TY - JOUR
T1 - The effect of preservative and temperature on the analysis of circulating tumor DNA
AU - Parpart-Li, Sonya
AU - Bartlett, Bjarne
AU - Popoli, Maria
AU - Adleff, Vilmos
AU - Tucker, Laura
AU - Steinberg, Rebecca
AU - Georgiadis, Andrew
AU - Phallen, Jill
AU - Brahmer, Julie
AU - Azad, Nilo
AU - Browner, Ilene
AU - Laheru, Daniel
AU - Velculescu, Victor E.
AU - Sausen, Mark
AU - Diaz, Luis A.
N1 - Publisher Copyright:
©2016 AACR.
PY - 2017/5/15
Y1 - 2017/5/15
N2 - Purpose: Analysis of genomic alterations in cell-free DNA (cfDNA) is evolving as an approach to detect, monitor, and genotype malignancies. Methods to separate the liquid from the cellular fraction of whole blood for circulating tumor DNA (ctDNA) analyses have been largely unstudied, although these may be a critical consideration for assay performance. Experimental Design: To evaluate the influence of blood processing on cfDNA and ctDNA quality and yield, we compared the cfDNA levels in serum with those in plasma. Given the limitations of serum for ctDNA analyses, we evaluated the effects of two plasma processing approaches, K2EDTA and Cell-Free DNA BCT (BCT) tubes, on cfDNA and ctDNA recovery. A total of 45 samples from nine patients with cancer were collected in both tube types. Once collected, blood was processed into plasma immediately or kept at room temperature and processed into plasma at 1, 3, 5, or 7 days. Results: As early as 24 hours after collection, plasma isolated from blood collected in K2EDTA tubes contained an elevated level of cfDNA that increased over time compared with BCT tubes where no significant increase in cfDNA levels was observed. When samples from an additional six patients with cancer, collected in the same manner, were stored at 4°C in K2 EDTA tubes over the course of 3 days, total cfDNA and ctDNA levels were comparable between samples collected in BCT tubes. At day 3, there was a trend toward a decrease in ctDNA levels in both tubes that was more pronounced when measuring the mutant allele fraction for cases stored at 4°C in K2EDTA tubes. Conclusions: In summary, methods of blood processing have a strong influence on cfDNA and ctDNA levels and should be a consideration when evaluating ctDNA in peripheral circulation.
AB - Purpose: Analysis of genomic alterations in cell-free DNA (cfDNA) is evolving as an approach to detect, monitor, and genotype malignancies. Methods to separate the liquid from the cellular fraction of whole blood for circulating tumor DNA (ctDNA) analyses have been largely unstudied, although these may be a critical consideration for assay performance. Experimental Design: To evaluate the influence of blood processing on cfDNA and ctDNA quality and yield, we compared the cfDNA levels in serum with those in plasma. Given the limitations of serum for ctDNA analyses, we evaluated the effects of two plasma processing approaches, K2EDTA and Cell-Free DNA BCT (BCT) tubes, on cfDNA and ctDNA recovery. A total of 45 samples from nine patients with cancer were collected in both tube types. Once collected, blood was processed into plasma immediately or kept at room temperature and processed into plasma at 1, 3, 5, or 7 days. Results: As early as 24 hours after collection, plasma isolated from blood collected in K2EDTA tubes contained an elevated level of cfDNA that increased over time compared with BCT tubes where no significant increase in cfDNA levels was observed. When samples from an additional six patients with cancer, collected in the same manner, were stored at 4°C in K2 EDTA tubes over the course of 3 days, total cfDNA and ctDNA levels were comparable between samples collected in BCT tubes. At day 3, there was a trend toward a decrease in ctDNA levels in both tubes that was more pronounced when measuring the mutant allele fraction for cases stored at 4°C in K2EDTA tubes. Conclusions: In summary, methods of blood processing have a strong influence on cfDNA and ctDNA levels and should be a consideration when evaluating ctDNA in peripheral circulation.
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U2 - 10.1158/1078-0432.CCR-16-1691
DO - 10.1158/1078-0432.CCR-16-1691
M3 - Article
C2 - 27827317
AN - SCOPUS:85020448949
SN - 1078-0432
VL - 23
SP - 2471
EP - 2477
JO - Clinical Cancer Research
JF - Clinical Cancer Research
IS - 10
ER -