Abstract 1870: Development of a standard operating procedure for exosome isolation and analysis using clinical samples: Application to cancer biomarker discovery

Abstract

Abstract Exosomes are small vesicles (30-150 nm) found in abundance in human body fluids which function as carriers of different species of RNA and protein between diverse locations in the body. The spectrum of current scientific interest in exosomes is wide and ranges from studying their functions and pathways to utilizing them in diagnostics and therapeutics development. As such, there is a growing need for quick and easy methods for both isolation of exosomes and analysis of their cargo. We present herein a workflow for exosome isolation and analysis which entails: (i) fast and efficient isolation of exosomes from serum, plasma, and urine of both healthy donors and patients with prostate cancer, using Total Exosome Isolation reagents; (ii) characterization of their size distribution and count with Nanosight LM10 instrument; (iii) extraction of exosome “cargo” with Total Exosome RNA and Protein Isolation kit; (iv) characterization of exosomal RNA content using the Ion Torrent PGM sequencing and qRT-PCR. The protocol described herein lays the groundwork for the development of a standardized operating procedure (SOP) for isolation of exosomes and downstream analysis of their constituents, using clinical samples. We demonstrate that cancer-specific RNA signatures residing within the exosomes can be delineated from different patient cohorts. This is the first step towards developing a method whereby performance characteristics can be measured and used to optimize a validated assay useful for routine testing of clinical samples. Citation Format: Robert A. Setterquist, Alex J. Rai, Emily Zeringer, Mu Li, Tim Barta, Jeoffrey Schageman, Susan Magdaleno, Alexander V. Vlassov. Development of a standard operating procedure for exosome isolation and analysis using clinical samples: Application to cancer biomarker discovery. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1870. doi:10.1158/1538-7445.AM2014-1870