Abstract 3966: Effect of anticoagulant and sample storage conditions on circulating tumor cells enrichment by vortex
Authors: Wilkerson Charles L. , Renier Corinne M. , Hanft Violet R. , Jeffrey Stefanie S. , Sollier Elodie 
Published: July 2016

Background

Circulating tumor cells (CTC) have recently emerged as powerful biomarkers for non-invasive diagnosis and prognosis of cancer [1]. The rarity and fragility of these cells has proven to be a significant impediment to their clinical use. Delay between phlebotomy and sample processing, together with blood sample storage, can impact the integrity of these cells and hamper their subsequent isolation. Addressing these issues could allow shipping between clinical facilities and significantly increase the use of CTCs. Here, we assessed the effects of various blood anticoagulants on the isolation of CTCs using Vortex technology. Vortex is a label-free, size-based technology that relies on the generation of laminar flow microvortices where CTCs can be trapped [2], and released off-chip, into a small volume compatible with various downstream assays.

Methods

To evaluate the effects of delayed processing on capture performances, samples from healthy donors were drawn into 4 different blood collection tubes (BCT): K3EDTA, ACD-B, CellSave and cell-free DNA Streck. Each BCT was spiked with MCF7 breast cancer cells and stored at room temperature for up to 3 days (72H). On days 0, 1, 2 and 3, aliquots of blood (0.5 ml; n = 3) from each BCT were diluted 10X and processed through Vortex chip. Captured cells were stained (CK, CD45, DAPI) and enumerated. Capture efficiency and purity were determined for each time point and BCT. To simulate over-sea shipping and processing of patient samples, larger blood volumes (6mL in EDTA, CellSave and Streck BCT) were also tested after 48H. Capture performances and device clogging were evaluated.

Results

Significant differences were observed for capture performances between the BCTs. Best results were obtained for CellSave tube, with an efficiency of 19.8 ± 2.5%, stable for up to 3 days, and a purity ranging from 40.6 to 52.8%. In comparison, performances for standard K3EDTA were maximal at 24H, with 19.1% efficiency decreasing to 10% at 72H. ACD-B and Streck showed lower efficiencies, reaching 4.6 and 9.7% respectively. Purity for EDTA, CellSave and Streck BCT was in a similar range (48.4, 46.5 and 34.9% respectively). No significant device clogging was observed between EDTA, CellSave and Streck BCT when larger blood volumes were processed.

Conclusion

Here we demonstrate the ability to isolate cancer cells spiked in blood for up to 48H post-phlebotomy, with higher overall performances obtained with CellSave tube. Standard EDTA BCT gives optimal performances up to 24H and would be the option to select whenever viable cells are required. This suggests that Vortex technology is compatible with various anticoagulants and preservatives containing blood collection tube; the election would depend on the assay performed downstream. Future work will include shipping blood samples from remote sites and effect on CTC enrichment by Vortex technology.

[1] Ignatiadis et al. Clin Cancer Res 2015.

[2] Sollier et al. Lab Chip 2014.

Citation Format: Charles L. Wilkerson, Corinne M. Renier, Violet R. Hanft, Stefanie S. Jeffrey, Elodie Sollier. Effect of anticoagulant and sample storage conditions on circulating tumor cells enrichment by vortex. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3966.