Optimizing RNA extraction yield from whole blood for microarray gene expression analysis.

Author(s): Wang J, Robinson JF, Khan HM, Carter DE, McKinney J, Miskie BA, Hegele RA

Publication: Clin Biochem, 2004, Vol. 37, Page 741-4

PubMed ID: 15329310 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to determine the effects of storage of specimens in PAXgene preservative and between day variability on RNA yield and transcript profiling using microarrays. 4-6 specimens were obtained from each of two volunteers after a 12 h fast on each of two consecutive days.

   Summary of Findings:

   Significantly more RNA was extracted from blood stored for 24 h than from blood stored for 4 h in PAXgene preservative solution (4.33 ug/mL versus 2.53 ug/mL, p<0.0001), but no differences in RNA quality were observed. The authors report 48.2-51.2% of genes were present after hybridization, but do not indicate if the percentage depended on storage duration. Using Affymetrix DMT, a greater than 1.5 fold increase was calculated in 0.58% of transcripts from specimens collected on day 1 and stored in PAXgene solution for 4 h compared to those collected from the same subject on day 2 and stored in PAXgene solution for 24 h. At the same time, a 1.5 fold or greater decrease was calculated in 3.03% of transcripts collected from specimens between the same two conditions. Further, using Affymetrix DMT, transcript levels from specimens from a single subject stored in PAXgene solution for 24 h showed a 1.5 fold or greater increase between day 1 and 2 for 0.16% of transcripts and a 1.5 fold or greater decrease between day 1 and 2 for 0.20% of transcripts. More transcripts displayed greater than 1.5 fold increases or decreases between collection days using GeneSpring or Bullfrog analysis tools than using Affymetrix DMT.

   Biospecimens

   • Bodily Fluid - Blood

   Preservative Types

   • PAXgene

   Diagnoses:

   • Normal

   Platform:

   Analyte	Technology Platform
   RNA	Spectrophotometry
   RNA	Automated electrophoresis/Bioanalyzer
   RNA	DNA microarray

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Storage	Time at room temperature	4 h 24 h
   Biospecimen Acquisition	Time of biospecimen collection	Day 1 Day 2
   DNA microarray Specific	Data handling	Affymetrix DMT GeneSpring Bullfrog

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