The present research focused on the development of a new methodology to assess the strength of the interaction between vaccine antigens and elicited polyclonal antibodies through SPR biosensors. Quantifying the binding strength of polyclonal antibodies is of first importance to evaluate the quality of the vaccine as well as to increase the scientific knowledge of immune protection mechanisms. To now the development of such tool has been complicated by the non-specific binding caused by high protein abundance in the blood and serum samples but also by the way of interpreting the data resulting from multi-interaction events measured at the same time. At first, we unsuccessfully tried to segregate the individual affinity contribution of each antibody population by measuring the signal as the sum of singular interactions. Differentiation of the singular contribution would have needed the fulfillment of the “additivity” hypothesis, meaning that each antibody bind identically alone or in mixture with other antibody. This hypothesis was not met and mathematical assessment by the sum of singular contribution led to fitting results that did not reflect the biological reality. It was therefore decided to switch the analysis method and to measure the end association binding level reached by the different samples injected at the same specific antibody content. The dissociation behavior was interpreted by the percentage of binding after long and fixed dissociation time. In a first application, we compared the antibodies elicited by two different commercially available vaccines and we showed that the binding interaction was not concentration dependent as, highly different levels were reached when injecting identical antibody concentration. No statistical significant difference was observed between both vaccines. Research firstly focused on the decrease of the non-specific binding and we found that ionic strength was a key parameter, increasing the buffer salt concentration reduced the non-specific binding without diminishing the binding strength. The sample composition was also a key parameter and purifying the IgG allowed to decrease dramatically the undesired binding events. A second application aimed at showing the equivalence between two different antigen constructions for two antibodies population. Even if identical antigen level immobilization is a challenge, the methodology is completely suitable to perform a 2-dimensional comparison (ligand and analyte). A last application was dedicated to the comparison between D and Q-pan Flu vaccines, and results showed that there was no statistical evidence of significant differences between both vaccines. End association level correlated well with haemagglutination inhibition assay at least when serum samples were not diluted at the same antibody content. This last application also showed that throughput may be extended to more than 50 samples per 80 hours / Doctorat en Sciences agronomiques et ingénierie biologique / info:eu-repo/semantics/nonPublished

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Point-of-care testing brings diagnosis and treatment monitoring to the site of the patient. It heavily relies on biosensors, which leverage the interactions between a target biomarker and a bioreceptor, to deliver fast and accurate results. However, non-specific binding of molecules and microorganisms on the biointerface can interfere with biomarker-bioreceptor interactions and diminish a biosensor’s sensitivity, specificity, and stability. In turn, this can lead to false diagnoses and ineffective treatments. Omniphobic-lubricant infused (OLI) coatings exhibit slippery, self-cleaning characteristics that repel untargeted molecules and microorganisms to augment the biosensor’s performance. In this work, we investigate the proficiency of OLI coatings in two specific applications: dissolved oxygen sensing and DNA biosensing. First, in water quality monitoring, an OLI coating is applied to the selectively permeable membranes of a dissolved oxygen sensor. Over a three-week incubation period in an environment with accelerated bacterial growth, the coated membranes exhibit a 160% higher reproducibility (10% deviation in sensitivity) and lower biofilm formation (96° static contact angle) in comparison to unmodified membranes (26%, 32°). The second application is in DNA biosensing, where a novel OLI coating uses carbon dioxide plasma activation to embed oligonucleotide probes. It demonstrates an optimized balance of slippery repellency (76° static contact angle, 10° sliding angle) and biosensing functionality, 19% longer clotting times than conventional blocking conditions, and equal sensitivity to PLL-PEG when capturing target DNA in whole blood. Going forward, our research will continue to expand the use of OLI coatings in biosensing applications, particularly exploiting its antibiofouling and anticoagulative capabilities. / Thesis / Master of Applied Science (MASc) / Biosensors are an integral tool in delivering quick and accurate point-of-care diagnosis and treatment monitoring. However, their performance can be impeded by the non-specific binding of undesirable molecules and microorganisms on the sensing surface. Omniphobic lubricant-infused (OLI) coatings have been shown to suppress biofouling and blood clotting on surfaces through exceptional repellency. This thesis focuses on the implementation of OLI coatings in biosensing applications. It investigates the antibiofouling capacity of an OLI coating on a membrane for dissolved oxygen detection. Then, it discusses a novel coating with integrated DNA biosensing functionality for working directly with blood samples. The enclosed work demonstrates that the OLI coating empowers biosensors to deliver more effective point-of-care testing.

While significant progress has been made in the treatment and management of cancer, challenges remain because of the complexity and the heterogeneous nature of the disease. The improvement that has been seen in survival rates reflects advancements not only in treatment, but also in early stage detection and diagnostics for certain cancers. In particular, early stage detection and treatment of cancer before it has metastasized to other organs has resulted in a dramatic improvement in patient survival rates. One area of research that has shown considerable promise in further advancing diagnostics and early cancer detection is nanotechnology. Specifically, semiconductor and metal nanoparticles have great potential to provide advanced technology platforms for ultrasensitive and multiplexed detection of disease markers and probe disease on the molecular level. Because they are in the same size regime as biological molecules, these nanoparticles exhibit unique interactions with proteins, nucleic acids and other biomarkers of interest for detecting and diagnosing disease. However, high-quality nanoparticles are often unsuited for use in complex biological environments because of their coatings and surface chemistry. In this work, we describe the design and development of polymer-coated nanoparticle imaging agents for use in blood, cell and tissue diagnostic applications. Low-molecular weight, amphiphilic polymers capable of noncovalent interactions with nanoparticle surface ligands and the aqueous environment were synthesized and characterized for use in nanoparticle coating applications. We demonstrate that the hydrophobic and hydrophilic interactions between the nanoparticle surface, the amphiphilic polymer and the aqueous solvent were able to drive the coating and water solubilization of quantum dots. Novel nanoparticle synthetic techniques were also developed using the amphiphilic polymers in a one-pot method to make high quality semiconductor and gold nanoparticles and stabilize and encapsulate the particles for transfer into water. Using the polymer functional groups as multidentate ligands, nanoparticles were synthesized with a high degree of size control and increased stability. In addition, by performing the synthesis in a noncoordinating amphiphilic solvent such as polyethylene glycol, nanoparticles were immediately transferred to water with the excess polymer forming a water soluble coating. Next, nanoparticle surface charge and how it relates to the nonspecific binding of nanoparticles in cells, tissues and other complex biological samples was studied. We have found that highly charged (negative and positive) particles exhibit significant nonspecific binding to biomolecules and other cellular components in biological environments. By reducing the surface charge through the incorporation of hydroxyl functional groups, we have nearly eliminated the nonspecific binding of quantum dots in blood, cells and tissues. Moreover, through crosslinking and altering the surface chemistry of the polymer-coated quantum dots, we have increased the stability of the nanoparticles while maintaining a small hydrodynamic size. Finally, we have investigated the use of the low-binding, hydroxyl quantum dots in tissue staining applications, where nonspecific binding presents a considerable challenge to detection sensitivity and specificity. A number of biomolecule conjugation techniques were examined for the coupling of quantum dots to antibody targeting molecules and preliminary staining experiments were performed.