Evaluation of Different Blood Collection Tubes and Blood Storage Conditions for the Preservation and Stability of Cell-F
For the subsequent analysis of the methylated mSEPT9 colorectal cancer screening marker in plasma, different blood collection tubes and blood storage conditions were investigated. The study demonstrated that methylated Septin 9 (mSEPT9) can be consistentl
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Jurgen Distler, Reimo Tetzner, Gunter Weiss, Thomas König, Anne Schlegel, and Michal Bagrowski
Abstract
For the subsequent analysis of the methylated mSEPT9 colorectal cancer screening marker in plasma, different blood collection tubes and blood storage conditions were investigated. The study demonstrated that methylated Septin 9 (mSEPT9) can be consistently detected in plasma samples derived from whole blood samples collected with S-Monovette® K3E and BD Vacutainer ® K2EDTA tubes stored at 2–8 °C for a maximum of 24 h and for samples collected in S-Monovette CPDA tubes stored at 18–25 °C for up to 48 h. Keywords
Blood • Stability • Collection tube • Septin 9 • Epigenetic marker • Colorectal cancer
Introduction
J. Distler (*) • R. Tetzner • G. Weiss • T. König A. Schlegel • M. Bagrowski Epigenomics AG, Geneststr. 5, Berlin 10829, Germany e-mail: [email protected]
The methylated Septin 9 DNA marker is highly correlated with the presence of colorectal cancer (CRC) (deVos et al. 2009). The analysis of m SEPT9 is performed with free circulating DNA (cfDNA) extracted from human plasma using the Epi proColon® 2.0 CE IVD kit (Epigenomics 2014; Jin et al. 2015). The shipment of blood samples requires the blood to be drawn in
© Springer International Publishing Switzerland 2016 P.B. Gahan et al. (eds.), Circulating Nucleic Acids in Serum and Plasma – CNAPS IX, Advances in Experimental Medicine and Biology 924, DOI 10.1007/978-3-319-42044-8_32
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collection tubes that preserve cfDNA and prevent lysis of blood cells before the plasma can be processed in the laboratory. Ideally, blood collection tubes should allow the shipment of blood at room temperature to avoid complex and costly logistics. The goal of the present studies was to identify a tube enabling the preservation of cfDNA and prevention of lysis of blood cells for at least 72 h at ambient temperature.
according to the manufacturers instruction for use (Epigenomics, Berlin, Germany). The analytical sensitivity (% mSEPT9 positive samples) and analytical specificity (100 % – % m SEPT9 positive samples) were determined with plasma of spiked and unspiked blood samples, respectively.
Results Materials and Methods Blood from male and female healthy donors (age 35–79 years) was collected with different blood collection tubes. To simulate a positive specimen, blood samples were spiked with 15 genome equivalents of HeLa DNA (BioChain Institute, Newark, USA), a genomic human tumor cell line DNA per millilitre blood. The blood draws with the following collection tubes were performed as recommended by the manufacturers: S-Monovette® K3E (Sarstedt, Nümbrecht, Germany) and S-Monovette® CPDA (Sarstedt, Nümbrecht, Germany), Cell-Free DNA BCT® (Streck, Omaha, USA), PAXgene® Blood DNA Tube (PreAnalytiX GmbH, Hombrechtikon, CH), and BD Vacutainer ® K2EDTA, (Becton Dickinson, Franklin Lakes, USA). Blood collected in BD Vacutainer ® PPT (Becton Dickinson, Franklin Lakes, USA) was not centrifuged directly after blood draw as recommende
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