Abstract LB-328: Highly sensitive detection of miRNA and mRNA from frozen, FFPE tissue and body fluid samples by expression microarray

Abstract

Abstract The RNA profiling from clinical samples readily obtainable in the routine care, such as blood or FFPE samples, is a promising method to find the unknown high-value diagnostic markers. However, as RNA is subjected to degradation even in properly-collected, and immediately frozen tissue, it is more difficult to obtain intact RNA from FFPE or body fluid samples for diagnostic analysis. “3D-Gene” is a highly sensitive gene expression microarray, featuring the unique pillar structure on the substrate and the beads agitation system during the hybridization reaction. Using “3D-Gene”, we achieved highly sensitive detection of mRNA or miRNA from diagnostically important clinical samples. Total RNA was extracted from human serum, plasma and frozen or FFPE tissue samples, with the recommended protocol for each sample. For mRNA detection, total RNA was reverse-transcribed to cDNA and labeled with fluorescent dye directly or after the amplification. For miRNA detection, total RNA was labeled with fluorescent dye directly. These pretreated target nucleotides were hybridized to “3D-Gene” while the hybridized buffer containing target nucleotides was agitated by beads during hybridization. The hybridized microarrays were washed and scanned for image acquisition. (1) With “3D-Gene”, we could detect the mRNA expression profile from FFPE samples with high reproducibility. We also showed high correlation of the expression profiles between FFPE and frozen tissue samples. With 500 ng total RNA obtained from frozen as well as FFPE tissue samples, miRNA was reproducibly detected at attomolar level. Some miRNA biomarkers for various cancers were found from FFPE samples. (2) Serum and plasma are suggested to contain microsomes in which miRNA is enclosed. miRNA from serum and plasma samples were detected with high sensitivity and reproducibility with “3D-Gene”. Some miRNA biomarkers for various cancers were found from patients' serum. “3D-Gene” is a potential tool for detecting transcriptome with high sensitivity and useful for quest and validation of diagnostic biomarkers from clinical samples. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-328. doi:10.1158/1538-7445.AM2011-LB-328