Cancer Diagnosis Program | Biorepositories and Biospecimen Research Branch

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miRNA Stability in Frozen Plasma Samples.

Author(s): Balzano F, Deiana M, Dei Giudici S, Oggiano A, Baralla A, Pasella S, Mannu A, Pescatori M, Porcu B, Fanciulli G, Zinellu A, Carru C, Deiana L

Publication: Molecules, 2015, Vol. 20, Page 19030-40

PubMed ID: 26492230 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to compare levels of 8 miRNAs in matched fresh plasma and after storage at -80°C for 6 and 12 months. The authors also compared miRNA levels in fresh plasma with archival frozen plasma. K₂EDTA blood was collected in the early morning from five healthy volunteers (30-50 years of age). Plasma was separated by centrifugation at 2500 g for 15 min at 4°C and immediately aliquoted.  miRNA was extracted from the fresh aliquot immediately and from the other aliquots after 6 and 12 months of storage at -80°C using the miRNeasy Serum/Plasma Kit. Levels of U6 snRNA, miR-125b-5p, miR-425-5p, miR-200b-5p, miR-200c-3p, miR-579-3p, miR-212-3p, miR-126-3p, and miR-21-5p were quantified by TaqMan real-time RT-PCR assays and levels were normalized to U6 snRNA. Additionally, miRNA was extracted from frozen plasma stored at -80°C since 1999 (5 volunteers), 2002 (4 volunteers), 2003 (5 volunteers), 2009 (5 volunteers), and 2010 (5 volunteers); and miRNA levels from these archival specimens were compared to those in fresh plasma (collected and promptly analyzed in 2013).

   Summary of Findings:

   Levels of all eight miRNAs investigated were unaffected by ≤ 12 months of storage at -80°C. However, miR-126-3p levels were significantly lower in frozen plasma collected in 1999 (P=0.007), 2002 (P-value not provided), 2003 (P-value not provided), and 2009 (P=0.021), than in unmatched fresh plasma.  However, miR-126-3p levels were comparable in frozen plasma collected and frozen in 2010 and unmatched fresh plasma (collected and promptly analyzed in 2013). In addition to the aforementioned reduction in miR-126-3p levels, frozen plasma from 1999 also had lower levels of miR-125b-5p (P=0.007), miR-425-5p (P=0.017), miR-200b-5p (P=0.008), miR-200c-3p (P<0.001), miR-579-3p (P=0.021), and miR-21-5p (P=0.002) in comparison to unmatched fresh plasma.

   Biospecimens

   • Bodily Fluid - Plasma
   • Bodily Fluid - Blood

   Preservative Types

   • None (Fresh)
   • Frozen

   Diagnoses:

   • Normal

   Platform:

   Analyte	Technology Platform
   RNA	Real-time qRT-PCR

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Preservation	Type of fixation/preservation	Frozen None (fresh)
   Real-time qRT-PCR Specific	Targeted nucleic acid	U6 snRNA miR-125b-5p miR-425-5p miR-200b-5p miR-200c-3p miR-579-3p miR-212-3p miR-126-3p miR-21-5p
   Storage	Storage duration	0 months 6 months 12 months 2010-2013 2009-2013 2003-2013 2002-2013 1999-2013
   Storage	Freeze/thaw cycling	0 cycles 1 cycle

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