The main goal of this proposal is to induce and detect in real time and at single-cell level the differentiation of human mesenchymal stem cells using mechanical loads in 2D and 3D conditions. This proposal integrates disciplines from nanotechnology, bioengineering and cell and molecular biology. First we propose to develop a method to track the differentiation of stem cells in real time and at the single cell level using cytoskeletal organization of actin, microtubules and intermediate filaments as a suitable cell biomarker. We will then establish mechanical loading protocols to induce, via direct force application onto cells, the first stages of stem cell differentiation towards specific cell lineages. We will apply cyclic tensile strain and compression to stem cells in 2D and 3D conditions, and track their differentiation status in real time using the cytoskeletal biomarkers that we will have identified before. The results of this proposal will have implications for the field of stem cell mechanobiology in particular, and some of the techniques developed will also contribute to the wider field of directed stem cell differentiation.

Due to the promising potential of evanescent optical waves and surface plasmon polaritons to successfully merge current photonic and electronic technology on the nanoscale, they are generally envisioned as the information carriers of the future. This, however, requires the advanced miniaturization of integrated optical circuitry, which demands great effort not only in fabrication and design, but even more so in the development of accurate control and, above all, a deepest scientific understanding of sub-wavelength-confined light. At present, the thorough understanding of the nanoscale behaviour of evanescent fields and the details of the underlying light-matter interactions are missing, but key elements in modern optoelectronics research. TRUEViEW aims to provide the currently missing fundamental knowledge, by implementing innovative electron imaging techniques as the ultimate tools to directly visualize and characterize photonic and plasmonic nanostructures in both space and time with nanometer and femtosecond resolution. The project takes a bottom-up approach throughout, yielding a systematic and consistent route towards unravelling the working principles of nanoscale-confined optical waves and gaining practical expertise on the manipulation of light in optoelectronic nanostructures. At the same time, the project will pioneer and establish the field of ultrafast electron microscopy in the European research community, as well as newly introducing the novel technique of in-situ Photon-Induced Near-field Electron Microscopy. On the whole, TRUEViEW will lay the groundwork for a myriad of future optoelectronic applications, which include - among many others - sub-wavelength optics, light generation and data storage, nanolithography, quantum computing, quantum cryptography, biophysical spectroscopy and nanosensing. As such, the project will have a strong impact not only all across the scientific board, but also in the European commercial and technological industry.

This project aims to design the first ever prototype of a plasmonic microscope for in vivo bio imaging. The principle behind the prototype consists in deep sub-wavelength focusing and raster scanning of multiple points to achieve imaging. On the one hand, using surface plasmons excitation for microscopy instead of light offers the advantage of ultra short plasmonic wavelengths (down to 100 nm) for visible light frequencies, enabling a plasmonic diffraction limit of 50 nm which sets the resolution of the microscopy. On the other hand, in this ultra short wavelength regime surface plasmons suffer losses, limiting the propagation length of plasmon waves. Losses limit the image size (field of view) to no more than 10 by 10 resolution points, a size which is completely insufficient for biological samples. The main scientific challenge of this proposal is to surpass the plasmonic losses which constitute a limitation for microscopy and most plasmonic applications. While previous attempts were based on reducing the losses (succeeded up by a factor of two), we propose a scheme that is not sensitive to these losses. The scheme consists in a 'network' of periodic plasmonic repeaters that regenerate the lossy signals, similarly to the standard method used for distributing cellular phones and TV/radio signals over long distances. In particular, we will use a Spatial Light Modulator (SLM) to create and scan multiple plasmonic foci in parallel. The image is acquired via raster scanning of all the plasmonic foci in parallel, yielding an image size limited only by the extension of the network, namely the number of pixels in the SLM. Moreover, this scheme also reduces the scanning time by up to two orders of magnitudes, making it suitable for in-vivo measurements. In conclusion, we propose a technological advancement for microscopy based on a novel scheme that can harvest the short plasmonic wavelengths for microscopy without compromising any other relevant parameters.

Electronic nanodevices have demonstrated to be versatile and effective tools for the investigation of exotic quantum phenomena under controlled and adjustable conditions. Yet, these have enabled to give access to the manipulation of charge flow with unprecedented precision. On the other hand, the wisdom dealing with control, measurements, storage, and conversion of heat in nanoscale devices, the so-called 'caloritronics' (from the Latin word 'calor', i.e., heat), despite a number of recent advances is still at its infancy. Although coherence often plays a crucial role in determining the functionalities of nanoelectronic devices very little is known of its role in caloritronics. In such a context, coherent control of heat seems at present still very far from reach, and devising methods to phase-coherently manipulate the thermal current would represent a crucial breakthrough which could open the door to unprecedented possibilities in several fields of science. Here we propose an original approach to set the experimental ground for the investigation and implementation of a new branch of science, the 'coherent caloritronics', which will take advantage of quantum circuits to phase-coherently manipulate and control the heat current in solid-state nanostructures. To tackle this challenging task our approach will follow three main separate approaches, i.e., the coherent control of heat transported by electrons in Josephson nanocircuits, the coherent manipulation of heat carried by electrons and exchanged between electrons and lattice phonons in superconducting proximity systems, and finally, the control of the heat exchanged between electrons and photons by coherently tuning the coupling with the electromagnetic environment. We will integrate superconductors with normal-metal or semiconductor electrodes thus exploring new device concepts such as heat transistors, heat diodes, heat splitters, where thermal flux control is achieved thanks to the use of the quantum phase.

The invention of new microscopy tools keeps transforming many areas of major economic importance. One example is the semiconductor industry, where sophisticated imaging tools are crucial for maintaining profitable high-volume manufacturing. State of the art tools are routinely used to characterize the dimensions and electrical functionality of manufactured devices, however, as device dimensions continue to reduce there is a perpetual need for the invention of new probing tools that could cope with the new challenges. The goal of this proposal is to make a Proof of Concept (POC) for a new type of nano detector and imaging apparatus, which addresses the future performance challenges for microscopy tools in the semiconductor, solar cell and novel materials industries. The nano detector is a spinoff from a unique nano-assembly technology developed in our ERC Starters project, capable of making the most advanced carbon-nanotube-based electronic devices to date. Based on this new nano detector we intend to demonstrate an imaging apparatus that enables ultra-sensitive imaging of electrical potentials on the nanoscale. The technology is foreseen to improve the current state of the art by orders of magnitudes. We will demonstrate detectors that operate at room temperature, work at high frequencies, are fast, accurate and inexpensive to manufacture - all desirable characteristics for rapid market adoption. In addition to the technological POC, the project comprises innovation protection tasks, the delivery of a relevant business model and networking actions for successful commercialization. The novel detectors would make a vast impact on an important productive sector in Europe. They will contribute especially to yield learning - necessary for maintaining the industry profit margins and competitiveness. The detectors also answer to the current nanoscale imaging challenges in most Key Enabling Technology fields in Europe.

In this work, I propose the use of a nano/microfluidic system to detect biomarkers relevant to Alzheimer's disease (AD). The use of this system could give earlier and more accurate diagnoses, as well as provide the opportunity for therapeutic interventions and effective disease monitoring. Prior to the diagnosis of dementia and even before the appearance of plaques and tangles, it is suspected that biochemical changes have begun to occur in the brain that eventually lead to AD. Due to the brain being a particularly difficult organ to access, the search for biomarkers has focused primarily on cerebrospinal fluid (CSF) and blood. Because the available sample and biomarker levels are low, clinical tools that can measure candidate biomarkers for reproducible detection are currently insufficient. Here we propose a nanofluidic system with well-defined surface chemistry to greatly improve the sensitivity, selectivity and reproducibility in detection of biomarkers found in CSF. Our device will be fabricated using standard silicon microfabrication procedures to produce highly controlled channels for sample handling and multiplex detection. By working under continuous-flow conditions rather than in batch format, we eliminate variability due to mass transport limitations while allowing for improved standardization. Concentrating the antibodies to a discrete area will increase selectivity and allow amyloid-beta quantitation in spiked serum and real CSF of normal and Alzheimer's donor samples to be achieved. I believe that development of a multiplex diagnostic tool will offer many more people access to better disease diagnosis management and assist in the search for better therapeutics that not only slow disease progression but potentially reverse it.

Staphylococcus aureus community-acquired (CA)-MRSA strains are highly virulent and can cause infections in otherwise healthy individuals and are a leading cause of death worldwide. Innate immunity is our primary defense against invading staphylococci. Blood-neutrophils migrate to the site of infection where they, in concert with the complement system, engulf and kill bacteria in a process called phagocytosis. Especially CA-MRSA strains seem to be very efficient in circumventing this neutrophil killing. Interestingly, only a relative small number of virulence factors have been associated with CA-MRSA, one of which are the phenol soluble modulins (PSMs). In vivo models of experimental infection with PSM-mutants have shown a critical role for PSMs in skin and soft tissue infections. PSMs are small alpha-helical peptides which have two distinct functions on the immune system, in vitro PSMs can attract neutrophils in the nanomolar range, whereas in the micromolar range they are cytolytic for neutrophils. Recent publications suggests that these two functions complement each other for full staphylococcal virulence, although it seems counter-intuitive for S. aureus to actively attract its mortal enemy: the neutrophil. To make matters even more complicated PSMs are functionally inactivated by host serum lipoproteins, most efficiently by high density lipoprotein (HDL). The goal of this research proposal is to determine the mechanism of action for PSMs in staphylococcal disease. To this end, I will use genomic and proteomic approaches combined with cutting edge in vivo spinning-disk confocal microscopy, to dissect the functions of PSMs in host-staphylococcal interactions in; 1) neutrophil recruitment, 2) neutrophil lysis 3) HDL neutralization and liver pathology, 4) Evasion of PSM-recognition by FLIPR-L

The detection of specific sequences of DNA is pivotal in the diagnosis of a number of infectious diseases, such as chlamydia. It is also important in detecting and identifying potential bio-warfare agents. Current methods of DNA detection typically involve laboratory based sample analysis, which is slow and costly. This proposal seeks to address these drawbacks by developing an alternative approach based on nanopore unzipping. In this method the current-time transient is recorded for a single molecule of dsDNA as it denatures and passes through a protein pore. The current as the DNA blocks the pore, as well as the time taken for the dsDNA to 'unzip' (denature) is characteristic of the DNA sequence, length and the presence of any modifications. The proiten pore itself must be suspended in a lipid bilayer across a nm-diameter orifice that acts as a platform. Whilst identification of DNA targets with nanopore unzipping is expected to be quick and efficient, the platforms typically used for such measurements (mainly silicates) are fragile and lack bio-compatibility. This proposal seeks to bring the power of nanopore unzipping outside of the laboratory through the development of a robust, ultra-stable, platform constructed from diamond. The research conducted during this proposal could lead to a commercially viable device capable of detecting and identifying DNA at the point need, for example in medical diagnostics in a hospital or identifying potential bio-warfare agents in the field of operation.

BENGRAS is a Marie Curie IIF project that focuses on multidisciplinary transfer of knowledge from a promising Australian early career researcher to KU Leuven towards the design and synthesis of novel functional nano-materials and the development of advanced analytical techniques for material analysis. The project will study bandgap engineering in graphene induced by physi- and chemi- sorption of self-assembled molecular monolayers, which is an interdisciplinary research topic centered at the interface between materials science, supramolecular chemistry, nanoscience and physics. Graphene, a material consisting of flat one-atom-thick sheets of carbon atoms has enormous potential for the use in electronic transistors because of the unique electronic properties and the reduced dimensionality. Graphene is a ‘zero-gap’ semiconductor and to unlock its electronic properties two basic requirements must be satisfied. Firstly, precise control over electronic band structure (bandgap) is needed. This can be achieved by adsorbing atoms and molecules (e.g. H, OH, K, NH3) on its surface thus generating local mid-gap states. Secondly, the means to control the degree of ordering and periodicity of modified graphene layers are to be derived. In other words, the regions where bandgap can be locally tuned have to be extended to a micron scale for practical applications. At present, this issue remains largely unexplored. This project will investigate the electronic structure of graphene the surface of which has been nano-patterned by physisorped (i.e. weak surface interactions) and covalent (i.e. strong surface interactions) molecular monolayers. Through BENGRAS the fellow will contribute extensive expertise in carbonaceous materials and spectroscopy towards controlled modification of electronic properties of graphene and, designing appropriate analytical methods for the study of low-dimensional materials using optical spectroscopy methods at the nanoscale.

Research neuronal signaling is the subject of a very large community, but progresses face a dense multi-scale dynamics involving signaling at the molecular, cellular and large neuronal network levels. Whereas the brain capabilities are most likely emerging from large neuronal networks, available electrophysiological methods limit our access to single cells and typically provides only a fragmented observation, on limited spatial/temporal scales. Therefore, broadening the spectrum of scales for observing neuronal signaling within large neuronal networks is a major challenge that can revolutionize our capability of studying the brain and its physio-pathological functions, as well as of deriving bio-inspired concepts to implement artificial system based on neuronal circuits. We propose the development of an innovative electro-plasmonic multifunctional platform that by combining different methodologies emerging from distant fields of Science and Technology will provide a radically new path for real time neurointerfacing at different scale levels: 1. The molecular scale: 3D plasmonic nanoantennas will give access to information at molecular level by means of enhanced spectroscopies with particular regard of time resolved Raman scattering. 2. The single-neuron scale within neuronal networks: by both in-cell and extra-cell couplings with 3D nanostructures which work at the same time as plasmonic antennas and CMOS 3D nanoelectrodes. 3. The scale of large neuronal networks: by CMOS high-density electrode arrays for spatially and temporally resolving neuronal signaling form thousands of measuring sites. This is achieved by exploiting an innovative nanofabrication method able to realize 3D nanostructures which can work at the same time as plasmonic nanoantennas and as nanoelectrodes. These structures will be integrated on CMOS multi-electrode arrays designed to manage multiscale measurements from the molecular level up to network level on several thousand of measurement sites.