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Unveiling Cellular Dynamics Through FluidFM-Mediated Nanoinjection of EVs

von Réka Enz

Precision Delivery at the Nanoscale

In their groundbreaking study, Kovács and colleagues present a novel approach to inject nano structures into individual live cells using robotic fluidic force microscopy (robotic FluidFM) with Cytosurge´s OMNIUM setup (cf. Figure 1a). They focus on nanoinjections of extracellular vesicles (EVs) into different cell types, since EVs play crucial roles in cell-to-cell communication by transporting bioactive molecules such as proteins, nucleic acids, and lipids. However, precise delivery of EV cargo into target cells remains a significant challenge. The authors address this challenge by leveraging the cutting-edge technology linking fluidic force sensing and robotic manipulation at the single cell level.

Figure 1: Photo of the FluidFM appliance, probe and the FluidFM Nanosyringe (a). Schematic illustration of the injection of EVs to the cytoplasm by FluidFM (b).

They present the capability of FluidFM to deliver EVs selectively and efficiently into live cells with high spatial and temporal resolution. By precisely controlling the injection process (injection volume: 100-200 fL), they achieve unprecedented control over the amount (10-100 EVs per cell) and type of cargo delivered into target cells. Furthermore, in this study the biological implications of EV nanoinjection is explored by fluorescence microscopy-based monitoring. This way the group around Kovács provides evidence for downstream effects like intracellular redistribution of the injected structures (cf. Figure 2a), and the encapsulation of the EVs by lipids to form membrane enclosed particles (cf. Figure 2b). The later can be understood as a precursor for the transport to the plasma membrane and a possible release. This assumption is supported by fluorescence images of non‑manipulated cells presenting internalized extracellular vesicles (highlighted by yellow arrows in Figure 2c). Thus, also intercellular transport was detectable after nanoinjection of EVs into selected single cells.

Figure 2: Microscopic images reveal inter- and intra-cellular trafficking of the injected vesicles and confirm viability of the injected cells. Localization and intercellular transfer of GFP positive particles (HEK 293T-PalmGFP cell-derived EV-like particles) 1 h after the nanoinjection (a,b). HeLa cells were injected by applying 40 mbar (a) and 50 mbar (b) pressure. The presence of GFP was validated by immunocytochemistry using unlabelled anti-GFP primary and ATTO550-labelled secondary antibody. DAPI was used for detection of DNA and Cy5- conjugated lactadherin was used for membrane staining. In the donor cells (*) green, and in the acceptor cells yellow arrows point to GFP containing EV-like particles in large (approx. 0.5-1 µm) membrane closed vesicles, and red the arrow shows similar DNA containing large vesicle (c).

The publication of Kovács et al. presents a significant advancement in the field of nanomedicine and cellular biology, introducing the FluidFM OMNIUM as a powerful tool for investigating the intricate mechanisms of intercellular communication and potentially revolutionizing therapeutic approaches reliant on EV-based therapies. The integration of robotic manipulation and fluidic force sensing in the OMNIUM not only enables precise nanoinjection but also opens avenues for further research into cellular dynamics and therapeutic interventions at the nanoscale.



Kovács, et al.. "Nanoinjection of extracellular vesicles to single live cells by robotic fluidic force microscopy." J. Extracell Vesicles, 2023, 12, 12388.