstract We developed a new approach for isolation of circulating cell-free DNA (cfDNA) from liquid biopsies (plasma or serum). Our technology is based on using the proprietary bi-functional substance X (SubX®) that binds DNA under physiological conditions (e.g. directly in biological liquids), followed by adsorption of [DNA- SubX®] complex on a solid phase matrix. Since SubX®) captures DNA via phosphate groups it allows for elimination of bias related to both AT/CG content and DNA fragments length, thus improving extraction efficacy and accuracy of downstream applications. All currently available commercial kits are [silica-chaotropic salt]-based and exhibit bias for either short or long DNA fragments, as well as for the GC content. In addition, such systems require at least 4- fold dilution of starting volume of bio liquid with concentrated (~5.0 M) guanidine thiocyanate (GTC) to keep resulted GTC concentration at about least 3.0 M. Thus, already low concentrations of cfDNA are significantly reduced and may negatively influence extraction efficacy of low abundance DNA fragments. Since our method does not require chaotropic salt for DNA capture, no dilution of starting material (and thus cfDNA) takes place. DNA-binding matrix SubX® allows either scaling the procedure to either large volumes of starting material or extra low volumes and adaptation of the protocol for 96/384 well format. Separation of cfDNA from the bulk of proteins occurs in a single vortex-spin step without employing chaotropic agents thus speeding-up isolation procedure and reducing the costs. The whole procedure can be completed within an hour and cfDNA is eluted in small volume of TE buffer. Average DNA yield from 0.2 ml of double centrifuged blood plasma from normal donors, cancer and multiple sclerosis patients was 7.5±4.2 ng (n=96). Citation Format: Andrei Malykh, Vladimir Evtushenko. Innovative approach for isolation of circulating cell-free DNA from liquid biopsies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5701. doi:10.1158/1538-7445.AM2017-5701