Cancer Diagnosis Program | Biorepositories and Biospecimen Research Branch

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Stabilization of circulating tumor cells in blood using a collection device with a preservative reagent.

Author(s): Qin J, Alt JR, Hunsley BA, Williams TL, Fernando MR

Publication: Cancer Cell Int, 2014, Vol. 14, Page 23

PubMed ID: 24602297 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   This study compared the stability of CTCs in peripheral blood collected in different tube types after transportation and storage of tubes for up to 4 days. Peripheral blood (10 mL) from seven healthy volunteers was collected into two of each type of blood collection tube (K3EDTA, CellSave, and Cell-free DNA BCT), spiked with 2,000 MCF-7 (breast cancer cell line) cells and inverted 10 times. Specimens were shipped and maintained at ambient temperature and analyzed for recovery using the CellSearch system one or four days after collection. An additional 10 mL of blood was collected from each donor into a K3EDTA tube and a BCT. Plasma was separated within 2 h by centrifugation at 300 × g for 20 min at RT. Cell-free plasma specimens (4-5 mL) were spiked with ~ 2,000 MCF-7 cells and stored at ambient temperature for zero or four days until immunofluorescent analysis for EpCAM and CK proteins and fluorescence in situ hybridization for c-fos and cyclin D1.

   Summary of Findings:

   Comparable percentages of cancer cells (spiked into peripheral blood using the MCF-7 cell line) were recovered from peripheral blood that was stored at ambient temperature for one and four days when blood was collected in BCT (60% and 58%, respectively) and CellSave tubes (52% and 54%, respectively); however, recovery was significantly lower when blood was collected and stored in K3EDTA tubes (32% at Day 1, P<0.001 and 16% at Day 4, P<0.0003). The percentage of cancer cells recovered from plasma transported and stored in K3EDTA tubes was statistically lower than BCT tubes after one and four days of storage (32% vs. 61% and 16% vs. 57%, respectively). EpCAM and cytokeratin proteins, cell nucleus and nuclear content, and c-fos and cyclin D1 mRNA were stable in plasma stored in BCTs at ambient temperature for four days; however, cancer cells stored in K3EDTA tubes showed degradation of EpCAM, cytokeratin, cell nucleus, nuclear content, and c-fos mRNA and increased cyclin D1 mRNA levels. The authors state that the storage-induced changes in mRNA levels when cancer cells were stored in K3EDTA tubes may be a result of cellular degradation.

   Biospecimens

   • Bodily Fluid - Blood

   Preservative Types

   • Streck/BCT
   • Other Preservative

   Diagnoses:

   • Normal

   Platform:

   Analyte	Technology Platform
   Cell count/volume	Fluorescent microscopy
   RNA	In situ hybridization
   Protein	Immunohistochemistry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Fluorescent microscopy Specific	Targeted peptide/protein	EpCAM cytokeratin
   In situ hybridization Specific	Targeted nucleic acid	DAPI c-fos cyclin D1
   Biospecimen Acquisition	Type of collection container/solution	CellSave tube K3EDTA tube BCT
   Immunohistochemistry Specific	Targeted peptide/protein	EpCAM cytokeratin
   Storage	Time at room temperature	0 days 1 day 4 days

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