Staff member

Dobryna Julia Valeria Zalvidea

Senior Researcher
Integrative Cell and Tissue Dynamics
+34 934 037 068
Staff member publications

Zalvidea, D., Trepat, X., Rebollo, E., (2019). Fast multiphoton microscope based on polymer ulenses array using a 3D printed mold CL Poster session 2019 Conference on Lasers and Electro-Optics Europe and European Quantum Electronics Conference , IEEE (Munich, Germany) OSA Technical Digest (Optical Society of America, 2019), paper cl_p_22

Multiphoton microscopy is a well-established technique for in vivo tissue imaging at high resolution and long penetration depth. As it uses a single point detector and a single scanning beam, image generation is slow. Several competitive solutions have been proposed during the last years, e.g. resonant scanning, that can reach 30 fps at 2048×2048 scanning resolution but relying on a fixed speed but it does not grant enough time for efficient collection of photons, or multiplexing beams, which show poor efficiency. Here, we propose a multiplexed beam solution that uses a rotating ulenses array for excitation and a camera for detection. The array of ulenses was built in polydimethilsiloxane (PDMS; > 90% transmission in the near IR wavelengths typically used in multiphoton microscopy), using a 3D-printed mold designed following a Nipkow disc pattern slightly modified to increase the lenses efficiency.

Zalvidea, D., Castano, O., Baker, S., Castro, N., Engel, E., Trepat, X., (2019). Time-lapse intravital imaging of biomaterials integration in tissues using a multicolor multiphoton microscope Novel lasers, instruments and technology 2019 Conference on Lasers and Electro-Optics Europe and European Quantum Electronics Conference , IEEE (Munich, Germany) OSA Technical Digest (Optical Society of America, 2019), paper cl_3_1

Different mechanisms are triggered when tissue is exposed to a biomaterial. The success of the biomaterial targeted process, like the release of chemicals, promoted angiogenesis, tissue regeneration, etc. depends on its integration in the tissue [1]. Studying this interaction in vivo requires the ability to image simultaneously deep immersed proteins and biomaterials with high resolution and low damage. Several methods offer solutions but only multiphoton microscopy (MM) has the ability to image with high resolution deep inside the sample. Why is not MM more extensively applied as a platform for investigating biomaterial integration in vivo? The high cost of the typical source for multiphoton microscopy is a clear limitation. Furthermore, imaging several channels simultaneously becomes out of reach for most of the labs.

Timmers, H., Kowligy, A., Lind, A. J., Nader, N., Shaw, J., Zalvidea, D., Biegert, J., Diddams, S. A., (2019). Hyperspectral microscopy with broadband infrared frequency combs Ultrafast Applications (SF1E) 2019 Conference on Lasers and Electro-Optics Europe and European Quantum Electronics Conference , IEEE (Munich, Germany) OSA Technical Digest (Optical Society of America, 2019), paper SF1E.4

We present a new modality for infrared, hyperspectral microscopy using dual-comb, electro-optic sampling of octave-spanning infrared frequency combs. We obtain hyperspectral images of SU8 test patterns on Si wafers with a spatial resolution of 12 µm.

Tekeli, I., Aujard, I., Trepat, X., Jullien, L., Raya, A., Zalvidea, D., (2016). Long-term in vivo single-cell lineage tracing of deep structures using three-photon activation Light: Science and Applications , 5, (6), e16084

Genetic labeling techniques allow for noninvasive lineage tracing of cells in vivo. Two-photon inducible activators provide spatial resolution for superficial cells, but labeling cells located deep within tissues is precluded by scattering of the far-red illumination required for two-photon photolysis. Three-photon illumination has been shown to overcome the limitations of two-photon microscopy for in vivo imaging of deep structures, but whether it can be used for photoactivation remains to be tested. Here we show, both theoretically and experimentally, that three-photon illumination overcomes scattering problems by combining longer wavelength excitation with high uncaging three-photon cross-section molecules. We prospectively labeled heart muscle cells in zebrafish embryos and found permanent labeling in their progeny in adult animals with negligible tissue damage. This technique allows for a noninvasive genetic manipulation in vivo with spatial, temporal and cell-type specificity, and may have wide applicability in experimental biology.

Keywords: Multi-photon microscopy, Photoactivation, Three-photon microscopy, Zebrafish

Casares, L., Vincent, R., Zalvidea, D., Campillo, N., Navajas, D., Arroyo, M., Trepat, X., (2015). Hydraulic fracture during epithelial stretching Nature Materials 14, (3), 343-351

The origin of fracture in epithelial cell sheets subject to stretch is commonly attributed to excess tension in the cells’ cytoskeleton, in the plasma membrane, or in cell–cell contacts. Here, we demonstrate that for a variety of synthetic and physiological hydrogel substrates the formation of epithelial cracks is caused by tissue stretching independently of epithelial tension. We show that the origin of the cracks is hydraulic; they result from a transient pressure build-up in the substrate during stretch and compression manoeuvres. After pressure equilibration, cracks heal readily through actomyosin-dependent mechanisms. The observed phenomenology is captured by the theory of poroelasticity, which predicts the size and healing dynamics of epithelial cracks as a function of the stiffness, geometry and composition of the hydrogel substrate. Our findings demonstrate that epithelial integrity is determined in a tension-independent manner by the coupling between tissue stretching and matrix hydraulics.

Zalvidea, D., Claverol-Tinturé, E., (2011). Second Harmonic Generation for time-resolved monitoring of membrane pore dynamics subserving electroporation of neurons Biomedical Optics Express , 2, (2), 305-314

Electroporation of neurons, i.e. electric-field induced generation of membrane nanopores to facilitate internalization of molecules, is a classic technique used in basic neuroscience research and recently has been proposed as a promising therapeutic strategy in the area of neuro-oncology. To optimize electroporation parameters, optical techniques capable of delivering time and spatially-resolved information on electroporation pore formation at the nanometer scale would be advantageous. For this purpose we describe here a novel optical method based on second harmonic generation (SHG) microscopy. Due to the nonlinear and coherent nature of SHG, the 3D radiation lobes from stained neuronal membranes are sensitive to the spatial distribution of scatterers in the illuminated patch, and in particular to nanopore formation.We used phase-array analysis to computationally study the SHG signal as a function of nanopore size and nanopore population density and confirmed experimentally, in accordance with previous work, the dependence of nanopore properties on membrane location with respect to the electroporation electric field; higher nanopore densities, lasting < 5 milliseconds, are observed at membrane patches perpendicular to the field whereas lower density is observed at partly tangent locations. Differences between near-anode and near-cathode cell poles are also measured, showing higher pore densities at the anodic pole compared to cathodic pole. This technique is promising for the study of nanopore dynamics in neurons and for the optimization of novel electroporation-based therapeutic approaches.