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BEAMing and Droplet Digital PCR Analysis of Mutant IDH1 mRNA in Tumor Extracellular Vesicles
Massachusetts General Hospital

Development of biofluid-based molecular diagnostic tests for cancer is an important step towards tumor characterization and real-time monitoring in a minimally invasive fashion. Extracellular vesicles (EVs) are released from tumor cells into body fluids and can provide a powerful platform for tumor biomarkers since they carry tumor proteins and nucleic acids. BEAMing (beads, emulsion, amplification, magnetics) PCR is substantially more sensitive than many other assays for detecting single nucleotide mutations when there is a significant background of wild-type sequences. Picodroplet digital PCR developed by RainDance Technologies is another highly sensitive technology that can be used to quantify rare mutations in a high wild type background. Here, we describe a novel approach that uses BEAMing droplet RT-PCR, as well picodroplet digital RT-PCR, to interrogate mRNA sequences contained within EVs from serum and cerebrospinal fluid (EV-BEAMing). Using both assays, we were able to reliably detect and quantify mutant and wild-type IDH1 transcripts in cerebrospinal fluid of patients with gliomas. EV-BEAMing and picodroplet dPCR are valuable new tools for cancer diagnostics which can be applied to a variety of biofluids and neoplasms.

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