Raman tweezers microspectroscopy of circa 100 nm extracellular vesicles

The technique of Raman tweezers microspectroscopy (RTM) for the global biomolecular content characterization of a single extracellular vesicle (EV) or a small number of EVs or other nanoscale bioparticles in an aqueous dispersion in the difficult-to-access size range of near 100 nm is described in detail. Researchers from the CNRS demonstrate the particularities and potential of RTM using the examples of DOPC liposomes, exosomes from human urine and rat hepatocytes, and a mixed sample of the transfection reagent FuGENE in diluted DNA solution. The approach of biomolecular component analysis for the estimation of the main biomolecular contributions (proteins, lipids, nucleic acids, carotenoids, etc.) is proposed and discussed. Direct Raman evidence for strong intra-sample biomolecular heterogeneity of individual optically trapped EVs, due to variable contributions from nucleic acids and carotenoids in some preparations, is reported. On the basis of the results obtained, the researchers are making an attempt to convince the scientific community that RTM is a promising method of single-EV research; to their knowledge, it is the only technique available at the moment that provides unique information about the global biomolecular composition of a single vesicle or a small number of vesicles, thus being capable of unravelling the high diversity of EV subpopulations, which is one of the most significant urgent challenges to overcome. Possible RTM applications include, among others, searching for DNA biomarkers, cancer diagnosis, and discrimination between different subpopulations of EVs, lipid bodies, protein aggregates and viruses.

Normalized Raman signal of the highest-intensity saturation plateau, within one experimental run of 10 min, versus particle concentration, for DOPC liposomes “50 nm” (a), “100 nm” (b), “200 nm” (c), and urinal exosomes (d). Dashed curves through the data points were drawn to highlight the tendencies.