Abstract Laura Kennedy¹, J. Keith Vass¹, D. Ross Haggart¹, Steve Moore¹, Michael E. Burczynski², Dan Crowther¹ and Gino Miele¹

¹Translational Medicine Research Collaboration Laboratory, Sir James Black Centre, University of Dundee, Dundee, DD1 5EH, U.K. ²Biomarker Laboratory, Wyeth Research, Collegeville, U.S.A.

Abstract

Peripheral blood as a surrogate tissue for transcriptome profiling holds great promise for the discovery of diagnostic and prognostic disease biomarkers, particularly when target tissues of disease are not readily available. To maximize the reliability of gene expression data generated from clinical blood samples, both the sample collection and the microarray probe generation methods should be optimized to provide stabilized, reproducible and representative gene expression profiles faithfully representing the transcriptional profiles of the constituent blood cell types present in the circulation. Given the increasing innovation in this field in recent years, we investigated a combination of methodological advances in both RNA stabilisation and microarray probe generation with the goal of achieving robust, reliable and representative transcriptional profiles from whole blood. To assess the whole blood profiles, the transcriptomes of purified blood cell types were measured and compared with the global transcriptomes measured in whole blood. The results demonstrate that a combination of PAXgene^TM RNA stabilising technology and single-stranded cDNA probe generation afforded by the NuGEN Ovation RNA amplification system V2^TM enables an approach that yields faithful representation of specific hematopoietic cell lineage transcriptomes in whole blood without the necessity for prior sample fractionation, cell enrichment or globin reduction. Storage stability assessments of the PAXgene^TM blood samples also advocate a short, fixed room temperature storage time for all PAXgene^TM blood samples collected for the purposes of global transcriptional profiling in clinical studies.

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