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Impact of Delayed Whole Blood Processing Time on Plasma Levels of miR-1 and miR-423-5p up to 24 Hours.

Author(s): Borges DP, Cunha-Neto E, Bocchi EA, Rigaud VOC

Publication: Microrna, 2018, Vol. , Page

PubMed ID: 29564990 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   This study investigated the effects of delayed centrifugation of blood on hemolysis and levels of miR-1, miR-423-5p, miR-23a, and miR-451. Blood was collected from 14 breast cancer patients by venipuncture into EDTA vacuum tubes. Blood was stored at room temperature for 0, 3, and 24 h before centrifugation at 2000 x g at 4˚C for 15 min. Plasma was transferred to microcentrifuge tubes and stored at -80˚C until analysis. Hemoglobin in plasma was determined by ultra-violet spectrophotometry. miRNAs were extracted from plasma using the miRNeasy serum/plasma kit and reverse transcribed using the TaqMan microRNA Reverse Transcription kit. miR-1, miR-423-5p, miR-23a, miR-451, and spiked-in Cel-miR-39 were quantified using the TaqMan microRNA assay.

   Summary of Findings:

   Unacceptable levels of hemolysis (λ414 >0.2) were found in 4 of the 14 specimens processed without delay and thus these specimens were excluded from further analysis. Hemolysis increased from a mean λ414 = 0.16±0.01 with increasing pre-centrifugation delay. No significant difference from immediately processed specimens were noted when centrifugation was delayed by only 3 h (mean λ414 = 0.19±0.02), but the mean λ414 increased to 0.30±0.02 (P<0.001) when centrifugation was delayed by 24 h. Levels of miR-1 and miR-423-5p were unaffected by delayed centrifugation. Levels of the hemolysis-related miR-451 increased with increasing delay to centrifugation (P<0.0001) and were modestly correlated with hemolysis (r=0.518, p<0.003). However, levels of the other hemolysis–related miRNA studied (miR-23a) were unaffected by delayed centrifugation and were not correlated with hemolysis in this study. In the four specimens with hemolysis when there was no processing delay, levels of miR-1 and miR-423-5p were comparable to those in non-hemolyzed specimens.

   Biospecimens

   • Bodily Fluid - Plasma
   • Bodily Fluid - Blood

   Preservative Types

   • Frozen

   Diagnoses:

   • Neoplastic - Carcinoma

   Platform:

   Analyte	Technology Platform
   Protein	Spectrophotometry
   RNA	Real-time qRT-PCR

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Storage	Time at room temperature	0 h 3 h 24 h
   Biospecimen Aliquots and Components	Hemolysis	Present Absent
   Real-time qRT-PCR Specific	Targeted nucleic acid	miR-1 miR-23a miR-423-5p miR-451

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