Abstract 4955: Advances in gene expression microarray profiling from small amounts of adenocarcinoma and normal total RNA samples

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC Gene expression profiling by microarray analysis provides an important avenue for understanding biological mechanisms, classifying tissue and tumor types, and identifying signs for diagnosis and prognosis. To address the need for high sensitivity gene expression profiling of low quantities of total RNA, we have developed a modified linear amplification procedure that generates quantities of Cy-labeled cRNA suitable for oligonucleotide microarray experiments from total RNA input amounts as low as 10 nanograms. This new procedure, available in the new Low Input Quick Amp Labeling Kit, employs the AffinityScript Reverse Transcriptase, a mutant MMLV-RT that binds primer-template complexes with 10-fold higher efficiency than wild type MMLV-RT, resulting in increased cDNA yields and improved sensitivity from smaller sample inputs. The protocol uses a single round of IVT amplification without purification of the cDNA product resulting in labeled cRNA in less than one day, enabling gene expression profile comparisons in less than two days. Comparisons of probe signals from technical replicate samples demonstrate high reproducibility with wide dynamic ranges, and generally comparable signals across a broad range of input amounts. This new labeling approach was used to detect differences in gene expression between cancerous and normal cells using new gene expression arrays with updated content resulting in gene expression profiling results consistent with the current literature. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4955.