While extracellular vesicles (EVs) are extensively studied by various practical applications in biomedicine, there is still little information on their biomechanical properties due to their nanoscale size. We identified isolated blood plasma vesicles that carried on biomarkers associated with exosomes and exomeres and applied atomic force microscopy (AFM) to study them at single particle level in air and in liquid. Air measurements of exosomes revealed a mechanically indented internal cavity in which highly adhesive sites were located. In contrast, the highly adhesive sites of exomeres were located at the periphery and the observed diameter of the particles was ~35 nm. In liquid, the reversible deformation of the internal cavity of exosomes was observed and a slightly deformed lipid bi-layer was identified. In contrast, exomeres were not deformed and their observed diameter was ~16 nm. The difference in diameters might be associated with a higher sorption of water film in air. The parameters we revealed correlated with the well-known structure and function for exosomes and were observed for exomeres for the first time. Our data provide a new insight into the biomechanical properties of nanoparticles and positioned AFM as an exclusive source of in situ information about their biophysical characteristics.

Cells of two molecular genetic types of breast cancer—hormone-dependent breast cancer (ZR-75 cell line) and triple-negative breast cancer (BT-20 cell line)—were studied using atomic force microscopy and an optical nanomotion detection method. Using the Peak Force QNM and Force Volume AFM modes, we revealed the unique patterns of the dependence of Young’s modulus on the indentation depth for two cancer cell lines that correlate with the features of the spatial organization of the actin cytoskeleton. Within a 200–300 nm layer just under the cell membrane, BT-20 cells are stiffer than ZR-75 cells, whereas in deeper cell regions, Young’s modulus of ZR-75 cells exceeds that of BT-20 cells. Two cancer cell lines also displayed a difference in cell nanomotion dynamics upon exposure to cytochalasin D, a potent actin polymerization inhibitor. The drug strongly modified the nanomotion pattern of BT-20 cells, whereas it had almost no effect on the ZR-75 cells. We are confident that nanomotion monitoring and measurement of the stiffness of cancer cells at various indentation depths deserve further studies to obtain effective predictive parameters for use in clinical practice.

This work studies the impact of the electrostatic interaction between analyte molecules and silver nanoparticles (Ag NPs) on the intensity of surface-enhanced Raman scattering (SERS). For this, we fabricated nanostructured plasmonic films by immobilization of Ag NPs on glass plates and functionalized them by a set of differently charged hydrophilic thiols (sodium 2-mercaptoethyl sulfonate, mercaptopropionic acid, 2-mercaptoethanol, 2-(dimethylamino)ethanethiol hydrochloride, and thiocholine) to vary the surface charge of the SERS substrate. We used two oppositely charged porphyrins, cationic copper(II) tetrakis(4-N-methylpyridyl) porphine (CuTMpyP4) and anionic copper(II) 5,10,15,20-tetrakis(4-sulfonatophenyl)porphine (CuTSPP4), with equal charge value and similar structure as model analytes to probe the SERS signal. Our results indicate that the SERS spectrum intensity strongly, up to complete signal disappearance, correlates with the surface charge of the substrate, which tends to be negative. Using the data obtained and our model SERS system, we analyzed the modification of the Ag surface by different reagents (lithium chloride, polyethylenimine, polyhexamethylene guanidine, and multicharged metal ions). Finally, all those surface modifications were tested using a negatively charged oligonucleotide labeled with Black Hole Quencher dye. Only the addition of copper ions into the analyte solution yielded a good SERS signal. Considering the strong interaction of copper ions with the oligonucleotide molecules, we suppose that inversion of the analyte charge played a key role in this case, instead of a change of charge of the substrate surface. Changing the charge of analytes could be a promising way to get clear SERS spectra of negatively charged molecules on Ag SERS-active supports.