High-accuracy adaptive modeling of the energy distribution of a meniscus-shaped cell culture in a Petri dish

Cylindrical Petri dishes embedded in a rectangular waveguide and exposed to a polarized electromagnetic wave are often used to grow cell cultures. To guarantee the success of these cultures, it is necessary to enforce that the specific absorption rate distribution is sufficiently high and uniform over the Petri dish. Accurate numerical simulations are needed to design such systems. These simulations constitute a challenge due to the strong discontinuity of electromagnetic material properties involved, the relative low field value within the dish cultures compared with the rest of the domain, and the presence of the meniscus shape developed at the liquid boundary. The latter greatly increases the level of complexity of the model in terms of geometry and intensity of the gradients/singularities of the field solution. In here, we employ a three-dimensional (3D) hp-adaptive finite element method using isoparametric elements to obtain highly accurate simulations. We analyze the impact of the geometrical modeling of the meniscus shape cell culture in the hp-adaptivity. Numerical results showing the error convergence history indicate the numerical difficulties arisen due to the presence of a meniscus-shaped object. At the same time, the resulting energy distribution shows that to consider such meniscus shape is essential to guarantee the success of the cell culture from the biological point of view

A summary of my twenty years of research according to Google Scholars

I am David Pardo, a researcher from Spain working mainly on numerical analysis applied to geophysics. I am 40 years old, and over a decade ago, I realized that my performance as a researcher was mainly evaluated based on a number called \h-index". This single number contains simultaneously information about the number of publications and received citations. However, dif- ferent h-indices associated to my name appeared in di erent webpages. A quick search allowed me to nd the most convenient (largest) h-index in my case. It corresponded to Google Scholars. In this work, I naively analyze a few curious facts I found about my Google Scholars and, at the same time, this manuscript serves as an experiment to see if it may serve to increase my Google Scholars h-index

A summary of my twenty years of research according to Google Scholars

I am David Pardo, a researcher from Spain working mainly on numerical analysis applied to geophysics. I am 40 years old, and over a decade ago, I realized that my performance as a researcher was mainly evaluated based on a number called \h-index". This single number contains simultaneously information about the number of publications and received citations. However, dif- ferent h-indices associated to my name appeared in di erent webpages. A quick search allowed me to nd the most convenient (largest) h-index in my case. It corresponded to Google Scholars. In this work, I naively analyze a few curious facts I found about my Google Scholars and, at the same time, this manuscript serves as an experiment to see if it may serve to increase my Google Scholars h-index

Microdevices and Microsystems for Cell Manipulation

Microfabricated devices and systems capable of micromanipulation are well-suited for the manipulation of cells. These technologies are capable of a variety of functions, including cell trapping, cell sorting, cell culturing, and cell surgery, often at single-cell or sub-cellular resolution. These functionalities are achieved through a variety of mechanisms, including mechanical, electrical, magnetic, optical, and thermal forces. The operations that these microdevices and microsystems enable are relevant to many areas of biomedical research, including tissue engineering, cellular therapeutics, drug discovery, and diagnostics. This Special Issue will highlight recent advances in the field of cellular manipulation. Technologies capable of parallel single-cell manipulation are of special interest

Numerical modelling of electrical stimulation for cartilage tissue engineering

In this thesis, the design and validity of numerical models of electrical stimulation for cartilage tissue engineering are critically assessed at different scales. In sum, the results of this thesis pave the way for experimentally validated numerical models of electrical stimulation devices for cartilage tissue engineering. Furthermore, models of tissue samples can be developed down to the cellular scale and will contribute to the development of patient-specific stimulation approaches

Autonomous capillary systems for life science research and medical diagnostics

In autonomous capillary systems (CS) minute amounts of liquid are transported owing to capillary forces. Such CSs are appealing due to their portability, flexibility, and the exceptional physical behavior of liquids in micrometer sized microchannels, in particular, capillarity and short diffusion times. CSs have shown to be a promising technology for miniaturized immunoassays in life science research and diagnostics. Building on existing experimental demonstrations of immunoassays in CSs, a theoretical model of such immunoassays is implemented, tools and CSs for performing immunoassays are developed, key functional elements of CSs such as capillary pumps and valves are explored experimentally, and a proof-of-concept of the ultimate goal of one-step immunoassays are given in this work. For the theoretical modeling of immunoassays in CSs a finite difference algorithm is applied to delineate the role of the transport of analyte molecules in the microchannel (convection and diffusion), the kinetics of binding between the analyte and the capture antibodies, and the surface density of the capture antibody on the assay. The model shows that assays can be greatly optimized by varying the flow velocity of the solution of analyte in the microchannels. The model also shows how much the analyte-antibody binding constant and the surface density of the capture antibodies influence the performance of the assay. We derive strategies to optimize assays toward maximal sensitivity, minimal sample volume requirement or fast performance. A method using evaporation for controlling the flow rate in CSs was developed for maximum flexibility for developing assays. The method allows to use small CSs that initially are filled by capillary forces and then provide a well defined area of the liquid-air interface from which liquid can evaporate. Temperature and humidity are continuously measured and Peltier-elements are used to adjust the temperatures in multiple areas of the CSs relative to the dew-point. Thereby flow rates in the range from ~1.2 nL s−1 to ~30 pL s−1 could be achieved in the microchannels. This method was then used for screening cells for surface receptors. CSs, that do not need any peripherals for controlling flow rates become even more appealing. We explored the filling behavior of such CSs having microchannels of various length and large capillary pumps. The capillary pumps comprise microstructures of various sizes and shapes, which are spaced to encode certain capillary pressures. The spacing and shape of the microstructures is also used to orient the filling front to obtain a reliable filling behavior and to minimize the risk of entrapping air. We show how two capillary pumps having different hydrodynamic properties can be connected to program a sequence of slow and fast flow rates in CSs. Liquid filling CSs can hardly be stopped, but in some cases it might be beneficial to do so. In a separate chapter we explore how microstructures need to be designed to use capillary forces to stop, time, or trigger liquids. Besides well-defined flow rates in CSs accurately patterned capture antibodies (cAbs) are key for performing high-sensitive surface immunoassays in CSs. We present a method compatible with mass fabrication for patterning cAbs in dense lines of up to 8 lines per millimeter. These cAbs are used with CSs that are optimized for convenient handling, pipetting of solutions, pumping of liquids such as human serum, and visualization of signals for fluorescence immunoassays to detect c-reactive protein (CRP) with a sensitivity of 0.9 ng mL−1 (7.8 pM) from 1 uL of CRP-spiked human serum, within 11 minutes, with 4 pipetting steps, and a total volume of sample and reagents of <1.5 uL. CSs for diagnostic applications have different requirements than CSs that are used as a research tool in life sciences, where a high flexibility and performance primes over the ease of use and portability of the CSs. We give a proof-of-concept for one-step immunoassays based on CSs which we think can be the base for developing portable diagnostics for point-of-care applications. All reagents are preloaded in the CSs. A sample loaded in the CSs redissolves and reconstitutes the detection antibodies (dAbs), analyte-dAb-complexes are formed and detected downstream in the CSs. A user only needs to load a sample and measure the result using a fluorescence microscope or scanner. C-reactive protein was detected in human serum at clinical concentrations within 10 minutes and using only 2 uL of sample

3D Bioprinting Hydrogel for Tissue Engineering an Ascending Aortic Scaffold

The gold standard in 2016 for thoracic aortic grafts is Dacron®, polyethylene terephthalate, due to the durability over time, the low immune response elicited and the propensity for endothelialization of the graft lumen over time. These synthetic grafts provide reliable materials that show remarkable long term patency. Despite the acceptable performance of Dacron® grafts, it is noted that autographs still outperform other types of vascular grafts when available due to recognition of the host\u27s cells and adaptive mechanical properties of a living graft. 3-D bioprinting patient-specific scaffolds for tissue engineering (TE) brings the benefits of non-degrading synthetic grafts and autologous grafts together by constructing a synthetic scaffold that supports cell infiltration, adhesion, and development in order to promote the cells to build the native extracellular matrix in response to biochemical and physical cues. Using the BioBots 3-D bioprinter, scaffold materials we tested non-Newtonian photosensitive hydrogel that formed a crosslinked matrix under 365 nm UV light with appropriate water content and mechanical properties for cell infiltration and adhesion to the bioprinted scaffold. Viscometry data on the PEGDA-HPMC 15%-2% w/v hydrogel (non-Newtonian behavior) informed CFD simulation of the extrusion system in order to exact the pressure-flow rate relationship for every hydrogel and geometry combination. Surface tension data and mechanical properties were obtained from material testing and provide content to further characterize each hydrogel and resulting crosslinked scaffold. The goal of this work was to create a basis to build a database of hydrogels with corresponding print settings and resulting mechanical properties in order to progress the field of tissue engineered vascular grafts fabricated by nozzle-based rapid prototyping

DEVELOPMENT OF HUMAN TISSUES ON A CHIP FROM PLURIPOTENT STEM CELLS

The development of new microscale technologies aimed at generating human tissues and organs on a chip is emerging as a novel effective strategy to perform cost effective and multi-parametric assays for disease modeling investigations and screening specific therapeutic strategies, as broadly recognized from the scientific community, the major pharmaceutical companies and the governmental agencies. The generation of human organs on a chip, in which microscale technologies, such as microfluidics, are combined with cultured human cells in order to mimic whole living organ environment, offers a unique opportunity to study human physiology and pathophysiology, resembling in vivo conditions. This technological perspective could provide an effective solution to the current limitations of animal models, which result highly expensive and non predictive of human physiology, and conventional cell culture models that fail to recapitulate complex, organ-level disease processes. The possibility of developing direct organogenesis on a chip from human pluripotent stem cells, which have the potential to generate all cellular types of the human body, could overcome the limited availability of human primary cells, such as human hepatocytes or cardiomyocytes. Moreover, the recent generation of human induced pluripotent stem cells (hiPSCs) from adult somatic cells through the ectopic expression of defined transcription factors, provides a new effective tool to obtain populations of patient-specific stem cells with distinctive genetic diversity. This work is aimed at the development of functional human tissues on a chip from pluripotent stem cells, to be used in developing new in vitro disease models or drug screening and toxicity assays, more predictive than current animal models, overcoming issues related to primary cell sources recruitment. To this purpose, an integrated microscale system, based on the microfluidic technology, was developed, in order to derive functional cardiac and hepatic tissues on a chip, from human pluripotent stem cells, through the accurate delivery of chemical compounds and growth factors, within cellular microenvironment, and proper regulation of exogenous and endogenous factors, which are known to affect embryonic development in vivo. These functionally differentiated cells on a chip can be directly used for multi-parametric and large-scale drug screening or for developing micro-engineered human organ models. This last aspect also requires to design new technology for assisting in vitro assays and developing new therapeutic strategies or for screening among potential clinical cures. In particular, design of proper functional assays to test cellular responses to drugs or external stimuli, such as mechanical stress or hypoxia, in a physiologic or pathologic context has been addressed. Thanks to the development of a microfluidic gas exchanger for generating rapid depletion of oxygen content in culture medium, the role of acute hypoxia in calcium handling machinery of a cardiomyocytes population was investigated, mimicking early effects of ischemia on cardiac microenvironment. Moreover, the role of cyclic mechanical stress, which plays a crucial role in the investigation of new physiological and pathological responses to cell culture microenvironment, was analyzed in a human Duchenne Muscular Dystrophy (DMD) in vitro model, through a microfluidic-based cell stretching device accurately reproducing cyclic mechanical deformations along time. The combination of tissues development on a chip and micro-technology assisting for functional assay on a chip opens new and substantial perespective in generating a human in vitro model that properly resemble the physiological and pathophysiological behaviro of a tissues or organs within humna body

Focussed MeV-Ion Micro- and Nano-Beams in the Life Sciences: Selected Applications

This work presents the development of a sub-micron nuclear microprobe for applications in the life sciences. It includes quantitative trace element analysis with sub-micron spatial resolution, 2D- and 3D-microscopy of density distributions and the targeted irradiation of living cells with counted single ions. The analytical methods base on particle induced X-ray emission spectrometry (PIXE), Rutherford backscattering spectrometry (RBS), scanning transmission ion microscopy (STIM) and STIM-tomography. The specific development of the existing nuclear microprobe LIPSION led to an improved performance of the capabilities for trace element analysis. For sub-micron analysis the spatial resolution could be improved to 300 nm at a sensitivity of about 1 µg/g for metal ions in biological matrices; for a resolution of 1 µm the sensitivity was improved to 200 ng/g (3 µmol/l). This habilitation thesis comprises a short general introduction including the motivation to utilize focussed high energy ion beams, an overview on the applications and actual research fields. The introduction is followed by the basic principles of the equipments and analytical methods. An estimation of the limits of resolution for element analytical and single ion techniques is given for the Leipzig system. Thereafter, selected studies from different research areas are presented. The first presented application is a study from environmental air pollution research. It is demonstrated that the microscopic elemental analysis of single aerosol particles can be used to assess the contributions from different sources. A further example is the analysis of the distribution of nanoparticles in skin cross-sections for a risk assessment of the applications of nanosized physical UV-filters in cosmetic products. The risk assessment is followed by the micro-analysis of trace elements, especially of bound metal ions, in brain sections on the cellular and sub-cellular level. After this the application of focussed MeV ion beams in low dose radiobiological research is presented. Finally, the analysis of 3D-density distributions by proton micro-tomography is demonstrated. A summary concludes on the applications and gives an outlook to further applications and methodological developments. The appendix comprises the relevant publications of the author.Die vorliegende Arbeit etabliert für Anwendungen in den Lebenswissenschaften den Einsatz hochfokussierter MeV-Ionenstrahlen für nuklear-mikroskopische Methoden der quantitativen Spurenelementanalyse, der 2D- und 3D-Dichtemikroskopie sowie für die gezielte Bestrahlung einzelner lebender Zellen für radiobiologische Experimente. Zur Anwendung kamen die Methoden ortsaufgelöste Protonen induzierte Röntgenfluoreszenzanalyse (particle induced X-ray emission - PIXE), Spektrometrie rückgestreuter Ionen (Rutherford backscattering spectrometry - RBS) und Rastertransmissionsionenmikroskopie (scanning transmission ion microscopy - STIM). Durch eine gezielte Weiterentwicklung des bestehenden Ionenstrahlmikroskops, der Hochenergie Ionennanosonde LIPSION, konnte die Ortsauflösung für Spurenelementanalyse auf unter 300 nm verbessert werden, beziehungsweise die Sensitivität für Metallionen in biologischen Proben auf unter 200 ng/g (3 µmol/l) bei einer Ortsauflösung von 1 µm verbessert werden. Die Habilitationsschrift umfasst eine kurze allgemeine Einleitung einschließlich der Motivation für den Einsatz fokussierter MeV-Ionenstrahlen sowie einen Überblick über die Anwendungsgebiete und aktuellen Forschungsschwerpunkte. Danach werden kurz die Grundlagen der Technik und Methoden vorgestellt, gefolgt von einer Abschätzung der A\u7fuflösungsgrenzen für Elementanalysen und Einzelionentechniken. Danach werden ausgewählte Anwendungen aus verschiedenen Forschungsgebieten vorgestellt. Das erstes Beispiel ist aus der Umweltforschung. Es wird dargestellt, wie mittels ortsaufgelöster Elementspektroskopie eine Abschätzung der Feinstaubbelastung nach Beiträgen einzelner Verursacherquellen erfolgen kann. Dann folgt als Beispiel eine ortsaufgelöste Analyse der Verteilung von Nanopartikeln aus Sonnencremes in Hautquerschnitten zur Risikoabschätzung der Anwendungen von Nanotechnologie in kosmetischen Produkten. Desweiteren werden Studien der Spurenelementverteilung, speziell der von gebundenen Metallionen, in Hirnschnitten auf zellulärer und subzellulärer Ebene erläutert. Das anschließende Beispiel erläutert die Anwendung niedriger Energiedosen in der Radiobiologie anhand des Beschusses einzelner lebender Zellen mit abgezählten einzelnen Ionen. Als letztes Beispiel wird die Anwendung hochfokussierter Ionenstrahlen für die Mikrotomographie gezeigt. Abschließend folgt eine zusammenfassende Bewertung der vorgestellten Anwendungen mit einem Ausblick auf weitere Anwendungen und methodische Entwicklungen. Der Arbeit sind die relevanten Veröffentlichungen mit Beteiligung des Autors als Anhang beigefügt