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Comparison of Roche Cell-Free DNA collection Tubes to Streck Cell-Free DNA BCTs for sample stability using healthy volunteers.

Author(s): Parackal S, Zou D, Day R, Black M, Guilford P

Publication: Pract Lab Med, 2019, Vol. 16, Page e00125

PubMed ID: 31289732 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   This study investigated the effects of storing blood in Streck and Roche cell-free DNA blood collection tubes on cfDNA yield and genomic DNA contamination and compared the electropherograms to those from blood stored in K₂EDTA tubes. Blood was collected from 20 healthy volunteers into five Streck cell-free DNA BCTs and five Roche cell-free DNA BCTs andfrom one volunteer into K₃EDTA Vacutainer tubes. Tubes were stored upright in a temperature controlled room (22˚C) for 0 days (<2 h from collection), 4 days, 7 days, 10 days, or 14 days. Plasma was obtained by centrifugation at 200 rcf for 10 min followed by 1600 rcf for 10 min with break and acceleration set to zero. The plasma was then transferred to a new tube and spun at 1600 rcf. DNA was extracted from plasma using the QIAamp Circulating Nucleic Acids Kit using centrifugation rather than vacuum and elution in DNAse-free/RNAse-free water rather than buffer. DNA concentration was determined using the Qubit dsDNA High Sensitivity Kit. Fragment size profile was evaluated by capillary electrophoresis on a bioanalyzer using the High Sensitivity DNA Analysis Kit.

   Summary of Findings:

   There was a high degree of inter-individual variability in cfDNA levels. Analysis of the data using the Kruskal-Wallis test showed no association with the changes in DNA concentration across days with tube type. The Wilcoxon rank-sum test showed no evidence that the distribution of concentration for the time-points was not centered around zero, but paired analysis identified significant increases in concentration between Day 0 and Days 7, 10, and 14 when stored in Roche tubes (P=0.01, P=0.046, and P=0.0016, respectively) but not Streck tubes (P>0.05). There were no significant differences using the Wilcoxon signed-rank test when specimens in Streck tubes at Day 0 were compared to specimens in Roche tubes at any time-point (P>0.05).

   Capillary electrophoresis showed obvious degradation of specimens stored in EDTA tubes for 14 days. For specimens stored in Streck tubes for 14 days, the cfDNA peak decreased and there was a slight increase in degradation products, but the cfDNA and other peaks (including a small genomic DNA peak) remained largely similar. For whole blood stored in Roche tubes, the only clear peak was from cfDNA and the peak was slightly smaller and the baseline more jagged after 14 days of storage, which the authors attribute to potential ethanol contamination. The authors conclude that genomic DNA contamination is largely avoided through the use of Streck or Roche tubes.

   Biospecimens

   • Bodily Fluid - Plasma
   • Bodily Fluid - Blood

   Preservative Types

   • Streck/BCT
   • None (Fresh)
   • Other Preservative

   Diagnoses:

   • Normal

   Platform:

   Analyte	Technology Platform
   DNA	Automated electrophoresis/Bioanalyzer
   DNA	Fluorometry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Acquisition	Type of collection container/solution	Streck cfDNA BCT K2EDTA tube Roche cfDNA BCT
   Biospecimen Aliquots and Components	Centrifugation	Centrifugation delays investigated
   Storage	Storage duration	0 days 4 days 7 days 14 days 10 days

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