The Quartz crystal microbalance (QCM) is a very sensitive mass-detecting device which is based on changes in to the vibrational frequency of quartz crystals after adsorption of substances to a modified crystal surface. In this study a QCM-based biosensor was developed for the rapid diagnosis of influenza viruses and its suitability and limitations were compared with currently available diagnostic methods on 67 clinical samples (nasal washes) received during the 2005 Australian winter. The type-specific and conserved viral M1 proteins of both A/PR/8/34 and B/Lee/40 viruses were used to prepare polyclonal antisera for the development of an ELISA. The limits of detection of ELISAs for the detection of purified A/PR/8/34 and B/Lee/40 ƒnviruses were 20ƒÝg/mL ƒnand 14 ƒÝg/mL using polyclonal antibodies, and 30 ƒÝg/mL and 20 ƒÝg/mL for monoclonal antibodies, respectively. The limit for detection of each virus was 104 pfu/mL, irrespective of whether antisera or monoclonal antibodies were used for capture. Non-purified cell culture-grown preparations of either virus could be detected at 103 pfu/mL The QCM utilised the same reagents used in ELISAs. However, a number of parameters were then further optimised to improve the sensitivity of the tests. These included blocking of non-specific binding, examination of the effects of flow-cell compression, the role of pH, flow rate, antibody concentration and the addition of protein A to the crystal surfaces of the biosensor. The lowest virus concentration that could be detected with the QCM was 104 pfu/mL for egg-grown preparations of both A/PR/8/34 and B/Lee/40, which could be detected within 30 min. However, conjugation of 13 nm gold nanoparticles to a second detector antibody resulted in a 10-fold increase in sensitivity and a detection limit of 103 pfu/mL that could be determined within 1 h. The direct detection of the influenza viruses in nasal samples was not possible by QCM because of the significant frequency fluctuation that was probably caused by the viscosity of the samples. Therefore, an additional culture step of 12 h was required, which increased the processing time to 2 days. The QCM/nanoparticle method was shown to be as sensitive as the standard cell culture method, and the QCM method as sensitive as the shell vial method. The QCM and QCM/nanoparticle methods were shown to be 81 and 87% as sensitive and both were 100% as specific as the real-time polymerase chain reaction. However, for use in rapid diagnosis, improvements are required to remove frequency fluctuation resulting from the direct use of nasal samples.

Résumé indisponible / This PhD work is dedicated to investigate potentialities of phthalocyanines materials to realize a Quartz Crystal Microbalance (QCM) sensor for Benzene, Toluene and Xylenes (BTX) detection in air. The goal is to develop a sensor-microsystem capable of measuring BTX concentrations quantitatively below the environmental guidelines with sufficient accuracy. To achieve these objectives, our strategies mainly focused on experimental works encompassing sensors realization, sensing material characterizations, development of gas-testing facility and sensor testing for different target gases. One of the main aims is to identify most appropriate phthalocyanine material for sensor development. After comparative sensing studies, tert-butyl-copper phthalocyanine based QCM device is found as most sensitive and detail metrological characteristics are further investigated. Results show repeatable, reversible and high magnitude of response, low response and recovery times, sub-ppm range detection limit, high resolutions and combined selectivity of BTX gases among common atmospheric pollutants. Special focus is given to understand the gas/material interactions which are achieved by (a) XRD and SEM characterizations of sensing layers, (b) formalization of a two-step adsorption model and (c) assessing extent of diffusion of target gas in sensing layer. At last, possible ageing of sensor and suitable storage conditions to prevent such effect are investigated.