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Most of the cancer deaths could be avoided by early detection of the tumor when it is confined to its primary site and it has not metastasized. To this aim, one of the most promising strategies is the discovery and detection of protein biomarkers shed by the young tumor to the bloodstream. Proteomic technologies, mainly mass spectrometry and multiplexed immunoassays, have rapidly developed during last years with improved limits of detection and multiplexing capability. Unfortunately, these developments together major investments and large international efforts have not resulted into new useful protein biomarkers. Here, we analyze the potential and limitations of current proteomic technologies for detecting protein biomarkers released into circulation by the tumor. We find that these technologies can hardly probe the deepest region of the plasma proteome, at concentrations below the pg/mL level, where protein biomarkers for early cancer detection may exist. This clearly indicates the need of incorporating novel ultrasensitive techniques to the proteomic tool-box that can cover the inaccessible regions of the plasma proteome. We here propose biological detectors based on nanomechanical systems for discovery and detection of cancer protein biomarkers in plasma. We review the modes of operation of these devices, putting our focus on recent developments on nanomechanical sandwich immunoassays and nanomechanical spectrometry. The first technique enables reproducible immunodetection of proteins at concentrations well below the pg/mL level, with a limit of detection on the verge of 10 ag/mL. This technology can potentially detect low abundance tumor-associated proteins in plasma at the very early stages of the tumor. The second technique enables the identification of individual intact proteins by two physical coordinates, the mass and stiffness, instead of the mass-to-charge ratio of the protein constituents. This technology enormously simplifies the identification of proteins and it can provide useful information on interactions and posttranslational modifications, that otherwise is lost in mass spectrometry. ; This study has been supported by European Union' s Horizon 2020 research and innovation programme under grant agreement No 731868 − VIRUSCAN; Spanish Ministry of Economy and Competitiveness (grant MAT2015-66904-R) and Spanish Association against Cancer (AECC) Scientific Foundation (grant PS15153089TAMA

Tumorigenesis induces actin cortex remodeling, which makes cancerous cells softer. Cell deformability is largely determined by myosin-driven cortical tension and actin fiber architecture at the cell cortex. However, it is still unclear what the weight of each contribution is, and how these contributions change during cancer development. Moreover, little attention has been paid to the effect of energy metabolism on this phenomenon and its reprogramming in cancer. Here, we perform precise two-dimensional mechanical phenotyping based on power-law rheology to unveil the contributions of myosin II, actin fiber architecture and energy metabolism to the deformability of healthy (MCF-10A), noninvasive cancerous (MCF-7), and metastatic (MDA-MB-231) human breast epithelial cells. Contrary to the perception that the actin cortex is a passive structure that provides mechanical resistance to the cell, we find that this is only true when the actin cortex is activated by metabolic processes. The results show marked differences in the nature of the active processes that build up cell stiffness, namely that healthy cells use ATP-driven actin polymerization whereas metastatic cells use myosin II activity. Noninvasive cancerous cells exhibit an anomalous behavior, as their stiffness is not as affected by the lack of nutrients and ATP, suggesting that energy metabolism reprogramming is used to sustain active processes at the actin cortex. ; This work was supported by the European Union's Horizon 2020 research and innovation program under European Research Council Grant 681275-LIQUIDMASS-ERCCoG-2015, under Grant Agreement No. 731868 – VIRUSCAN and by the Spanish Science, Innovation and Universities Ministry through project CELLTANGLE reference RTI2018-099369-B-I00; by the Comunidad de Madrid (iLUNG B2017/BMD-3884) with support from EU (FEDER, FSE) and Ramón y Cajal grant RYC-2017-21640 to P.M.K. The service from the X-SEM Laboratory is funded by MCIU (CSIC13-4E-1794) and EU (FEDER, FSE).

Ultrasensitive physical, chemical and biological sensors have emerged in the last decade based on the measurement of the eigenfrequencies of micro- and nanosized cantilever plates. Surface stress is omnipresent in these devices due to a variety of factors such as the fabrication process, temperature variations and analyte adsorption. How surface stress influences on the eigenfrequencies of cantilever plates has remained as an unsolved question in physics that has raised a long debate since first experiments in 1975. Recent theoretical models have shed light on the role of the net surface stress. Still, there exists a discrepancy between theory and some experimental reports, affecting to the capability for quantification of these sensors. In this Letter, we present a theoretical framework that demonstrates that the cantilever bending due to differential surface stress between opposite faces of the cantilever, a neglected effect in classical beam theory, plays a relevant role in the stiffness and eigenfrequencies of cantilevers. We develop a new theoretical framework that provides analytical equations that accurately describe the effect of surface stress on the first three vibration modes of cantilevers. Our findings provide the final piece of the puzzle for solving this long-standing problem in physics ; This study has been supported by European Union' s Horizon 2020 research and innovation programme under grant agreement No 731868 – VIRUSCAN and European Research Council grant 681275 – LIQUIDMASS- ERC- CoG-2015; by Spanish Ministry of Economy and Competitiveness (grant MAT2015-66904-R). PMK acknowledges financial support by Fundación General CSIC (ComFuturo Program) ; Peer reviewed

There is a need for noninvasive techniques for simultaneous imaging of the stress and vibration mode shapes of nanomechanical systems in the fields of scanning probe microscopy, nanomechanical biological and chemical sensors and the semiconductor industry. Here we show a novel technique that combines a scanning laser, the beam deflection method and digital multifrequency excitation and analysis for simultaneous imaging of the static out-of-plane displacement and the shape of five vibration modes of nanomechanical systems. The out-of-plane resolution is at least 100 pm Hz−1/2 and the lateral resolution, which is determined by the laser spot size, is 1–1.5 μm. The capability of the technique is demonstrated by imaging the residual surface stress of a microcantilever together with the shape of the first 22 vibration modes. The vibration behavior is compared with rigorous finite element simulations. The technique is suitable for major improvements in the imaging of liquids, such as higher bandwidth and enhanced spatial resolution. ; The authors acknowledge financial support from the Spanish Science Ministry through the projects ANEM! (TEC2009-14517-C02) and INMUNO-swing (IPT-2011-0821-010000), and from the European Union through the project NANOFORCELLS (ERC-StG-2011-278860). ; Peer reviewed

The real-time analysis of single analytes in flow is becoming increasingly relevant in cell biology. In this work, we theoretically predict and experimentally demonstrate hydrodynamic focusing with hollow nanomechanical resonators by using an interferometric system which allows the optical probing of flowing particles and tracking of the fundamental mechanical mode of the resonator. We have characterized the hydrodynamic forces acting on the particles, which will determine their velocity depending on their diameter. By using the parameters simultaneously acquired: frequency shift, velocity and reflectivity, we can unambiguously classify flowing particles in real-time, allowing the measurement of the mass density: 1.35 ± 0.07 g·mL-1 for PMMA and 1.7 ± 0.2 g·mL-1 for silica particles, which perfectly agrees with the nominal values. Once we have tested our technique, MCF-7 human breast adenocarcinoma cells are characterized (1.11 ± 0.08 g·mL-1) with high throughput (300 cells/minute) observing a dependency with their size, opening the door for individual cell cycle studies. ; This work was supported by the European Union's Horizon 2020 research and innovation program under European Research Council Grant 681275-LIQUIDMASS-ERC-CoG-2015, project CELLTANGLE reference RTI2018-099369-B-I00, project MOMPs reference TEC2017-89765-R, Asociación Elena Torres (contract OTR04893) and FERO Foundation (Grant OTR06145). The service from the Micro and Nanofabrication Laboratory (MiNa) and X-SEM Laboratory, is funded by MCIU (CSIC13-4E-1794) and EU (FEDER, FSE). ; Peer reviewed

Resumen del trabajo presentado en la Conferencia Española de Nanofotónica (CEN2021), celebrada de forma virtual del 20 al 22 de septiembre de 2021 ; Vibration modes of biological particles such as proteins, viruses and bacteria involve coherent structural vibrations at frequencies in the THz and GHz domains. These vibration modes carry information on its structure and mechanical properties that play a pivotal role in many relevant biological processes. Despite the rapid advances of optical spectroscopy techniques, detection of vibration modes of single bioparticles has remained elusive. Here we harness a particular regime in the physics of mechanical resonator sensing that serves for detecting them. By depositing single bacterium on the surface of ultra-high frequency optomechanical disk resonators, we demonstrate that the vibration modes of the disk and bacterium hybridize when their associated frequencies are similar (Figure). A general theoretical framework is developed to describe the different regimes that can be found when an analyte adsorbs on a mechanical resonant sensor. The model recovers the classical inertial mass sensing regime as a limit case of a more general problem. Theory, numerical calculations and experiments show excellent agreement, allowing to retrieve the eigenfrequencies and mechanical loss of the bacterium vibration modes. Our method is applied for analysis of the effect of hydration on the vibrational properties of a single bacterium. This work opens the door for a new class of vibrational spectrometry based on the use of high frequency mechanical resonators with the unique capability to obtain information on single biological entities ; This work was supported by the European Union's Horizon 2020 under grants agreement no. 731868 – VIRUSCAN, 681275 – LIQUIDMASS- ERC- CoG-2015 and 770933-NOMLI-ERC-CoG 2017. E. Gil-Santos acknowledge financial support by the Spanish Science and Innovation Ministry through the projects PID2019-109765RA-I00 and RYC2019-026626-I.

The study of biophysical properties of single cells is becoming increasingly relevant in cell biology and pathology. The measurement and tracking of magnitudes such as cell stiffness, morphology, and mass or refractive index have brought otherwise inaccessible knowledge about cell physiology, as well as innovative methods for high-throughput labelfree cell classification. In this work, we present hollow resonator devices based on suspended glass microcapillaries for the simultaneous measurement of single-cell buoyant mass and reflectivity with a throughput of 300 cells/minute. In the experimental methodology presented here, both magnitudes are extracted from the devices' response to a single probe, a focused laser beam that enables simultaneous readout of changes in resonance frequency and reflected optical power of the devices as cells flow within them. Through its application to MCF-7 human breast adenocarcinoma cells and MCF-10A nontumorigenic cells, we demonstrate that this mechano-optical technique can successfully discriminate pathological from healthy cells of the same tissue type. ; This work was supported by the European Union's Horizon 2020 research and innovation program under European Research Council Grant 681275-LIQUIDMASS-ERC-CoG2015 and by the Spanish Science, Innovation and Universities Ministry through project CELLTANGLE reference RTI2018- 099369-B-I00, project MOMPs reference TEC2017-89765-R and Ramon y Cajal grant RYC-2017-21640 to P.M.K.; by the ́ Comunidad de Madrid (iLUNG B2017/BMD-3884) with support from EU (FEDER, FSE). E.G.-S. received funding from Fundacion General CSIC (Programa ComFuturo), as ́ well as Marie-Sklodowska Curie Actions (H2020-MSCA-IF2015) under NOMBIS project (703354).The service from the Micro and Nanofabrication Laboratory (MiNa) and X-SEM Laboratory, is funded by MCIU (CSIC13-4E-1794) and EU (FEDER, FSE). We have received support for part of the publication fee by CSIC open access publication support initiative through URICI. ; Peer reviewed

Free-standing dielectric structures with a substrate underneath, as the SiN microdrums studied here, are the basis of optomechanical devices. Optimization of the optomechanical coupling is demanding in order to enhance the performance of these devices. The optomechanical coupling in the case of drum resonators critically depends on the thickness of the drum, the surface stress, the cavity length and the wavelength. Here, we develop a methodology that combines spatially multiplexed microspectrophotometry, optical modelling of the cavity and finite element method simulation of the mechanical eigenmodes that enables obtaining the properties of the optomechanical cavity and predicting the wavelength that optimizes the optomechanical coupling. By choosing this illumination wavelength we are able to spatially resolve the thermal motion of the microdrums up to the fourth mechanical mode ~ 20 MHz. The presented study opens the door for further optimization pathways for optomechanical detection and actuation. ; This work was supported by the Spanish Science Ministry (MINECO) through project MAT2015-66904-R and by the European Union's Horizon 2020 research and innovation program under grant agreement No 731868–VIRUSCAN. D.R. acknowledges the RyC fellowship supported by MINECO. ; Peer reviewed