Abstract 3155: High-throughput profiling of miRNA expression in plasma and cerebrospinal fluid using OpenArray® platform for biomarker discovery

Abstract

Abstract OpenArray® platform can be used for expression profiling using TaqMan® assays that enables higher throughput, less assay consumption, and lower sample input than other low-density assay platforms. One OpenArray® plate has 3,072 through-holes, each of which requires only 33nl of reaction volume. We combined the workflow of Megaplex™ RT primer pools and pre-amplification with TaqMan® MicroRNA assays on OpenArray® plates. Each OpenArray® plate provides the capacity of 3 samples, and each run can take up to 3 plates with a run time about 2.5 hours. Therefore, each typical work day can have 4 instrument runs generating expression data of 754 miRNAs from 36 samples. This platform is ideal to identify potential miRNA biomarkers within a short period of time by screening a large cohort of clinical specimens that might have limitations in sample size, contamination of PCR inhibitors, or low abundance of target miRNAs. We optimized the original workflow of Megaplex™ RT primer pools and pre-amplification before applying to the OpenArray® plates to ensure increasing detection sensitivity while maintaining the linearity in plasma samples. In a proof-of-principle investigation, we applied the new protocol to a small panel of plasma and cerebrospinal fluid (CSF) specimens derived from pediatric patients diagnosed with either germinoma or non-germinoma germ cell tumors of the brain, and benchmarked with another miRNA expression profiling platform, TaqMan® Array Card (TAC), and OpenArray® plates demonstrated good concordance with the TAC. Expression patterns of miRNAs detected in our CSF samples are largely consistent with previously published miRNAs differentially expressed in primary cancerous tissues between these two types of intracranial germ cell tumors. (For Research Use Only. Not for use in diagnostic procedures.) Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3155. doi:1538-7445.AM2012-3155