#C11885: miRNA Analysis Method
Unmet Need
The gold standard for miRNA profiling is quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). While qRT-PCR is fast, reproducible, and can handle smaller input samples than other profiling methods (Chugh et al.), it can only evaluate less than 10 miRNAs at a time. Other methods can profile thousands of miRNAs, but they are not as reproducible and are more expensive and time-consuming. Thus, there is a need for a miRNA profiling method that combines the reproducibility and speed of qRT-PCR with the scale of other methods. Such a method would facilitate exploration of miRNAs’ roles in the cell and their use in diagnostics and treatments.
Value Proposition
· Performs 4-hour profiling of 10-100 microRNAs (miRNAs) compared to 12 hours for less than 10 miRNAs via qRT-PCR.
· miRNA detection limit is comparable to that of existing technologies.
· Reagents cost 25-50-fold less than those for competing technologies.
· Does not require purification, separation, or wash steps. Additionally, assay readout requires only a standard gel electrophoresis apparatus and gel reader as opposed to expensive (>$200k) genomic analyzers needed for some multiplex methods.
· Minimal sample volume required.
Technology Description
Researchers at Johns Hopkins have developed Ligo-miR, a platform of miRNA profiling assays that enables analysis of 10-100 miRNAs in a single reaction. The assay uses a novel 2-step ligation process, successively adding different probes to create size encoded miRNA products that can be individually identified and quantified by gel separation (Ligo-miR EZ) or single molecule analysis (Ligo-miR HD). Ligo-miR EZ is intended for low-cost applications such as clinical diagnostics; it requires only a standard gel apparatus and reader for detection. Ligo-miR HD maximizes detection sensitivity (as low as 20 molecules) and minimizes sample consumption (million-fold reduction in assay volume). It is intended for extremely sensitive samples such circulating tumor cells (CTC) or single cell spatial profiling of tumors. Ligo-miR Bead combines magnetic bead-based RNA isolation with the Ligo-miR assay to detect miRNA directly from clinical samples without requiring separate RNA isolation.
Data Availability
Data available upon request.
Publication
Kilburn, D., Song, Y., Wang, T. H., & Liu, K. J. (2017). A Multiplex Ligation Assay for miRNA Copy Number Profiling. In MicroRNA Profiling (pp. 185-193). Humana Press, New York, NY.
