Motility of flagellated bacteria has been a topic of increasing scientific interest over the past decades, attracting the attention of mathematicians, physicists, biologists and engineers alike. Bacteria and other micro-organisms cause substantial damage through biofilm growth on submerged interfaces in water cooling systems, ship hulls and medical implants. This gives social and economic motivations for learning about how micro-organisms swim and behave in different environments. Fluid flows on such small scales are dominated by viscosity and therefore behave differently from the inertia-dominated flows that we are more familiar with, making bacterial motility a physically intriguing phenomenon to study as well. We use the boundary element method (BEM) to simulate the motion of singly flagellated bacteria in a viscous, Newtonian fluid. One of our main objectives is to investigate the influence of external surfaces on swimming behaviour. We show that the precise shape of the cell body and flagellum can be important for determining boundary behaviour, in particular, whether bacteria are attracted or repelled from surfaces. Furthermore, we investigate the types of motion that may arise between two parallel plates and in rectangular channels of fluid and show how these relate to the plane boundary interactions. As an extension to original models of flagellar propulsion in bacteria that assume a rotation of the rigid helical flagellum about an axis fixed relative to the cell body, we consider flexibility of the bacterial hook connecting the aforementioned parts of the swimmer. This is motivated by evidence that the hook is much more flexible than the rest of the flagellum, which we therefore treat as a rigid structure. Elastic dynamics of the hook are modelled using the equations for a Kirchhoff rod. In some regimes, the dynamics are well described by a rigid hook model but we find the possibility of additional modes of behaviour.

This work deals with the development of a new label-free detection technique called Pulsed Streaming Potential (PSP). Its novelty relies on the adaptation of a classical electrokinetic phenomenon (streaming potential) into a tool which can evaluate molecular interplay in label-free fashion. Implementation of PSP to microfluidic platforms allowed the label-free sensing of binding events to plastic (modified and unmodified) surfaces. It was demonstrated the use of real time PSP in plastic microfluidic platforms for determination of kinetic parameters of the interaction of proteins and plastic surfaces. Moreover, initial change of PSP after adsorption of proteins showed to be proportional to the bulk concentration of proteins and it was used for quantification of Lysozyme in the nanomolar range. Several approaches were studied to manipulate the surface of microfluidic channels in order to improve selectivity of PSP through reduction of non-specific adsorption. These approaches included the fabrication of composite surface of polyacrilic acid (PAA) and polyethylene glycol acrylate (PEGA) on cyclic olefin copolymer microchannels, as well as adsorption of nanospheres on COC-PEGA channels.

<p>Cancer comprises a collection of diseases, all of which begin with abnormal tissue growth from various stimuli, including (but not limited to): heredity, genetic mutation, exposure to harmful substances, radiation as well as poor dieting and lack of exercise. The early detection of cancer is vital to providing life-saving, therapeutic intervention. However, current methods for detection (e.g., tissue biopsy, endoscopy and medical imaging) often suffer from low patient compliance and an elevated risk of complications in elderly patients. As such, many are looking to “liquid biopsies” for clues into presence and status of cancer due to its minimal invasiveness and ability to provide rich information about the native tumor. In such liquid biopsies, peripheral blood is drawn from patients and is screened for key biomarkers, chiefly circulating tumor cells (CTCs). Capturing, enumerating and analyzing the genetic and metabolomic characteristics of these CTCs may hold the key for guiding doctors to better understand the source of cancer at an earlier stage for more efficacious disease management.</p><p> The isolation of CTCs from whole blood, however, remains a significant challenge due to their (i) low abundance, (ii) lack of a universal surface marker and (iii) epithelial-mesenchymal transition that down-regulates common surface markers (e.g., EpCAM), reducing their likelihood of detection via positive selection assays. These factors potentiate the need for an improved cell isolation strategy that can collect CTCs via both positive and negative selection modalities as to avoid the reliance on a single marker, or set of markers, for more accurate enumeration and diagnosis.</p><p> The technologies proposed herein offer a unique set of strategies to focus, sort and template cells in three independent microfluidic modules. The first module exploits ultrasonic standing waves and a class of elastomeric particles for the rapid and discriminate sequestration of cells. This type of cell handling holds promise not only in sorting, but also in the isolation of soluble markers from biofluids. The second module contains components to focus (i.e., arrange) cells via forces from acoustic standing waves and separate cells in a high throughput fashion via free-flow magnetophoresis. The third module uses a printed array of micromagnets to capture magnetically labeled cells into well-defined compartments, enabling on-chip staining and single cell analysis. These technologies can operate in standalone formats, or can be adapted to operate with established analytical technologies, such as flow cytometry. A key advantage of these innovations is their ability to process erythrocyte-lysed blood in a rapid (and thus high throughput) fashion. They can process fluids at a variety of concentrations and flow rates, target cells with various immunophenotypes and sort cells via positive (and potentially negative) selection. These technologies are chip-based, fabricated using standard clean room equipment, towards a disposable clinical tool. With further optimization in design and performance, these technologies might aid in the early detection, and potentially treatment, of cancer and various other physical ailments.</p> / Dissertation

Surfactant coated microbubbles are widely used as contrast agents (UCA) in medical ultrasound imaging, due to their high echogenicity and non-linear response to acoustic excitation. Controlling the stability of microbubbles in vivo represents a considerable challenge. Understanding the characteristics of the bubble surface and how they change with production method, composition and environment is key to addressing this problem. The aim of this thesis is to investigate viscosity, bubble dissolution, and acoustic response as functions of their composition, manufacturing method and environment. Bubbles were made using combinations of phospholipid and an emulsifier in different molar ratios. Adding the emulsifier decreased both the size and the surface viscosity of the bubbles and caused changes in the scattered pressure amplitude of bubbles under ultrasound. To increase microbubble stability, solid inorganic nanoparticles were adsorbed on to the microbubble surface. These particles behaved as Pickering stabilisers, and deterred Ostwald ripening. The nanoparticles also enhanced the nonlinear behaviour of bubbles at low acoustic pressures. Three manufacturing methods (sonication, cross-flow and flow focusing) were investigated in order to verify stability differences. Sonication produced bubbles with surface viscosities hundreds of centipoise greater than those produced by microfluidics. Both pressure amplitude and harmonic content for sonicated bubbles were found to be much larger due to a higher liposomal adhesion rate at the surface. Solution temperature and bubble age were also investigated. When the solutions were heated above the phospholipid gelling temperature, microfluidic bubbles showed an increased surface viscosity, due to increased liposome adhesion caused by the increased temperature. Bubble composition, manufacturing method and environment were found to vary the surface characteristics of the microbubbles. Further investigations into the affects of the filling gas, in vitro studies, and low temperature TEM characterisation should be conducted to produce a microbubble with the full range of desired characteristics.

Le but de cette thèse est de développer un outil microfluidique d'étude de la cristallisation (et plus particulièrement de la nucléation) le plus adapté aux contraintes de l'industrie. C'est-à-dire un outil permettant de réaliser un grand nombre d'expériences de cristallisation tout en utilisant le moins de produits possible et en restant simple à mettre en place expérimentalement. Seule la microfluidique permet, en utilisant des volumes de l'ordre du nL, de répondre simultanément à ces deux contraintes. Les systèmes microfluidiques permettent en effet de générer des gouttes de quelques nanolitres qui sont autant de microcristallisoires permettant l'étude de la nucléation. Ce travail présente la mise au point de systèmes microfluidiques et des méthodes analytiques associées dédiés à l'étude de la nucléation de principes actifs pharmaceutiques. Un système microfluidique existant a été adapté afin de répondre dans un premier temps aux problèmes posés par la cristallisation de protéines dans l'eau. Ce système a permis de mesurer la limite de zone métastable ainsi que la fréquence de nucléation d'une protéine modèle, le lysozyme, également utilisée comme principe actif. Puis ce système a été à nouveau adapté afin de permettre l'étude de la nucléation dans des solvants organiques variés et donc l'étude d'un grand nombre de principes actifs pharmaceutiques. À l'occasion de cette nouvelle adaptation, des méthodes plus polyvalentes d'études de la nucléation ont dû être mises au point afin de résoudre les nouveaux problèmes soulevés. / The aim of this work is to develop a microfluidic tool to study crystallization (and specifically nucleation) adapted to industrial issue, that's mean doing a lot of experiment with only few materials. Microfluidic, thanks to using nanoliters volume, are able to solve simultaneously both issues. Microfluidic system allows us to generate plenty of nanoliters droplets and each droplet is a microcristallizer to study nucleation. Here I present the development of a microfluidic system and the related analytical method dedicated to nucleation study of active pharmaceutical ingredient. As a first step we adapt an existing microfluidic system to study the nucleation of protein in water. Thanks to this system we are able to measure the metastable zone width and nucleation frequency of model protein used as an active pharmaceutical ingredient, the lysozyme. In a second step we modify this system in order to allow nucleation study in organic solvent. Thanks to this new system we can study the nucleations of different APIs using polyvalent methods develop to avoid nucleation problems due to the crystallization of API. This microfluidic system and the method develop to study nucleation of API are really polyvalent and let us imagine to extend their applicative field to all industrial field where using nanoliter volume can reduce the cost (protein crystallization) and/or risk (explosives, radioactive hazard).

<p>This thesis demonstrates a new way to achieve sparse biological sample detection, which uses magnetic bead manipulation on a digital microfluidic device. Sparse sample detection was made possible through two steps: sparse sample capture and fluorescent signal detection. For the first step, the immunological reaction between antibody and antigen enables the binding between target cells and antibody-­‐‑ coated magnetic beads, hence achieving sample capture. For the second step, fluorescent detection is achieved via fluorescent signal measurement and magnetic bead manipulation. In those two steps, a total of three functions need to work together, namely magnetic beads manipulation, fluorescent signal measurement and immunological binding. The first function is magnetic bead manipulation, and it uses the structure of current-­‐‑carrying wires embedded in the actuation electrode of an electrowetting-­‐‑on-­‐‑dielectric (EWD) device. The current wire structure serves as a microelectromagnet, which is capable of segregating and separating magnetic beads. The device can achieve high segregation efficiency when the wire spacing is 50µμm, and it is also capable of separating two kinds of magnetic beads within a 65µμm distance. The device ensures that the magnetic bead manipulation and the EWD function can be operated simultaneously without introducing additional steps in the fabrication process. Half circle shaped current wires were designed in later devices to concentrate magnetic beads in order to increase the SNR of sample detection. The second function is immunological binding. Immunological reaction kits were selected in order to ensure the compatibility of target cells, magnetic bead function and EWD function. The magnetic bead choice ensures the binding efficiency and survivability of target cells. The magnetic bead selection and binding mechanism used in this work can be applied to a wide variety of samples with a simple switch of the type of antibody. The last function is fluorescent measurement. Fluorescent measurement of sparse samples is made possible of using fluorescent stains and a method to increase SNR. The improved SNR is achieved by target cell concentration and reduced sensing area. Theoretical limitations of the entire sparse sample detection system is as low as 1 Colony Forming Unit/mL (CFU/mL).</p> / Dissertation

Nous présentons plusieurs expériences d'hydrodynamique interfaciale illustrant des procédés de création, stabilité et rupture d'interfaces fluides, liquide/liquide ou liquide/gaz. Dans un premier temps, nous étudions le processus de fragmentation d'objets déformables, gouttes et bulles, par un obstacle rectangulaire ou une boucle asymétrique placés dans un canal microfluidique. La deuxième partie de ce travail se consacre à des expériences menées avec des bulles et films liquides minces formés à l'aide d'eau savonneuse ou de liquides très visqueux. Après avoir revisité quelques surfaces minimales adoptées par un film de savon à l'équilibre, nous étudions divers mécanismes de formation de bulles et de bulles interfaciales. Dans ces deux parties, nos études sont menées en faisant varier de façon systématique les paramètres hydrodynamiques, physicochimiques et géométriques contrôlant chaque expérience. Nous interprétons les résultats obtenus en microfluidique à l'aide d'arguments simples basés notamment sur l'analogie électro-hydraulique aux bas nombres de Reynolds, tandis que l'analyse dimensionnelle et des lois d'échelle permettent de décrire la plupart des comportements expérimentaux des bulles et films liquides minces. / We report hydrodynamic experiments illustrating the creation, stability and rupture of either liquid/liquid or liquid/gas interfaces. First, we investigate the fragmentation of deformable objects, such as drops and bubbles, against a rectangular obstacle or at the entry node of an asymmetric loop placed in a microfluidic channel. In the second part of this work, we report experiments conducted with bubbles and thin-liquid films either made with soapy water or highly viscous fluids. After having described a few minimal surfaces sought by a soap film at equilibrium, we study a variety of mechanisms that yield the formation of bubbles and interfacial bubbles. In both parts, our investigations are conducted by systematically varying the parameters (hydrodynamic, physicochemical and geometric) controlling each experiment. We interpret microfluidic results with simple physical arguments based on the electro-hydraulic analogy at low Reynolds numbers. Experimental findings on bubbles are rationalized mostly using dimensional analysis and scaling laws.

<p>Microscale environments provide significant resolution and distortion challenges with respect to measurement techniques; however, with improvements to existing techniques, it is possible to gather relevant data to better understand the thermal and fluidic mechanisms at such small scales in evaporating droplets.</p> <p> </p> <p>Infrared thermography provides several unique challenges at small scales. A primary issue is that the low native resolution of traditional infrared cameras significantly hamper the collection of details of microscale features. Furthermore, surfaces exhibiting vastly different emissivities, results in inaccurate temperature measurements that can only be corrected with irradiance-based emissivity maps of the surface; however, due to the resolution limitations of infrared thermography, these emissivity maps can also display significant errors. These issues are overcome through the use of multi-frame super-resolution. The enhanced resolution allows for better capture of microscale features, therefore, enhancing the emissivity map. A quantitative error analysis of the system is conducted to quantify the feature size resolution improvement as well as the smoothing effect of super-resolution reconstruction. Furthermore, a sensitivity analysis is conducted to quantify the impact of registration uncertainty on the accuracy of the reconstruction. Finally, the improved emissivity map from super-resolution is demonstrated to show the increased accuracy over low-resolution mapping.</p> <p> </p> <p>When applied to water droplets, particularly on nonwetting surfaces, infrared thermography is confounded by the presence of nonuniform reflectivities due to the spherical curvature of the liquid-air interface. Thus, when measuring the temperature along the vertical axis of a water droplet, it is necessary to correct the reflection. Using a controlled background environment, in conjunction with the Fresnel equations, it is possible to correct the reflective effects on the interface and calculate the actual temperature profile. This allows for a better understanding of the governing mechanisms that determine the thermal transport within the droplet. While thermal conduction is the primary transport mechanism along the vertical axis of the droplet, it is determined that the temperature drop is partially dampened by the convective transport from the ambient air to the liquid interface. From this understanding revealed by the measurements, the vapor-diffusion-based model for evaporation was enhanced to better predict evaporation rates.</p> <p> </p> <p>Further exploration into the mechanisms behind droplet evaporation on nonwetting surfaces requires accurate knowledge of the internal flow behavior. In addition, the influence of the working fluid can have a significant impact on the governing mechanisms driving the flow and the magnitude of the flowrate. While water droplet evaporation has been shown to be governed by buoyancy-driven convection on nonwetting substrates, similar studies on organic liquid droplets are lacking. Particle image velocimetry is effective at generating a velocity flow field, but droplets introduce distortion due to the refraction from the spherical interface of the droplet. As such, velocity correction using a ray-tracing approach was conducted to correct the velocity magnitudes and direction. With the velocity measurements, the flow was determined to be surface-tension-driven and showed speeds that are an order of magnitude higher than those seen in buoyancy-driven flow in water droplets. This resulted in the discovery that advection plays a significant role in the transport within the droplet. As such, the vapor-diffusion-governed evaporation model was adjusted to show a dramatic improvement at predicting the temperature gradient along the vertical axis of the droplet.</p> <p> </p> <p>Armed with the knowledge of flow behavior inside droplets, it is expected that droplets with aqueous solutions should exhibit buoyancy-driven convection. The final part of this work, therefore, leverages this phenomenon to enhance mixing during reactions. Colorimetry is a technique that is widely utilized to measure the concentration of a desired sample within some liquid; the sample reacts with a reagent dye the color change is measured, usually through absorbance measurements. In particular, the Bradford assay is used to measure protein concentration by reacting the protein to a Coomassie^TM Brilliant Blue G-250. The absorbance of the dye increases, most significantly at the 590 nm wavelength, allowing for precise quantitation of the amount of protein in the solution. A droplet-based reaction chamber with buoyancy-enhanced mixing has the potential to speed up the measurement process by removing the need for a separate pre-mixing step. Furthermore, the reduced volume makes the process more efficient in terms of reactant usage. Experimental results of premixed solutions of protein sample and reagent dye show that the absorbance measurement through a droplet tracks strongly with the protein concentration. When the protein sample and dye reagent are mixed <i>in situ</i>, the complex interaction between the reactants, the mixing, and the adsorption of protein onto the substrate creates a unique temporal evolution in the measured absorbance of the droplet. The characteristic peaks and valleys of this evolution track strongly with concentration and provide the framework for measurement of concentration in a droplet-based system.</p> <p> </p> <p>This thesis extends knowledge about droplet thermal and fluidic behavior through enhanced measurement techniques. This knowledge is then leveraged in a novel application to create a simple, buoyancy-driven colorimetric reaction setup. Overall, this study contributes to the field of miniaturized, efficient reaction and measurement devices.</p>

Ce travail de thèse est consacré à la manipulation et la déformation optique d'interfaces liquides molles, cela dans deux géométries fondamentales: plane et sphérique. Nous montrons alors que les déformations induites par pression de radiation optique permettent de déduire les propriétés des interfaces, comme la tension interfaciale par exemple. Dans le cadre de la déformation d'une interface liquide plane par pression de radiation, nous généralisons pour la première fois la manifestation électro-hydrodynamique des cônes de Taylor au régime optique, en montrant que des cônes liquides peuvent émerger sous fortes excitation laser. Nous avons alors caractérisé la morphologie de ces « cônes optiques » et nous montrons que l'angle de ces derniers dépend à la fois des paramètres de l'excitation laser mais aussi des caractéristiques des fluides. Une étude analytique ainsi qu'une étude numérique ont alors été menées afin de rendre compte des comportements observés.Afin d'étudier la déformation d'interfaces molles en géométrie sphérique, nous avons développé un double piège optique fibré en dispositif microfluidique dans une configuration inédite en termes de longueur d'onde excitatrice et de puissance laser. Nous avons alors appliqué notre dispositif à la déformation de vésicules en tant qu'objets modèles mous et nous montrons que notre double piège est bien adapté à la caractérisation rhéologique d'objets micrométriques déformables. Grâce à l'utilisation de faisceaux laser de forte puissance, nous mettons ici en évidence expérimentalement l'apparition d'un régime non-linéaire de déformation au sein de notre double piège optique. / This thesis work is devoted to the optical manipulation and deformation of soft liquid interfaces, in two fundamental geometries: plane and spherical. We then show that the deformations induced by optical radiation pressure allow to deduce the properties of interfaces, such as interfacial tension for example. In the framework of the deformation of a plane liquid interface by optical radiation pressure, we generalize for the first time the electro-hydrodynamic manifestation of Taylor cones to the optical regime, showing that liquid cones can emerge under intense laser excitation.We then characterized the morphology of these "optical cones" and we show that their angle depends both on the parameters of the laser excitation and on the characteristics of the fluids. An analytical study as well as a numerical investigation were then conducted to account for the observed behaviors. In order to study the deformation of soft interfaces in spherical geometry, we have developed a fiber-based dual-beam optical trap in a microfluidic device in a novel configuration in terms of excitation wavelength and laser power. We then applied our device to the deformation of vesicles as soft model objects and we show that our dual-beam trap is well adapted to the rheological characterization of deformable micron-sized objects. Thanks to the use of high laser power beams, we experimentally highlight the appearance of a non-linear deformation regime within our double optical trap.

Este trabalho descreve o desenvolvimento de um biossensor contendo transdução condutométrica sem contato (C4D, capacitively coupled contactless conductivity detection) e ácido fólico (FA) como biorreceptor em microchip, uma nova alternativa que poderá ser utilizada na determinação do biomarcador tumoral FR-&alpha;. Essa espécie exibe interações com FA altamente específicas, com constantes de formação da ordem de 109-1010. Os dispositivos microfluídicos, os quais consistiram de uma lâmina de vidro (integrando os eletrodos), dielétrico (contendo a fase biossensora) e substrato de poli(dimetilsiloxano) (PDMS, incorporando os microcanais), foram fabricados utilizando-se processos de fotolitografia e deposição de filmes finos em fase vapor. Objetivando melhorias nos níveis de detecção da C4D, estudos de sensibilidade com base em parâmetros da curva analítica foram conduzidos alterando-se a natureza do dielétrico e a configuração dos eletrodos. Posteriormente, estudos de caracterização foram realizados para as superfícies modificadas com os intermediários de imobilização; condições reacionais distintas (reagente, concentração, solvente e tempo) foram consideradas. As técnicas de microscopia eletrônica de varredura e espectroscopia de fotoelétrons excitados por raios-X foram usadas, respectivamente, a fim de se verificar a possível formação de aglomerados e permitir determinações qualitativas e quantitativas sobre as composições químicas das superfícies. Como resultado dos experimentos de sensibilidade e caracterização de superfície, adotamos os parâmetros seguintes para os ensaios de interações biomoleculares posteriores: filme de SiO2 como dielétrico, eletrodos seletivos à C4D com formato retangular e orientação antiparalela e monocamadas automontadas do reagente 3-aminopropil(trietoxisilano) como intermediário de imobilização de FA. As duas etapas finais do trabalho foram: otimização do tempo de funcionalização com FA (3, 5 e 7 h) e caracterização da fase biossensora, realizada a partir de medidas de C4D e microscopia de força atômica (AFM). Para o primeiro caso, os microchips foram aplicados a um padrão de anticorpo monoclonal específico a FA (&alpha;-FA). Os ensaios biomoleculares indicaram uma adsorção efetiva de FA junto à superfície de SiO2 silanizada, sem a ocorrência (ao menos em níveis significativos) de impedimentos estéricos de sua espécie bioativa. Dentre os tempos de funcionalização investigados, 3 h foi aquele que resultou em uma maior sensibilidade do método. Em termos da etapa de caracterização eletroquímica da fase biossensora, seus resultados mostraram haver correlação entre a resposta analítica e as interações FA/&alpha;-FA. Em adição, conforme indicaram as medidas de AFM, não houve alterações drásticas na morfologia do substrato (SiO2) em função dos processos de modificação química de superfície. Por fim, o uso da C4D como uma técnica de transdução em biossensores mostrou-se uma alternativa promissora para a análise do biomarcador tumoral FR-&alpha;. Dentre outros aspectos, essa plataforma analítica requer uma instrumentação simples, barata e portátil, não apresenta inconvenientes relacionados ao contato eletrodo/solução, dispensa o uso de mediadores redox e permite a determinação simultânea de multianalitos. Neste ínterim, alterações no transdutor devem ser implementadas visando um aumento na sensibilidade do método, o qual representa seu fator limitante principal. / This work describes the development of a biosensor containing capacitively coupled contactless conductivity transduction (C4D) and folic acid (FA) as bioreceptor in microchip, a new alternative that can be used in FR-&alpha; tumor biomarker analysis. FR-&alpha; exhibits highly specific interactions with FA, showing formation constants of the order of 109-1010. The microfluidic devices consisted of a glass layer (integrating the electrodes), dielectric (containing the biosensor phase), and poly(dimetilsiloxane) substrate (PDMS, incorporating microchannel). The microfabrication stage evolved photolithography processes, metal adsorption via sputtering, and plasma-enhanced vapor film deposition. In order to improve detection levels of C4D, sensitivity studies were conducted by changing the dielectric nature and electrode configuration. Through flow analysis with given electrolyte standards, the limits of detection and quantification were calculated based on analytical curve parameters. Subsequently, researches were performed to characterize the modified surfaces with immobilization intermediate considering reaction conditions distinct (reagent, concentration, solvent, and time). The techniques of scanning electron microscopy and X-ray photoelectron spectroscopy were employed, respectively, aiming to verify the clusters formation and allow qualitative and quantitative determinations about the surfaces chemical composition. From the results of sensitivity experiments and surface characterization, we adopt the following parameters for the biomolecular interactions assays: SiO2 film as dielectric, C4D selective electrodes with rectangular shape and antiparallel orientation, and self-assembled monolayers of 3-aminopropyl(triethoxysilane) as intermediary for immobilization of FA. The two final steps of the work were: optimizing the FA functionalization time (3, 5, and 7 h) and phase biosensor characterization, made from measures of C4D and atomic force microscopy (AFM). For the first case, due to the absence of FR-&alpha; standard for purchase, the microchips were applied to FA specific monoclonal antibody (&alpha;-FA). The biomolecular assay indicated effective adsorption of FA, without occurrence (at least in significant levels) of steric hindrance of its bioactive specie. Among the investigated times of functionalization, 3 h resulted in a higher sensitivity of the method. In terms of biosensor phase electrochemical characterization stage, their results evidenced correlation between analytical response and FA/&alpha;-FA interactions. Additionally, as the AFM measurements showed, drastic changes in the morphology of the substrate (SiO2) with the surface modification processes did not occur. Finally, the use of the C4D as transduction technical in biosensors proved to be a promissory alternative for FR-&alpha; tumor biomarker analysis. Among other features, this platform has not drawbacks related to the electrode/solution contact, dispenses the use of redox mediators, allows the simultaneous determination of multianalytes, and employs an instrumentation that is simple, cheap, and portable. Nevertheless, changes in the transducer should be implemented to increase the method sensitivity, which represents its main limiting factor.