Kang Qing, Henry N Lynn, Paoletti Costanza, Jiang Hui, Vats Pankaj, Chinnaiyan Arul M, Hayes Daniel F, Merajver Sofia D, Rae James M, Tewari Muneesh
Department of Internal Medicine, Division of Hematology/Oncology, University of Michigan, Ann Arbor, MI, USA.
Department of Biostatistics, School of Public Health, University of Michigan, Ann Arbor, MI, USA.
Clin Biochem. 2016 Dec;49(18):1354-1360. doi: 10.1016/j.clinbiochem.2016.03.012. Epub 2016 Apr 27.
Optimal conditions for blood collection for circulating tumor DNA (ctDNA) are still being developed. Although both Streck and EDTA tubes are commonly used, their ability to stabilize ctDNA as a function of time and temperature post-collection has not been thoroughly studied. Additionally, the potential utility of CellSave tubes (commonly used for circulating tumor cell) for ctDNA measurements has not been studied.
Blood was collected into Streck, EDTA, and CellSave tubes from ten patients with metastatic breast cancer enrolled in the MI-ONCOSEQ tumor sequencing program at the University of Michigan and kept either on ice or at room temperature until plasma isolation. Plasma was processed after 2, 6, and 48h post-collection. We used droplet digital PCR (ddPCR) to quantify plasma ctDNA and wild-type DNA for six patients who had tumor tissue mutations represented in commercially available ddPCR assays.
ctDNA abundance was similar and stable for up to 6h in all tube types, and there was no effect of storage temperature on the yield for Streck and EDTA tubes. After 48h, however, one out of four patients with detectable ctDNA showed a ~50% decline in ctDNA in the EDTA tube, and three out of six patients showed a 2-3-fold increase in wild-type DNA in the EDTA tube.
Streck, EDTA, and CellSave tubes showed similar performance in preserving ctDNA for up to 6h before plasma isolation. Streck and CellSave tubes more consistently stabilized ctDNA and wild-type DNA at 48h than EDTA tubes.
循环肿瘤DNA（ctDNA）采血的最佳条件仍在探索之中。尽管Streck管和乙二胺四乙酸（EDTA）管都常用，但它们在采血后随时间和温度稳定ctDNA的能力尚未得到充分研究。此外，用于循环肿瘤细胞检测的CellSave管在ctDNA检测中的潜在用途也未被研究。
从密歇根大学MI-ONCOSEQ肿瘤测序项目中招募的10例转移性乳腺癌患者采集血液，分别放入Streck管、EDTA管和CellSave管中，置于冰上或室温下直至分离血浆。采血后2、6和48小时对血浆进行处理。我们使用液滴数字PCR（ddPCR）对6例在市售ddPCR检测中有肿瘤组织突变的患者的血浆ctDNA和野生型DNA进行定量。
在所有管型中，ctDNA丰度在6小时内相似且稳定，储存温度对Streck管和EDTA管的产量没有影响。然而，48小时后，在4例可检测到ctDNA的患者中，有1例患者的EDTA管中ctDNA下降了约50%，在6例患者中，有3例患者的EDTA管中野生型DNA增加了2至3倍。
在分离血浆前，Streck管、EDTA管和CellSave管在保存ctDNA方面表现相似，长达6小时。在48小时时，Streck管和CellSave管比EDTA管更能持续稳定ctDNA和野生型DNA。
