To do this, the UCSF research group designed their own microRNA preAMP and PCR primers specifically to be used on the BioMark platform. Through this method, they were able to identify mRNA signatures that correlate with prognosis.
One in six men will be diagnosed with prostrate cancer during their lifetime. Recent prostate specific antigen (PSA) based screening trials indicate an urgent need for novel and non-invasive biomarker identification strategies to improve the prediction of prostate cancer behaviour. Non-coding microRNAs (miRNAs) in the serum and plasma have been shown to have potential as non-invasive markers for physiological and pathological conditions. To identify serum miRNAs that diagnose and correlate with prognosis of prostate cancer, the researchers at UCSF developed a multiplex quantitative reverse transcription PCR (qRT-PCR) method involving purification of multiplex PCR products followed by uniplex analysis on a Fluidigm microfluidics chip to evaluate 384 human miRNAs. Using Dgcr8 and Dicer knockout (small RNA - deficient) mouse ES cells (mESC) as the benchmark, the researchers confirmed the validity of their technique, while uncovering a significant lack of accuracy in previously published methods. Profiling 48 sera from healthy men and untreated prostate cancer patients with differing CAPRA (Cancer of the Prostate Risk Assessment) scores, UCSF researchers identified miRNA signatures that allowed them to diagnose cancer patients and correlate with prognosis. These serum signatures included oncogenic and tumor suppressive miRNAs suggesting functional roles in prostate cancer progression.
The Fluidigm BioMark System changes the way that biologists can conduct high-throughput real-time qPCR studies. Using microfluidic chips, the BioMark System features low input requirements and provides the sensitivity and assay throughput necessary for qPCR. Using standard TaqMan and DNA binding dye assays, Fluidigm's proprietary Integrated Fluidic Circuit (IFC) technology enables single-cell discovery with the ability to screen hundreds of genes from single cells. Now with this unique technology it is possible to conduct up to 36,000 PCR reactions in one day, with one technician, and one BioMark System.
Fluidigm SARL
