The detection of circulating molecular biomarkers is useful for monitoring pathogenic processes and response to therapeutic intervention1,2. In cancer, genomic and proteomic biomarkers are used to assess tumour sub-type and stage, and determine patient stratification. Research has shown that using a combined signature of multiple biomarkers can better account for patient and epidemiological heterogeneity, and provide a more accurate indication of patient health2. In addition, the desire to leverage minimally-invasive samples has necessitated the development of technologies that can profile biomarkers directly from biofluids such as plasma, serum, and urine. To address this need, we developed the FirePlex Technology platform, which allows for the sensitive and accurate multiplexed detection of protein analytes or miRNAs directly from a wide array of biological samples. The platform uses patented, multi-functional hydrogel particles that allow for in-well multiplexing and panel design flexibility.
For the detection of protein analytes, FirePlex immunoassays use high-performance matched antibody pairs that provide minimal cross-reactivity between individual analytes, up to 5 logs dynamic range, and single-digit pg/ml sensitivity, while requiring only 12.5 µl biofluid input. Similarly, the FirePlex miRNA assay can reliably detect miRNAs directly from as little as 10 µl of biofluid without the need for RNA purification. This assay utilizes single¬ step RT-PCR signal amplification with universal primers, thus leveraging PCR sensitivity while eliminating an independent reverse transcription step and mitigating amplification biases introduced by target¬-specific qPCR.
FirePlex assays are validated across a wide range of biological samples including plasma, serum, urine and cell culture supernatant. The 96-well plate assay format enables high-throughput sample screening with readout on standard flow cytometers, thereby omitting the need for complex and expensive dedicated instrumentation. Finally, the integrated FirePlex Analysis Workbench software enables rapid data analysis, visualization, and export, and includes key features such as standard curves as well as publication-quality heatmaps and graphs.
Here we present data from studies using the FirePlex Platform to profile miRNAs in cancer. Together, the combination of multiplexed, high¬-sensitivity assays with powerful bioinformatics tools enables rapid discovery and validation of biomarker signatures from liquid and solid specimens.