Self-immobilisation enzyme technologies, such as SphereZyme™, suffer from the lack of applicability to hydrolyse large substrates. Solid support immobilisation is usually a method of choice, to produce a stable biocatalyst for large substrates hydrolysis in the industry. In order to investigate this limitation, a commercial protease called Alcalase® was chosen as a model enzyme due to its natural activity (hydrolysis of large substrates-proteins). Prior to immobilising through the SphereZyme™ technology, Alcalase® was partially purified through dialysis followed by CM Sepharose™ FF cation exchanger. Sample contaminants, such as salts and stabilisers can inhibit protein crosslinking by reacting with glutaraldehyde. Alcalase® was successfully separated into 3 proteases with the major peak correlating to a positive control run on native PAGE, indicating that it was likely subtilisin Carlsberg. A 16% alkaline protease activity for azo-casein hydrolysis was retained when 5% v/v PEI: 25% v/v glutaraldehyde solution was used as a crosslinking agent in Alcalase® SphereZyme™ production. An increase in activity was also observed for monomeric substrates (PNPA) where the highest was 55%. The highest % activities maintained when 0.33 M EDA: 25% v/v glutaraldehyde solution was initially used as crosslinking agent were 4.5% and 1.6% for monomeric and polymeric substrates, respectively. PEI is a hydrophilic branched polymer with an abundance of amine groups compared to EDA. A comparison study of immobilisation efficiencies of SphereZyme™, Eupergit® and Dendrispheres was also performed for large substrate biocatalysis. The two latter technologies are solid-support immobilisation methods. Dendrispheres reached its maximum loading capacity in the first 5 minute of the one hour binding time. Twenty minutes was chosen as a maximum binding time since there was constant protein maintained on the solid support and no enzyme loss was observed during the 1 hour binding time. PEI at pH 11.5, its native pH, gave the highest immobilisation yield and specific activity over the PEI pH range of 11.5 to 7. SphereZyme™ had the highest ratio for azocasein hydrolysis followed by Dendrispheres and Eupergit®. The SphereZyme™ was also shown to be applicable to biosensors for phenol detection. Different modifications of glassy carbon electrode (GCE) were evaluated as a benchmark for the fabrication of SphereZyme™ modified phenol biosensor. GCE modified with laccase SphereZyme™ entrapped in cellulose membrane was the best modification due to the broad catechol range (<0.950 mM), high correlation coefficient (R2, 0.995) and relative high sensitivity factor (0.305 μA.mM-1). This type of biosensor was also shown to be electroactive at pH 7.0 for which its control, free laccase, lacked electroactivity. From the catalytic constants calculated, GCE modified with laccase SphereZyme™ entrapped in cellulose membrane also gave the highest effectiveness factor (Imax/Km app) of 1.84 μA.mM-1. The modified GCE with Alcalase® SphereZyme™ was relatively more sensitive than GCE modified with free Alcalase®.

Receptor-based biological detection techniques often suffer from the problem of non-specific interactions. This is largely due to the presence of weak electrostatic and Van der Waals forces between the receptor and the non-target substances in the analyte that are not easily dissociated in practice. Most existing detection techniques are unable to probe the interaction between the bound entity and the surface and differentiate between specific and non-specific interactions in terms of bond strength or activation energy. The resulting false positive responses lead to various issues, such as misdiagnosis and mistreatment in clinical diagnostics and false alarms in biosecurity. The problem is even more significant with direct direction techniques, such as the resonant frequency shift based detection using quartz crystal microbalance (QCM) or micro-cantilevers, which involve minimal sample processing and washing steps. The work presented in this thesis investigates, through modeling and experiments, the mechanical interactions of a resonator with microparticles attached via biomolecular linkers and analyses the resulting nonlinear acoustic modulation of the resonator from the transduced electrical signal. Physisorbed and specific interactions both in air and liquid medium are studied using thickness shear mode quartz crystal resonators and streptavidin-coated polystyrene microbeads (SCPM) of various sizes. It is found that the modification in the transduced electrical signal measured at the third harmonic (3f), or three times the driving frequency f, is significant in presence of the attached particles and approximately proportional to the number of particles. A detection limit of approximately 2 SCPM of 5.6 µm diameter in air and 6700 SCPM of 0.39 µm diameter in liquid is demonstrated, which corresponds to a mass detection limit of ~200 pg. Most interestingly, the deviation in the magnitude of the 3f signal as a function of the resonator oscillation amplitude is found to hold a distinct relationship with the type of particle-surface interaction. This provides a basis for selectivity in detection over and above the efficacy of the receptor. The function is also found to correlate well with the event of SCPM diffusion on the surface. This detection technique, based on the measurement of deviation in magnitude of the transduced electrical signal measured at a higher odd harmonic of the drive frequency due to the presence of surface-bound particles on a resonator, is termed as the anharmonic detection technique (ADT). A feasibility study with Bacillus subtilis spores in phosphate buffer saline (PBS) is carried out successfully where the modeling and experimental results with SCPM are successfully reproduced. A detection limit of 430 spores is demonstrated, which corresponds to a mass detection limit of ~650 pg. Capability for differentiation of the specifically-captured spores from unwashed physisorbed SCPM of similar dimensions is demonstrated using the shape of the ADT signal. These results indicate that the spore immobilization step may be directly followed by the detection step, which are 9 mins and 2 mins respectively in these experiments. ADT thus potentially enables a rapid, sensitive, reliable and direct detection without the need for any sample processing. Moreover, being an entirely electronic technique, ADT suitably lends itself to multiplexing, large scale fabrication and implementation on a miniaturized low-cost point-of-care detection platform that is of immense need in clinical diagnostics, food and environmental monitoring and biosecurity. Furthermore, fitting the experimental results with modeling estimates enables ADT to determine the force-extension characteristics of the binding biomolecular linker. The force-extension characteristics and the estimated unbinding force for a streptavidin-biotin complex estimated using ADT agrees well with those computed using molecular dynamics (MD) simulation at similar loading rates. Thus ADT contributes a unique force-spectroscopic method, which unlike conventional techniques such as the atomic force microscopy (AFM) provides statistically averaged data for multiple biomolecules in a relatively quicker and simpler experimental format. A method for determination of activation energy of the interaction is also proposed using ADT. This potentially enables a method for rapid and large scale biomolecular screening and studying of interaction networks, which have important applications in drug discovery and individualized therapy.

In this work, a chemiresistor-type sensing platform based on aligned arrays of nitrogen-doped multi-walled carbon nanotubes (N-MWCNTs) was developed. Our N-MWCNT based sensors can be made on both rigid and flexible substrates; they are small, have low power consumption and are suitable for highly efficient and reliable detection of different biomolecules and gases, at room temperature. The performance of these sensors was demonstrated for avian influenza virus (AIV) subtype H5N1 DNA sequences and toxic gases NO and NH3 at low concentrations. In our study, chemical vapor deposition (CVD) method was applied to synthesize vertically aligned nitrogen doped carbon nanotube arrays on a large area (> 1 cm2) on Si/SiO2 substrate using Fe/Al2O3 layer as a catalyst and a mixture of ethanol and acetonitrile as a C/N source. Especially, the diameter, length, nitrogen-doping concentration and morphology of the nanotubes were controllably tailored by adjusting the thickness of catalyst film, reaction duration and temperature as well as the amount of nitrogen-containing precursor. For integrating N-MWCNTs into chemiresistor devices, we developed a direct contact printing method for a dry, controllable and uniform transferring and positioning of the CVD-grown vertical nanotubes onto well-defined areas of various rigid and flexible substrates. After horizontally aligned N-MWCNT arrays were formed on a target substrate, interdigitated metallic microelectrodes with an interspacing of 3 µm were deposited perpendicular to the nanotube alignment direction to fabricate chemiresistor devices for biomolecule and gas sensing. This way, well-aligned nanotubes were laid across the Au/Cr interdigitated electrode fingers, had a strong adhesion with the electrodes and served as conducting channels bridging the electrodes. The N-MWCNT based chemiresistor device was applied as a label-free DNA sensor for a highly sensitive and fast detection of AIV subtype H5N1 DNA sequences. For this, the nanotubes were functionalized with probe DNA, which was non-covalently attached to sidewalls of the N-MWCNTs via π-π interaction. Such functionalized sensors were applied to quantitatively detect complementary DNA target with concentration ranging from 20 pM to 2 nM after 15 min incubation at room temperature. The sensors showed no response to non-complementary DNA target for concentrations up to 2 µM showing an excellent selectivity. Investigations on the efficient gas sensing of N-MWCNT-based chemiresistor of reducing/ oxidizing gases NH3 and NO were also reported in this work. The aim was to assess the possibility for N-MWCNTs to be applied as innovative sensing materials for room temperature gas sensing. N-MWCNTs with varying doping levels (N/C ratio of 5.6 to 9.3at%) were used as sensing materials and exposed to NH3 (1.5-1000 ppm) and NO (50-1000 ppm) for exploring and comparing their sensing performance. This study offered an effective route for further modification of CNTs according to various sensing application. Finally, our investigations showed a high potential of the developed N-MWCNT-based sensing platform for various applications ranging from environmental monitoring to point-of-care medical diagnostics.

Advancements in the micro-and nano-scale fabrication techniques have opened up new avenues for the development of portable, scalable and easier-to-use biosensors. Over the last few years, electrodes made of carbon have been widely used as sensing units in biosensors due to their attractive physiochemical properties. The aim of this research is to investigate different strategies to develop functionalized high surface carbon micro/nano-structures for electrochemical and biosensing devices. High aspect ratio three-dimensional carbon microarrays were fabricated via carbon microelectromechanical systems (C-MEMS) technique, which is based on pyrolyzing pre-patterned organic photoresist polymers. To further increase the surface area of the carbon microstructures, surface porosity was introduced by two strategies, i.e. (i) using F127 as porogen and (ii) oxygen reactive ion etch (RIE) treatment. Electrochemical characterization showed that porous carbon thin film electrodes prepared by using F127 as porogen had an effective surface area (Aeff 185%) compared to the conventional carbon electrode. To achieve enhanced electrochemical sensitivity for C-MEMS based functional devices, graphene was conformally coated onto high aspect ratio three-dimensional (3D) carbon micropillar arrays using electrostatic spray deposition (ESD) technique. The amperometric response of graphene/carbon micropillar electrode arrays exhibited higher electrochemical activity, improved charge transfer and a linear response towards H2O2 detection between 250μM to 5.5mM. Furthermore, carbon structures with dimensions from 50 nano-to micrometer level have been fabricated by pyrolyzing photo-nanoimprint lithography patterned organic resist polymer. Microstructure, elemental composition and resistivity characterization of the carbon nanostructures produced by this process were very similar to conventional photoresist derived carbon. Surface functionalization of the carbon nanostructures was performed using direct amination technique. Considering the need for requisite functional groups to covalently attach bioreceptors on the carbon surface for biomolecule detection, different oxidation techniques were compared to study the types of carbon–oxygen groups formed on the surface and their percentages with respect to different oxidation pretreatment times. Finally, a label-free detection strategy using signaling aptamer/protein binding complex for platelet-derived growth factor oncoprotein detection on functionalized three-dimensional carbon microarrays platform was demonstrated. The sensor showed near linear relationship between the relative fluorescence difference and protein concentration even in the sub-nanomolar range with an excellent detection limit of 5 pmol.

Neste trabalho são elaborados biossensores para a detecção amperométrica de glicose. Para isso, imobilizou-se a enzima glicose oxidase em matrizes de polímeros condutores. Foram construídos sensores utilizando-se poli(pirrol) e poli(N-metilpirrol). Com o objetivo de substituir o oxigênio molecular na etapa de transdução do sinal, o ferroceno foi incorporado dentro do polímero condutor. Para isso, os polímeros foram elaborados utilizando misturas água-etanol como meio de polimerização. A inclusão do ferroceno no sensor resulta em maior sensibilidad à glicose (4,33 &#181;A Mm-1 cm-2 para o biossensor preparado a partir da mistura água-etanol contendo o ferroceno e de 0,23 &#181;A mM-1 cm-2 para o sensor sem ferroceno ). Por outro lado, permite o funcionamento do sensor a potenciais menores que no caso do sensor sem ferroceno (0,4 V para o sensor com ferroceno vs. 0,65 V para o caso sem ferroceno). O deslocamento do potencial de detecção para valores menos positivos não foi suficiente para evitar as interferências causadas pelos íons ascorbato e ureato. Para isto, mostrou-se 100% efetivo o recobrimento dos sensores com uma película de Nafion&#174;. A sobreoxidação do poli(pirrol) também mostrou potencialidade para a eliminação de interferentes, embora o processo resulte na perda de sensibilidade, provavelmente causada pela desnaturação da enzima. Quando usado o poli(N-metilpirrol) como suporte para a enzima, obtiveram se melhores respostas, causadas pela possibilidade de se preparar filmes mais espessos, consequentemente de imobilizar maior quantidade de enzima, sem observar perda de resposta causada por problemas difusionais. / The present work describes the elaboration of a biosensor for glucose detection. The enzyme, glucose oxidase, was immobilized in different conducting polymers. Two different polymers were used: polypyrrole and poly(N-methilpyrrole ). With the aim of replacing the molecular oxygen in the transduction step, ferrocene has been immobilized within the conducting polymer. Once the ferrocenium was insoluble in water, in order to develop a different route, the electropolymerization was carried out in a mixture of water and ethanol (1:1). This procedure leads to a polymer with a poor electroactivity, detected by Raman experiments. The ferrocene addition in the sensor increases the sensitivity to the glucose determination (4,33 &#181;A mM-1 cm-2 for the biosensor with ferroecene and 0,23 &#181;A mM-1 cm-2 for the sensor without ferrocene). Alternatively, the sensor containing ferrocene allows to operate at less positive potentials than that one without ferrocene (+ 0,40 V and + 0,65 V, respectively). This potential shift was not enough to inhibit the interference caused by ascorbate and ureate ions. One method to avoid the interference problem was to recover the sensor with a very thin layer of Nafion. Also poly(pyrrole) overoxidation is a very efficient method to eliminate this interference, but this process leads to a sensitivity decrease dueto enzyme denaturation. A better response was observed for sensor assembled using the poly(Nmethyl-pyrrole) as the support for enzyme immobilization. This behavior was provoked by the thicker of polymer film formed leading to higher amount of immobilized enzyme. Even though, no diminution in the response was caused by diffusion problems.

Nuisance odor levels produced by solid waste management operations are subject to regulatory standards due to their impacts on the quality of life of the residents living nearby the facility. Failure to meet regulatory standards may result in fines, litigation, inability to acquire permits, mitigation, and re-siting operations. Since measurement of environmental nuisance odors is currently limited to subjective techniques, monitoring odor levels to meet such standards is often problematic. This is becoming more acute as increasing residential populations begin to encroach on properties adjacent to landfills. In order to ensure that nuisance odor issues are minimized, it is necessary to provide an objective measurement. The objective of the current research is to develop a biosensor for providing an objective, standard measurement of odors. The approach is to modify the human odorant binding protein (hOBPIIa), isolated using published biomolecular techniques, by fluorescently tagging it with a chromophore functional group. When this protein is tagged with a fluorophore marker and excited in a spectrofluorometer, it emits light of a certain wavelength that can be detected and quantified. Once odorant molecules are exposed to this complex, they start replacing the fluorophore, and as a result, the emitted light intensity decreases in proportion to the number of odorant molecules. Since the protein response depends on odorant concentration, following an inverse Beer’s Law relationship, the odorants can be quantified accurately and rapidly using fluorometric measurements. The results establish quantitation ranges for different pure and mixture of odorant gases as well as the amount of gas that can be quantified across various flow rates. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2020. / FAU Electronic Theses and Dissertations Collection

The development of molecular tools that enable precise manipulation and control of biological systems would allow for a broader understanding of cellular functions and applications in biotechnology, synthetic biology, and therapeutic research. The discovery of CRISPR/Cas systems and the understanding and repurposing of their mechanisms have revolutionized the field of molecular biology. Here, I identified and characterized novel CRISPR/Cas systems and applied them for different in vivo and in vitro applications. In this work, I interrogated various Cas13 effector proteins and identified the most efficient Cas13 effector (CasRx) for in planta applications. I adapted CasRx to engineer plant immunity against different plant RNA viruses. CasRx showed robust activity and specificity against RNA viruses, demonstrating its suitability for studying key questions relating to virus biology. To expand the Cas13 toolbox and enable new applications, I performed a homology search of Cas13 enzymes in prokaryotic genomes and metagenomes, and identified previously uncharacterized, novel CRISPR/Cas13 effector proteins. I first identified and functionally characterized a small size, miniature Cas13 effector (named here as mCas13) and combined it with isothermal amplification to develop a simple and sensitive CRISPR-based SARS-CoV-2 diagnostic platform. In addition, I discovered and biochemically characterized the first known thermostable Cas13 proteins and showed that these thermostable proteins are phylogenetically related. I harnessed the unique features of these thermostable enzymes to develop the first one-pot, RT-LAMP coupled Cas13-based nucleic acid detection assay, which was utilized for highly sensitive, specific, and easily programmable detection of SARS-CoV-2 and other viruses. Lastly, I utilized CRISPR/Cas12a to develop a detection assay of plant ssDNA geminiviruses with easy-to-interpret visual readouts, making it suitable for point-of-use applications. In addition, I leveraged the self vs. non-self-discrimination and pre-crRNA processing capabilities of CRISPR/Cas12a, with the allosteric transcription factors (aTFs)- regulated expression of CRISPR array to engineer a field-deployable small molecule detection platform. I demonstrated the ability of the developed platform to detect different tetracycline antibiotics with high sensitivity and specificity. In conclusion, my work demonstrates that the discovery and characterization of programmable nucleic acid targeting systems could enable their utility for biotechnological innovations, including technologies for inhibition of viral replication and diagnostics.

La tuberculose est une maladie contagieuse qui s’attaque habituellement aux poumons, mais parfois aussi à d’autres parties du corps, comme les reins, les ganglions et les os. La tuberculose tue près 1,8 millions de personnes chaque année dans le monde. Il y a par conséquent un besoin urgent de mettre des moyens analytiques pour détecter l’ADN de la bactérie Mycobacterium tuberculosis, responsable de la propagation de la tuberculose. La recherche développée dans le cadre de cette thèse consiste en l'élaboration d'un outil de diagnostic pour la détection d’ADN génomique de bactérie M.Tuberculosis après la lyse et amplification par PCR. Dans cette perspective, nous nous sommes intéressés au développement des biocapteurs basés sur des différentes morphologies de polypyrrole comme transducteur pour la détection électrochimique d'ADN du gène rpob de Mycobacterium tuberculosis. Une étude de l’impact des différentes morphologies sur les propriétés électriques des matériaux et également sur les performances des biocapteurs électrochimiques d’ADN a été effectuée. Nous avons aussi étudié l’effet de la fonctionnalisation par différentes chaine linéaire et ramifiée sur la structure des nanomatériaux de polypyrroles et leurs effets sur la quantité de biomolécules immobilisées. La stratégie adoptée pour le développement de ces matériaux est une construction du biocapteur réalisée étape par étape. Les différentes couches le constituant ont été caractérisées par différentes techniques de surface telles que les techniques électrochimiques et optiques, la microscopie électronique à balayage et la microscopie électronique à force atomique. Les propriétés des biocapteurs ont été suivies à travers les propriétés redox des groupes ferrocényles et naphtoquinone. La détection électrochimique de l’ADN cible évaluée avec ces biocapteurs à base de polypyrrole nanostructure montre une sensibilité de détection plus élevée que celle du biocapteur à base de polypyrrole couche compacte. Cela démontre le potentiel d'utilisation de la surface élevée des nanostructures polypyrrole comme transducteur et l’utilisation des approches de modification douce. Les biocapteurs ont été appliqués à la détection de l'ADN dans des échantillons réels d'ADN génomique de Mycobacterium tuberculosis et de l'ADN muté présentant la résistance de la rifampicine. Les biocapteurs basés sur des nanostructures de polypyrrole démontre une application potentielle dans la détection d'ADN et la capacité à discriminer le gène rpoB du mutant et pourrait être utilisé comme plate-forme dans la technologie des biocapteurs. / Tuberculosis is a contagious disease that usually attacks the lungs but can sometimes reach other parts of the body such as the kidneys, ganglia and bones. This disease is responsible for killing nearly 1.8 million people each year worldwide. Therefore, we have an urgent need to put analytical means to detect the DNA of the bacterium Mycobacterium tuberculosis, responsible for the spread of tuberculosis. The research developed in this thesis consists in the development of a diagnostic tool for the detection of genomic DNA of M.Tuberculosis bacteria after lysis and amplification by PCR. In this perspective, we focused on the development of biosensors based on different polypyrrole morphologies as transducers for the electrochemical detection of DNA of the rpob gene of Mycobacterium tuberculosis. A study of the impact of different morphologies on the electrical properties of materials as well as the performance of electrochemical DNA biosensors was carried out. We also studied the effect of the functionalization by different linear and branched chains on the structure of polypyrrole nanomaterials and their effects on the number of immobilized biomolecules. The strategy adopted for the development of these materials is a biosensor construction carried out step by step. The various layers constituting it have been characterized by different surface techniques such as electrochemical and optical techniques, scanning electron microscopy and atomic force electron microscopy. The properties of the biosensors were monitored through the redox properties of the ferrocenyl groups. The electrochemical detection of the target DNA evaluated with these biosensors based on polypyrrole nanostrcutrue compared to the results obtained with a biosensor based on polypyrrole compact layer shows a higher detection sensitivity.This demonstrates a potential for using the high surface area of polypyrrole nanostructures as a transducer and the use of soft modification approaches. The biosensors were applied to the detection of DNA in real samples of genomic DNA of Mycobacterium tuberculosis and mutated DNA with resistance to rifampicin. The biosensors based on polypyrrole nanostructures demonstrate potential application in DNA detection and the ability to discriminate the mutant rpoB gene and could be used as a platform in biosensor technology.

Finite-Difference Time-Domain method (FDTD) is based on numerical solution of Maxwell's equations, nowadays widely used for simulating optical response of photonic structures. This paper provides brief introduction to the FDTD method and several important extensions which make the basic code much more versatile. In order to broaden analysis of photonic structures, transfer matrix method (TMM) is also involved. The code is firstly tested using simple model structures which optical response might be compared with different numerical or even analytical approaches. Debugged code is used to improve photonic crystals for enhanced sensitivity of biosensing devices based on refractive index changes of sensed medium. Last but not the least, properties (sensitivity and Q-factor of resonant peak) of holey waveguide are investigated in one-, two- and three-dimensional simulation. It is shown here, that even this simple structure may compete with complex photonic crystals in the field of biosensors. Powered by TCPDF (www.tcpdf.org)

La détection précoce de biomarqueurs de maladies telles que le cancer représente un intérêt majeur dans le processus de traitement. En effet, un diagnostic avancé augmente considérablement les chances de réussite du traitement. En pratique, cela nécessite des outils permettant de détecter rapidement d'infimes quantités de composants biologiques (anticorps, protéines, ADN...) au sein d'échantillons réels tels que du sang ou du sérum.Ces dernières années, les avancées et progrès technologiques en matière de micro et nanofabrication ont permis le développement des Micro et Nano Systèmes Electro-Mécaniques (M/NEMS) dans de nombreux domaines d'application et notamment celui de la détection de masse. Ainsi, des nano-capteurs de masse atteignant des résolutions de l'ordre du yoctogram ($10^{-24}g$), soit la masse d'un seul proton ont été développés. De telles résolutions permettraient d'utiliser ces capteurs à des fins de biodétection. Ces résultats ont cependant été obtenus sous vide ce qui est incompatible avec le monde biologique. Immergés en liquide, les performances des M/NEMS traditionnels sont drastiquement réduites notamment à cause de l'amortissement du au fluide. Un nouveau type de résonateur à base de microdisques optomécaniques résonants a ainsi vu le jour démontrant un fort potentiel pour la détection en milieu liquide. Là où les méthodes classiques de transduction électriques des M/NEMS éprouvent des difficultés en liquide, l'exceptionnelle sensibilité de la transduction optomécanique permet de surmonter ce problème.Dans ce cadre, ces travaux de thèse visent à développer un biocapteur à base de microdisques optomécaniques résonants en silicium pour la détection biologique en milieu liquide. Le design, la fabrication ainsi que la caractérisation complète de ces capteurs est décrite. Enfin, une preuve de concept de détection de virus T5 à une concentration de quelques pM à l'aide de ces microdisques est également présentée. / Early detection of disease's biomarkers such as cancer represents a major interest in the treatment process. Indeed, a diagnosis at an early stage considerably increases the chance of the treatment to be successful. Practically, tools allowing the rapid detection of tiny amount of biological compounds (antibodies, proteins, DNA...) in real samples such as blood or serum are needed.Over the last years, the advances and progresses of micro and nanofabrication techniques have allowed the development of Micro-Nano Electro Mechanical Systems (M/NEMS) in various fields of application including mass sensing. Thus, nano mass sensors reaching resolution down to the yoctogram level, the equivalent of a single proton have been demonstrated. Such resolution limit would theoretically allow these sensors to be used as potential biosensors. These results were nonetheless obtained in vacuum conditions which is incompatible with the biological world. Immersed in fluid, the performance of traditional M/NEMS are drastically degraded mostly due to the large viscous damping. A new type of object in the form of optomechanical microdisk resonators have recently emerged, demonstrating a huge potential for sensing in liquid. While M/NEMS classical electrical or optical transduction methods become very challenging in liquid, the astonishing sensitivity of the optomechanical transduction overcomes this major issue.In this context, this thesis work aims at developing a biosensor based on silicon optomechanical microdisk resonators for biosensing in liquid. Design, fabrication along with the complete characterization of theses devices is described. Eventually, a proof-of-concept of T5 virus detection at the pM level using these microdisks is presented.