That being said, protein arrays are expensive, there’s quantitation bias and, again, you’re only able to ID known markers.įinally, there’s mass spectrometry which is the most comprehensive approach. Protein arrays, however, have the advantage of being scalable with low to mid-throughput and there are many commercially available panels. However, if you’re involved in biomarker discovery then you should keep in mind that the use of western blotting is pretty limiting it’s low-throughput and you can only assess the IDs of known markers. So if you know which protein you’re targeting, western blotting is great. Western blotting is easy to perform and there are plenty of antibodies available. The current approaches used to characterize the proteins in EVs include western blotting, protein arrays, and mass spectrometry. They include cytoskeletal proteins, enzymes, and signal transduction proteins among many others (Diagram 3.5). Just as well, the way one treats the sample, the processing, and the isolation all of these factors are going to affect the final make-up and composition of the exosomal RNA and thus the entire process of biomarker discovery.Īs for proteins, there’s a wide range of them within exosomes as well. Thus, the number of samples that one uses to try to assess or come up with a novel biomarker is very important. We found that, just as before, the composition of exosomal RNA can vary enormously between different types of samples (Diagram 3.4).ĭiagram 3.4: Class Make-up of Exosomal RNAs in Body Fluids. Yet there was plenty of intraindividual variabilities as well (Diagram 3.3).ĭiagram 3.3: Class Make-up of Exosomal RNAs.Īt SBI we generated some interesting data using our Exo-NGS service with respect to all of this. For instance, the serum sample was far richer in microRNAs than the urine sample. There was a clear distinction between the two sample types. At the same time, we need to be sure that we’re identifying the RNA found within exosomes instead of other possibilities, like freely circulating RNA.Īdditionally, it’s important to take into account that the make-up of exosomal RNAs is going to vary a lot between individuals and between sample types.Īs a case in point: in one paper, exosomal RNA was assessed from serum and urine samples. One of the challenges here is that concentrations are going to be very low. The two main components of EVs are RNAs and proteins we’ll start with the challenges surrounding the former and then move on to those involving the latter.Įxosomal RNA is mainly comprised of species between 20-200 nt including microRNAs, lncRNAs, mRNAs, and so on (Diagram 3.2).ĭiagram 3.2: Exosomal RNA Challenges: Concentration & Composition. Custom miRNA Precursor and anti-miRNA (miRZip) ServicesĪs you can imagine given the aforementioned discussion, there are numerous challenges with respect to the use of EVs for biomarker discovery.Custom Luciferase-labeled Stable Cell Lines.Custom Overexpression Stable Cell Lines.
Exosome Lipidomics & Metabolomics Services.Cumate Inducible Gene Expression Systems.Lentiviral Expression Plasmids & Lentiviral Vectors.LentiSuite & LentiStarter High-Titer Lentivirus Kits.CLOuD9 Gene Expression Regulation System.
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