Valeria Caprettini

Spatially, Temporally, and Quantitatively Controlled Delivery of Broad Range of Molecules into Selected Cells through Plasmonic Nanotubes

A Universal plasmonic/microfluidic platform for spatial and temporal controlled intracellular delivery is described. The system can inject/transfect the desired amount of molecules with an efficacy close to 100%. Moreover, it is highly scalable from single cells to large ensembles without administering the molecules to an extracellular bath. The latter enables quantitative control over the amount of injected molecules.

Intracellular and Extracellular Recording of Spontaneous Action Potentials in Mammalian Neurons and Cardiac Cells with 3D Plasmonic Nanoelectrodes

Three-dimensional vertical micro- and nanostructures can enhance the signal quality of multielectrode arrays and promise to become the prime methodology for the investigation of large networks of electrogenic cells. So far, access to the intracellular environment has been obtained via spontaneous poration, electroporation, or by surface functionalization of the micro/nanostructures; however, these methods still suffer from some limitations due to their intrinsic characteristics that limit their widespread use. Here, we demonstrate the ability to continuously record both extracellular and intracellular-like action potentials at each electrode site in spontaneously active mammalian neurons and HL-1 cardiac-derived cells via the combination of vertical nanoelectrodes with plasmonic optoporation. We demonstrate long-term and stable recordings with a very good signal-to-noise ratio. Additionally, plasmonic optoporation does not perturb the spontaneous electrical activity; it permits continuous recording even during the poration process and can regulate extracellular and intracellular contributions by means of partial cellular poration.

Plasmonic meta-electrodes allow intracellular recordings at network level on high-density CMOS-multi-electrode arrays

The ability to monitor electrogenic cells accurately plays a pivotal role in neuroscience, cardiology and cell biology. Despite pioneering research and long-lasting efforts, the existing methods for intracellular recording of action potentials on the large network scale suffer limitations that prevent their widespread use. Here, we introduce the concept of a meta-electrode, a planar porous electrode that mimics the optical and biological behaviour of three-dimensional plasmonic antennas but also preserves the ability to work as an electrode. Its synergistic combination with plasmonic optoacoustic poration allows commercial complementary metal–oxide semiconductor multi-electrode arrays to record intracellular action potentials in large cellular networks. We apply this approach to measure signals from human-induced pluripotent stem cell-derived cardiac cells, rodent primary cardiomyocytes and immortalized cell types and demonstrate the possibility of non-invasively testing a variety of relevant drugs. Due to its robustness and easiness of use, we expect the method will be rapidly adopted by the scientific community and by pharmaceutical companies.A combination of planar porous electrodes, optoacoustic poration and laser scanning allows large-scale recording of intracellular action potentials of cardiomyocyte cultures on CMOS-MEAs.

Cells adhering to 3D vertical nanostructures: cell membrane reshaping without stable internalization

The dynamic interface between the cellular membrane and 3D nanostructures determines biological processes and guides the design of novel biomedical devices. Despite the fact that recent advancements in the fabrication of artificial biointerfaces have yielded an enhanced understanding of this interface, there remain open questions on how the cellular membrane reacts and behaves in the presence of sharp objects on the nanoscale. Here we provide a multifaceted characterization of the cellular membranes mechanical stability when closely interacting with high-aspect-ratio 3D vertical nanostructures, providing strong evidence that vertical nanostructures spontaneously penetrate the cellular membrane to form a steady intracellular coupling only in rare cases and under specific conditions. The cell membrane is able to conform tightly over the majority of structures with various shapes while maintaining its integrity.

Live Intracellular Biorthogonal Imaging by Surface Enhanced Raman Spectroscopy using Alkyne-Silver Nanoparticles Clusters

Live intracellular imaging is a valuable tool in modern diagnostics and pharmacology. Surface Enhanced Raman Spectroscopy (SERS) stands out as a non-destructive and multiplexed technique, but intracellular SERS imaging still suffers from interfering background from endogenous components. Here we show the assembly of small colloidal SERS probes with Raman signal in the cell-silent window of 1800–2900 cm−1 for biorthogonal intracellular SERS imaging of dopamine that was undistinguishable from the endogenous cell background. By linking colloidal silver nanoparticles with alkyne-dopamine adducts, clusters are formed by 2–6 nanoparticles spaced by tight interparticle gaps that exhibited high electric field enhancement and strong SERS signals of alkyne and dopamines. Due to the cell-silent signals of the alkyne, intracellular in-vitro Raman imaging shows that the dopamines on the internalized clusters remain distinguishable across the cytoplasm with good spatial resolution. Our method can be a general-purpose method for real-time imaging of biomolecules, such as proteins, peptides, DNA and drugs.

Author Correction: Live Intracellular Biorthogonal Imaging by Surface Enhanced Raman Spectroscopy using Alkyne-Silver Nanoparticles Clusters

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Author Correction: Plasmonic meta-electrodes allow intracellular recordings at network level on high-density CMOS-multi-electrode arrays

In the version of this Article originally published, the affiliation for the author Francesca Santoro was incorrectly given; it should have been ‘Center for Advanced Biomaterials for Healthcare, Istituto Italiano di Tecnologia, Napoli, Italy’. This has now been corrected in all versions of the Article.

Enhanced Raman Investigation of Cell Membrane and Intracellular Compounds by 3D Plasmonic Nanoelectrode Arrays

Abstract 3D nanostructures are widely exploited in cell cultures for many purposes such as controlled drug delivery, transfection, intracellular sampling, and electrical recording. However, little is known about the interaction of the cells with these substrates, and even less about the effects of electroporation on the cellular membrane and the nuclear envelope. This work exploits 3D plasmonic nanoelectrodes to study, by surface‐enhanced Raman scattering (SERS), the cell membrane dynamics on the nanostructured substrate before, during, and after electroporation. In vitro cultured cells tightly adhere on 3D plasmonic nanoelectrodes precisely in the plasmonic hot spots, making this kind of investigation possible. After electroporation, the cell membrane dynamics are studied by recording the Raman time traces of biomolecules in contact or next to the 3D plasmonic nanoelectrode. During this process, the 3D plasmonic nanoelectrodes are intracellularly coupled, thus enabling the monitoring of different molecular species, including lipids, proteins, and nucleic acids. Scanning electron microscopy cross‐section analysis evidences the possibility of nuclear membrane poration compatible with the reported Raman spectra. These findings may open a new route toward controlled intracellular sampling and intranuclear delivery of genic materials. They also show the possibility of nuclear envelope disruption which may lead to negative side effects.

Cell electroporation and intracellular delivery by mean of 3D vertical hollow nanoelectrodes

In this work we present a device able to electroporate and deliver molecules inside adherent cells using hollow nanoelectrodes that act also as nanofluidic channels. Our fabrication approach allows to greatly reduce the amplitude of the applied electrical pulse train and to selectively deliver molecules only to the targeted cells.

3D plasmonic nanostructures for in-vitro applications in neuroscience and cell biology

In the study of neuronal networks or other cell cultures it is essential to combine information about a single element with the connections developed throughout the whole system. Those data come from different space and time scales, therefore requiring the employment of various techniques to be acquired and combined. Unfortunately, connecting data from more than one experiment is often challenging and the many variables involved make the results difficult to correlate. Here we present a platform able to merge different techniques on the same device, reducing the variability of the system under investigation thanks to the simultaneous study of the single element and the collective behavior of various in-vitro mammalian cell cultures.