Extracellular vesicles, including exosomes, are currently being investigated to better understand their biogenesis and biological functions. There is also a rapidly growing interest in utilizing exosomes present in patient biofluids for molecular diagnostics in the clinic. Exosomes are natural shuttles of RNA and protein cargo, making them attractive as potential therapeutic delivery vehicles.
Here, the authors describe the methods for using the latest tools and technologies to study exosomes to better understand their roles in cell-to-cell communication, for discovery of clinical biomarkers and to engineer exosomes for therapeutic applications.
Microarray analysis of exoRNA samples. Exosome RNA was purified from human normal control vs. colon cancer patient sera. (A) The exoRNA was converted to cDNA and amplified using the SeraMir kit. The amplified libraries were visualized on a 2% agarose-TAE ethidium bromide gel. A sample profile is shown with sizes labeled in base pairs (bp). The amplified cDNAs were then fluorescently labeled and hybridized to an LC Sciences mirBase ver. 16 microarray chip containing about 1200 different miRNA probes as a custom service. (B) The miRNAs identified to be more abundant in the colon cancer exosomes are depicted in red color on the heatmaps and the less abundant miRNAs are shown in green. Unsorted data are shown on top heatmap and the lower array heatmap image shows the sorted miRNAs identified as significantly, differentially expressed between the two sample sets. This experiment elucidated 79 microRNAs that are capable of stratifying normal serum exosome miRNA from colon cancer serum exosome miRNA samples.
- Peterson MF, Otoc N, Sethi JK, Gupta A, Antes TJ. (2015) Integrated systems for exosome investigation. Methods [Epub ahead of print]. [abstract]