We investigate experimentally the effects of acoustic forces on submicron aerosol in a channel flow. This technique can potentially overcome some of the limitations of conventional separation systems and provide advanced manipulation capabilities such as sorting according to size or density. The theoretical framework for acoustophoresis at such small length scales where molecular effects are expected to be significant is still incomplete and in need of experimental validation. The main objectives of this thesis are to identify the physical limitations and capabilities of acoustophoretic manipulation for submicron aerosol particles. Two sets of experiments were carried out: first, qualitative results revealed that acoustic manipulation is possible for submicron particles in air and that the acoustic force follows the trend expected by theoretical models developed for particles in inviscid fluids. The acoustic force on submicron particles was estimated in a second set of measurements performed with quantitative diagnostic tools. Comparison of these results with available theoretical models for the acoustic radiation forces demonstrates that for such small particles additional forces have to be considered. At submicron length scales, the magnitude of the forces observed is orders of magnitude higher than the predictions from the inviscid theory. One potential application for acoustophoresis is specifically investigated in this thesis: assist electrostatic precipitation (ESP) samplers to target very small aerosols, such as those carrying airborne viruses. To identify the shortcomings of ESP samplers that acoustophoresis should overcome, two ESP designs have been investigated to quantify capture efficiency as a function of the particle size and of the air velocity in a wind tunnel. The results reveal that both designs have limitations when it comes to sampling submicron aerosol particles. When exposed to polydispersed suspensions they behave as low-pass filters. / <p>QC 20161125</p>

Acoustophoresis is a label free method where the acoustic radiation force is used to manipulate microparticles inside microfluidic channels. The magnitude of the force is dependent of several parameters, which include the density, speed of sound and size of the microparticles, as well as the amplitude of the pressure waves. Recently, acoustophoresis has been used in microfluidics to manipulate microparticles inside moving droplets. In this Master's thesis project, two microfluidic chip designs are used to enrich droplets with polystyrene beads (10 μm in diameter) using acoustophoresis. The microchips have been fabricated with two different fabrication methods; crystalline dependent wet etching and crystalline independent dry etching. In the microchips, water droplets in oil are generated with microparticles suspended in them. By using a channel width that is half a wavelength of the incoming acoustic waves, pressure nodal lines are created in the middle of the channel in which the microparticles align. The droplets then enters a droplet splitting feature, where they are divided into three daughter droplets. Since the majority of the incoming particles are recovered in the center daughter droplet while some of the droplet volume is removed, the center droplet is enriched with the microparticles. For the wet etched design stable droplet splitting was observed when the volumetric flow was 18 μL/min and the incoming droplets had a length-to-width ratio larger than 3. The maximum recovery for this design was 81.1% ± 13.8% with an applied voltage at 10 Vpp. Stable droplet splitting was observed for the dry etched chip at 10.5 μL/min and 18 μL/min at 10 and 20 Vpp, when the incoming droplet had a length-to-width ratio of 3. In this chip the maximum recovery was 93.2% ± 8.3% at the volumetric flow of 10.5 μL/min and an applied voltage of 20 Vpp.

Diagnostic ultrasound (US) is safer, quicker and cheaper than other diagnostic imaging modalities. Over the past two decades, the applications of US imaging has been widened due to the development of injectable, compressible and encapsulated microbubbles (MBs) that provide an opportunity to improve conventional echocardiographic imaging, blood flow assessment and molecular imaging. The encapsulating material is manufactured by different biocompatible materials such as proteins, lipids or polymers. In current research, researchers modify the encapsulated shell with the help of advanced molecular chemistry techniques to load them with dyes (for fluorescent imaging), nanoparticles and radioisotopes (for multimodal imaging) or functional ligands or therapeutic gases (for local drug delivery). The echogenicity and the radial oscillation of MBs is the result of their compressibility, which undoubtedly varies with the encapsulated shell characteristics such as rigidity or elasticity. In this thesis, we present acoustic properties of novel type of polyvinyl alcohol (PVA)-shelled microbubble (PVA-MB) that was further modified with superparamagnetic iron oxide nanoparticles (SPIONs) to work as a dual-modal contrast agent for magnetic resonance (MR) imaging along with US imaging. Apparently, the shell modification changes their mechanical characteristics, which affects their acoustic properties. The overall objective of the thesis is to investigate the acoustic properties of modified and unmodified PVA-MBs at different ultrasound parameters. The acoustic and mechanical characterization of SPIONs modified PVA-MBs revealed that the acoustical response depends on the SPION inclusion strategy. However they retain the same structural characteristics after the modification. The modified MBs with SPIONs included on the surface of the PVA shell exhibit a soft-shelled behavior and produce a higher echogenicity than the MBs with the SPIONs inside the PVA shell. The fracturing mechanism of the unmodified PVA-MBs was identified to be different from the other fracturing mechanisms of conventional MBs. With the interaction of high-pressure bursts, the air gas core is squeezed out through small punctures in the PVA shell. During the fracturing, the PVA-MBs exhibit asymmetric (other modes) oscillations, resulting in sub- and ultra-harmonic generation. Exploiting the US imaging at the other modes of the oscillation of the PVA-MBs would provide an opportunity to visualize very low concentrations of (down to single) PVA-MBs. We further introduced the PVA-MBs along with particles mimicking red blood cells in an acoustic standing-wave field to observe the acoustic radiation force effect. We observed that the compressible PVA-MBs drawn toward pressure antinode while the solid blood phantoms moved toward the pressure node. This acoustic separation method (acoustophoresis) could be an efficient tool for studying the bioclearance of the PVA-MBs in the body, either by collecting blood samples (in-vitro) or by using the extracorporeal medical procedure (ex-vivo) at different organs. Overall, this work contributes significant feedback for chemists (to optimize the nanoparticle inclusion) and imaging groups (to develop new imaging sequences), and the positive findings pave new paths and provide triggers to engage in further research. / <p>QC 20150827</p> / 3MiCRON

Changes in the biomechanical properties of cells accompanying the development of various pathological conditions have been increasingly reported as biomarkers for various diseases and as a predictor of disease progression stages. For instance, cancer cells have been found to be less stiff compared to their healthy counterparts due to the proteomic and lipidomic dysregulations conferred by the underlying pathology. The separation and selective recovery of cells or extracellular vesicles secreted from such cells that have undergone these changes have been suggested to be of diagnostic and prognostic value. This dissertation first describes the implementation of a stiffness-based separation of phosphatidylcholine-based vesicles using a method first introduced based on the research in this work and was dubbed thermally-assisted acoustophoresis, or thermo-acoustophoresis. By tuning the temperature, we achieved the separation of vesicles of the same size, shape, and charge but with different stiffness values. It was observed that at a specific transition point, the acoustic contrast factor of vesicles changed sign from positive to negative. This change was mainly due to change in the compressibility of the vesicles, which is inversely proportional to stiffness. The acoustic contrast temperature (Tϕ), corresponding to the temperature at which the contrast factor switches sign, was determined to be unique to the composition of the vesicles. This unique temperature signature allowed us to develop this separation method of vesicles with distinct membrane stiffness with target outlet purities exceeding 95%. We have further explored the functionality of this method by experimenting with cholesterol-containing vesicles. In cells, the cholesterol content plays a crucial role in determining stiffness. Changes in the cholesterol content in cellular membranes can be an indication of pathological disorders. We evaluated the Tϕ of vesicles at different cholesterol molar ratios (Xchol) and developed a multi-stage lab-on-a-chip method to accomplish for the first time the separation of a three-vesicle mixture. Using Xchol = 0.1, 0.2, and 0.3 vesicles, we obtained efficiencies exceeding 93%. The simplicity, rapidity, and label-free nature of this approach holds promise as a diagnostic and separation tool for cells affected by diseases that affect the stiffness and extracellular vesicles such as exosomes and microvesicles.

Cette thèse s’inscrit dans le contexte du développement des laboratoires sur puce (LOC, « Lab On a Chip », permettant de réaliser plusieurs opérations nécessaires à l’analyse d’un échantillon biologique à l'intérieur d'un seul microsystème. Dans ce type de dispositif, de nombreuses étapes sont nécessaires avant d’arriver au résultat d’une analyse donnée (introduction de l'échantillon, concentration, mélange, purification, séparation, etc.). L’équipe microsystèmes du laboratoire Ampère étudie depuis plusieurs années différentes techniques de manipulation sans contact de particules, pour le tri ou de manipulation de particules individuelles dans les laboratoires sur puce, telles que la diélectrophorèse ou la magnétophorèse. Dans cette thèse, nous nous intéressons à la manipulation acoustique de micro particules. Cette technique se révèle notamment avantageuse pour la manipulation d’objets biologiques comme des bactéries, car elle permet de s’affranchir de certaines contraintes de marquage ou de changement de milieu. Notre choix s’est porté sur l’emploi des ondes acoustiques de surface (SAW, « Surface Acoustic Waves »), compatibles avec la filière PDMS très utilisée dans la communauté des LOC. Outre la possibilité de simplifier l’intégration microfluidique de la pince acoustique, la technologie SAW offre une alternative aux dispositifs à pièges acoustiques fixes existant dans la littérature en permettant un contrôle en temps réel des particules piégées. C’est ce que nous avons réalisé expérimentalement : en jouant sur le déphasage entre les signaux d’alimentation électriques des transducteurs électromécaniques, nous pouvons modifier la position des noeuds et des ventres de l’onde acoustique résultante. Ainsi, nous avons pu contrôler en temps réel la position d’une bille en latex de 3 μm ou encore d’un faisceau de bactéries E.coli. Par ailleurs, nous avons réalisé une simulation par éléments finis de la puce acoustofluidique dans son ensemble permettant une meilleure compréhension de tous les phénomènes en jeu et l’optimisation du transfert énergétique entre la source électrique et la particule manipulée. Cette simulation nous indique notamment que l’amplitude de l’onde acoustique stationnaire sur le substrat piézoélectrique varie environ d’un facteur deux en fonction du déphasage imposé entre les deux sources électriques. Cela impacte donc dans la même proportion la force acoustique résultante. Cette variation semble être validée par nos dernières expériences. / In lab-on-a-chip (LOC) technologies, many sample preparation steps are required before achieving a biological analysis on a single chip (sample introduction, concentration, mixing, purification, separation, etc.). The microsystem team of the Ampere Lab has studied for many years different contactless particle manipulation techniques, for sorting or manipulating bioparticles in LOC platforms, such as dielectrophoresis and magnetophoresis. In this thesis, we focus on acoustic manipulation of microparticles. This technique is advantageous for the manipulation of biological objects such as bacteria, because labelling and medium exchange can be avoided. We chose to work with surface acoustic waves (SAW), because this approach is consistent with the use of PDMS, widely used in microfluidics. Besides an easier microfluidic integration of the acoustic tweezers, the SAW technology provides an alternative to the existing devices with fixed acoustic traps, allowing a real time control of the trapped particles. This was experimentally achieved by playing on the phase shift between the two electrical signals driving the IDT, thereby modifying the position of nodes and antinodes of the resulting pressure wave. As a result, we could control in real time the position of a 3 μm latex bead or an E.coli bacteria alignment. We have also developed a finite-element model of the whole acoustofluidic chip allowing a better understanding of the physics and the optimization of the energy transfer between the electrical source and the trapped particle. Among different results, this model informs us that the magnitude of the acoustic radiation force varies by a factor of two with the phase shift between the electrical sources. This result seems to be validated by our last experiments.

Microfluidics is a field of research that enables the manipulation of fluids in the submillimetre length scale. The technology allows the development of lab-on-a-chip devices, which are miniaturized systems for chemical and biological analysis. Currently, the conventional manufacturing methods for these systems require multiple time-consuming steps. Therefore, focus has shifted towards laser micromachining as an alternative method. Direct laser writing would circumvent many of the steps required for the conventional methods, drastically reducing the process time. In this Master thesis project, it was shown that microfluidic chips can be manufactured using a Nd:YVO4 (532 nm) nanosecond laser system. The process was optimized for silicon and borosilicate glass substrates. Acoustic focusing of polystyrene beads was demonstrated for a system etched in silicon. The optimized process used a power of 50%, a frequency of 10 kHz, a scan speed of 60 mm/s with triple lines as fill type and it had an etch rate of 4.3 μm/pass. Processed wafers were cleaned in buffered HF and bonded using anodic bonding as well as adhesive bonding. Processing of glass proved unpredictable, resulting in cracks and chippings. However, in- and outlets were successfully etched through thin glass wafers. It was found that crucial factors for the process were to control the focus, positioning of structures, structure orientation and the pulse separation for a uniform distribution of pulses. Based on the results, it is estimated that the manufacturing process could be done in two to three days using the laser micromachining process.

This project aimed at exploring new hybrid materials to be used for acoustophoresis applications. Acoustophoresis can be used to manipulate particles inside a microfluidic channel by creating ultrasound standing waves within the channel [1]. This can be used for cell separation [2] or trapping of particles [3]. The intent of this project was to create materials for use in microfluidic channels that would be cheaper and easier to manufacture than those traditionally used, while still having adequate acoustic properties to allow for use in acoustopheresis. This was done by investigating whether the addition of glass-beads or glass-bubbles could increase the acoustic properties of an off-stoichiometry-thiol-enes (OSTE) based polymer. Hybrid samples with different volume fractions of glass-beads or glass-bubbles added to the OSTE polymer were manufactured and characterised according to their acoustic properties using the pulse-echo buffer-rod method. The acoustic properties measured were the density, attenuation, acoustic impedance and the reflection coefficient between water and the material. The addition of glass-beads was found to increase the acoustic impedance while the inverse was found for the addition of glass-bubbles. Both the addition of glass-beads and glass-bubbles were found to increase the attenuation. The hybrid material that was found to have the most suitable acoustic properties was OSTE/Glass-beads 40%, whose acoustic impedance had been increased ∼60% compared to pure OSTE. Consequently, the OSTE/Glass-beads 40% material was used to manufacture a microfluidic channel. A particle trapping experiment showed that the OSTE/Glass-beads 40% microfluidic channel was able to obtain bead trapping. This means that a standing wave was able to be generated within the channel and that it was strong enough to trap particles in the centre of the channel. However, evaluation of the particle trapping efficiency of the channel showed that it was not as effective as those using traditional materials. Therefore, future work is recommended to optimise a channel design for the OSTE/Glass-beads 40% material to increase the particle trapping efficiency. / I detta projekt undersöktes ett nytt hybridmaterial för användning i applikationer inom akustofores. Akustofores kan användas till att manipulera partiklar inuti mikrofluidkkanaler genom att generera ståendevågor i kanalen med hjälpav ultraljud [1]. Detta kan användas till cellseparation [2] eller till att fånga partiklar [3]. Målet i detta projekt var att skapa material som skulle bli billigare och möjliggöra enklare fabricering av kanalerna som används inom akustofores än de material som traditionellt används, med bibehållande av tillräckliga akustiskaegenskaper. Detta genomfördes genom att undersöka om tillsättning av glaspärlor eller glasbubblor kunde förbättra de akustiska egenskaperna av en off-stoichiometry-thiol-enes (OSTE) baserad polymer. Hybridprover gjorda på OSTE-polymeren med olika volymandelar av glaspärloroch glasbubblor tillverkades och kategoriserades med avseende på deras akustiska egenskaper med hjälp av pulseeko buffertstång metoden. De akustiska egenskaperna som uppmättes var densitet, attenuering, akustisk impedans och reflektions koefficienten mellan vatten och materialet. Resultatet av projektet visade att tillsättning av glaspärlor ökade den akustiska impedansen  i motsatts till glasbubblorna som visade sig minska den. Vidare visade det sig att både tillsättningen av glaspärlor och glasbubblor ökade attenueringen. Det hybridmaterial som visade sig ha de mest lämpliga akustiska egenskaperna var OSTE/glaspärlor med en 40% volymandel av glaspärlor. Den akustiska impedansen hade förhöjts med cirka 60% jämfört med vanlig OSTE. Därför valdes det hybrid-materialet till att tillverka en mikrofluidikkanal. Därefter genomfördes ett partikelfångstexperiment som visade att, OSTE/glaspärlor med en 40% volymandel av glaspärlor, kunde erhålla partikelfångst i kanalen. Detta innebär att en stående våg kunde genereras i kanalen och att den var tillräckligt stark för att kunna fånga partiklarna i mitten av kanalen. Däremot visade utvärdering av kanalens partikelfångsteffektivitet att den inte var lika effektiv som kanaler gjorda av traditionellt använda material. Därför rekommenderas framtida arbete till att designa en optimerad kanaldesign med OSTE/Glas-pärlor 40% materialets egenskaper i åtanke för att förhoppningsvis kunna öka partikelfångst effektivitet.

Droplet microfluidics is an emerging technology for analysis of single cells and biomolecules at high throughput. The controlled encapsulation of particles along with the surrounding microenvironment in discrete droplets, which acts as miniaturized reaction vessels, allows millions of particles to be screened in parallel. By utilizing the unit operations developed to generate, manipulate and analyze droplets, this technology platform has been used to miniaturize a wide range of complex biological assays including, but not limited to, directed evolution, rare cell detection, single cell transcriptomics, rare mutation detection and drug screening. The aim of this thesis is to develop droplet microfluidics based methods for analysis of single cells and nucleic acids. In Paper I, a method for time-series analysis of mammalian cells, using automated fluorescence microscopy and image analysis technique is presented. The cell-containing droplets were trapped on-chip and imaged continuously to assess the viability of hundreds of isolated individual cells over time. This method can be used for studying the dynamic behavior of cells. In Paper II, the influence of droplet size on cell division and viability of mammalian cell factories during cultivation in droplets is presented. The ability to achieve continuous cell division in droplets will enable development of mammalian cell factory screening assays in droplets. In Paper III, a workflow for detecting the outcome of droplet PCR assay using fluorescently color-coded beads is presented. This workflow was used to detect the presence of DNA biomarkers associated with poultry pathogens in a sample. The use of color-coded detection beads will help to improve the scalability of the detection panel, to detect multiple targets in a sample. In Paper IV, a novel unit operation for label-free enrichment of particles in droplets using acoustophoresis is presented. This technique will be useful for developing droplet-based assays that require label-free enrichment of cells/particles and removal of droplet content. In general, droplet microfluidics has proven to be a versatile tool for biological analysis. In the years to come, droplet microfluidics could potentially be used to improve clinical diagnostics and bio-based production processes. / <p>QC 20160926</p>

Recent advances and interest in ultrasound particle manipulation calls for theoretical understanding of acoustic radiation force and torque exerted on a configuration of multiple particles. In this thesis we theoretically study the acoustic radiation force and torque exerted by an arbitrary acoustic beam on a cluster of spherical particles in an inviscid fluid. The method is based on the partial-wave expansion (PWE) and the translational addition theorem for spherical wave functions. The combination of (PWE) and addition theorem Method enable us to solve the associated multiple scattering problem by numerically computing the (PWE) coefficients in a system of linear equations. On the other hand, when we consider the radiation force and torque exerted on a single sphere, the addition theorem has the advantage to solve this problem in a closed form. After obtaining the PWE coefficients, the acoustic radiation force and torque is computed through the farfield series solution. To illustrate the method, the acoustic radiation force and torque exerted on a single or multiple spheres are analyzed. In the case of a single sphere, the force is generated by a spherically focused ultrasound beam, where as the torque is generated by a Bessel vortex beam. For the multiple spheres configuration, the radiation force is induced by a traveling and a standing plane wave. In a specific configuration of three olive oil droplets suspended in water, with radii of the order of the wavelength, we found that rescattering events produce an acoustic interaction force, which significantly changes the radiation force on each droplet depending on the inter-droplet distance. In addition, we have found for the first time that an acoustic interaction torque due to the nonsymmetric spatial distribution of the acoustic energy density to the droplets. Further more, our study does not have restrictions on the spheres size compared to the wave length, nor on their composition material, which includes rigid, void, compressional liquid, elastic and viscoelastic solids, and layered material. Finally, this study has direct applications on methods for noncontact object handling by acoustic waves such as acoustic levitation, acoustical tweezers, and acoustophoresis in lab-on-a-chip devices. / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Recentes avanços e interesse em manipulação de partículas necessitam de uma maior compreensão teórica da força de radiação e torque acústico exercidos sobre uma configuração de múltiplas partículas. Nesta tese, nós estudamos teoricamente a força de radiação e torque acústico exercido por um feixe acústico arbitrário em um conjunto de partículas esféricas suspensas em um fluido não viscoso. O método baseia-se na expansão de ondas parciais (EOP) e no teorema translacional da adição para funções de onda esférica. A combinação do método de ondas parciais com o teorema da adição nos permitir resolver o problema de espalhamento mútiplo computando numericamente os coeficientes da expansão em um sistema de equações lineares. Por outro lado, quando consideramos a força e torque de radiação exercidos sobe uma única esfera, o teorema da adição tem a vantagem para resolver este problema exatamente. Após a obtenção dos coeficientes, a força e o torque de radiação são calculados usando um método em séries no campo distante. Para ilustrar o método, a força e o torque exercidos sobre uma ou multiplas esferas são analisados. Para o de uma única esfera, a força de radiação é gerada por um feixe de ultrassom focalizado. Para uma configuração de multiplas esferas, a força de radiação é induzida por ondas planas e estacionarias. Numa configuração específica de três gotas de azeite suspensas em água, com raios da ordem do comprimento de onda, verificou-se que as ondas reespalhadas produzem uma força de interação acústica, o que altera significativamente a força de radiação em cada gota em função da distância inter-gota. Além disso, verificou-se, pela primeira vez que um torque de interação acústico devido a uma distribuição espacial não simétrica da densidade de energia acústica para as gotas. Além disso, nosso estudo não tem restrições quanto ao tamanho esferas em comparação com o comprimento de onda, nem sobre a sua composição, que inclui rígida, líquida, elástica e sólidos viscoelásticos. Por fim, este estudo tem aplicações diretas sobre os métodos de manipulação de objetos sem contato por ondas acústicas, tais como a levitação acústica, pinças acústicas e acoustophoresis em dispositivos lab-on-a-chip.

Cette thèse de doctorat porte sur la description analytique des phénomènes acoustophorétiques, en solutions et suspensions. L'acoustophorèse est la création d'un champ électrique par une onde acoustique.La première partie porte sur les solutions d'électrolytes, et est basée sur l'analyse critique de la littérature. A partir des différents articles, un modèle original, basé sur la résolution des équations de la dynamique pour les ions est trouvé, lequel permet la prédiction, sans paramètres ajustables, de l'acoustophorèse des sels simples pour des solutions allant de très diluées à assez concentrées. Ce modèle est ensuite étendu aux cas de solutions avec trois espèces ioniques différentes, et un programme informatique calculant l'acoustophorèse en fonction de la concentration est en annexe. Une seconde extension est faite pour les liquides ioniques, et permet de déduire le volume des ions. Des tentatives d'extension du modèle sont faites pour les micelles et les colloïdes, en précisant les écueils. Une deuxième approche, basée sur la thermodynamique irréversible et les relations de réciprocité d'Onsager, est faite dans le cas des suspensions colloïdales. Les principaux résultats sont la proportionnalité entre l'acoustophorèse et la mobilité électrique des colloïdes, et donc l'applicabilité de cette technique à la caractérisation des suspensions, y compris concentrées ; le lien rigoureux entre l'acoustophorèse et son corollaire, la création d'une onde acoustique et son champ électrique ; enfin une procédure pour séparer le signal des colloïdes du signal de l'électrolyte support dans l'acoustophorèse des suspensions réelles. / This Ph.D. thesis is on the analytical description of acoustophoretic phenomena, in solutions and suspensions. Acoustophoresis is the creation of an electric field by an acoustic wave.First part is on electrolytic solutions, and it begins by a critical review of literature, from Debye first paper to a recent Ph.D. thesis on the same subject. Hypotheses are carefully selected, and a new model is deduced. This model, using pressure, friction, electric, inertia and corrective force, allows the prediction of acoustophoresis up to 0,3 molar for a simple salt, without any fitting parameter. An extension to solutions with three ionic species is done, and a Fortran program to compute the acoustophoresis as a function of the concentration is given in annex. Extension of the model, in the case of ionic liquid, allows the measurement of the volume of ions. A brief point is done on micellar and colloidal suspensions. A second part is on the application of non-equilibrium thermodynamic, especially Onsager reciprocal relation, to the acoustophoresis of suspensions. Acoustophoresis is shown to be proportional to the electric mobility, which allows the measurement of the latter in dark and concentrated suspensions. A link between acoustophoresis and the creation of acoustic wave by an electric field is also found, and a process to isolate contributions of colloids in real suspensions, with a supporting electrolyte, is proposed.