title: Global transcriptomic profiling using small volumes of whole blood: a cost-effective method for translational genomic biomarker identification in small animals.
abstract: Blood is an ideal tissue for the identification of novel genomic biomarkers for toxicity or efficacy. However, using blood for transcriptomic profiling presents significant technical challenges due to the transcriptomic changes induced by ex vivo handling and the interference of highly abundant globin mRNA. Most whole blood RNA stabilization and isolation methods also require significant volumes of blood, limiting their effective use in small animal species, such as rodents. To overcome these challenges, a QIAzol-based RNA stabilization and isolation method (QSI) was developed to isolate sufficient amounts of high quality total RNA from 25 to 500 μL of rat whole blood. The method was compared to the standard PAXgene Blood RNA System using blood collected from rats exposed to saline or lipopolysaccharide (LPS). The QSI method yielded an average of 54 ng total RNA per μL of rat whole blood with an average RNA Integrity Number (RIN) of 9, a performance comparable with the standard PAXgene method. Total RNA samples were further processed using the NuGEN Ovation Whole Blood Solution system and cDNA was hybridized to Affymetrix Rat Genome 230 2.0 Arrays. The microarray QC parameters using RNA isolated with the QSI method were within the acceptable range for microarray analysis. The transcriptomic profiles were highly correlated with those using RNA isolated with the PAXgene method and were consistent with expected LPS-induced inflammatory responses. The present study demonstrated that the QSI method coupled with NuGEN Ovation Whole Blood Solution system is cost-effective and particularly suitable for transcriptomic profiling of minimal volumes of whole blood, typical of those obtained with small animal species.
authors: Global Pharmaceutical Research and Development, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, Il 60064, USA; E-Mails: meagan.fricano@abbott.com (M.M.F.); amy.ditewig@abbott.com (A.C.D.); paul.jung@abbott.com (P.M.J.); michael.liguori@abbott.com (M.J.L.); eric.blomme@abbott.com (E.A.G.B.).
authors: Meagan M
authors: MM
authors: Fricano
authors: Amy C
authors: AC
authors: Ditewig
authors: Paul M
authors: PM
authors: Jung
authors: Michael J
authors: MJ
authors: Liguori
authors: Eric A G
authors: EA
authors: Blomme
authors: Yi
authors: Y
authors: Yang
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000097:blood*; D016000:Cluster Analysis; D020869:Gene Expression Profiling / Q000191:economics / Q000379:methods*; D007249:Inflammation / Q000378:metabolism; D008070:Lipopolysaccharides / Q000633:toxicity; D008297:Male; D020411:Oligonucleotide Array Sequence Analysis; D012313:RNA / Q000097:blood / Q000302:isolation & purification; D020871:RNA Stability / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects; D059467:Transcriptome
chemical_list: D015415:Biomarkers; D008070:Lipopolysaccharides; D012313:RNA