Procedural optimization of the quartz crystal microbalance for rapid detection of Escherichia coli O157:H7

Yimei Angelina Lim

    Research output: ThesisMaster's Thesis

    112 Downloads (Pure)

    Abstract

    [Truncated abstract] The applications of biosensors are rapidly expanding with the increased emphasis placed on the use of technology in the evaluation of food safety and also in military use. The United States food industry carried out 144.3 million microbiological tests in 1999 (Alocilja and Radke, 2003). These numbers are expected to rise with the recently implemented regulatory measures for food safety in the United States. In fact, similar trends in food safety are occurring on a global scale. Furthermore, with the recognition and establishment of Microbial Forensics as a new field of forensics, the interest in biosensor development for the detection of microbes will thrive. Moreover, the recent spate of biocrimes, notably the anthrax scares, has called for newer and improved techniques for the sensitive, rapid and reproducible detection of microbes. Biosensors have the capability to fill this role as an efficient device for microbial detection. There is a wide range of biosensors available for different purposes. In addition, their versatility allows for their overlap in many fields. The quartz crystal microbalance (QCM) is a biosensor that is cost-efficient, sensitive, field-deployable with the ability to perform automated, real-time assays within minutes. The QCM is a mass sensitive device that works on the principle where a change in mass deposited on the crystal is inversely proportional to the change in the resonant frequency of the crystal. Therefore, frequency decreases with increasing mass deposited. The QCM has been used in several studies as a biosensor for the detection of a number of viral and bacterial species. ... High antibody incubation concentration required a shorter antibody incubation duration. Conversely, low antibody incubation concentration required a longer antibody incubation duration. Furthermore, regardless of antibody incubation concentration, a distinct pattern in the rate of antibody binding with time was observed. One hour antigen incubation at ambient room temperature (22.5oC) was sufficient for the efficient binding of the antigens to the immobilized antibody layer. Extension of antigen binding time to 15 hours produced inconsequential differences in readings. The binding efficiency of the quartz crystals after a storage period of 2 to 4 weeks at ambient room temperature (22.5oC) fared better than the crystals that were refrigerated at 4oC. Results showed that 0.2M glycine hydrochloride is a poor reagent for the removal of the antigen layer on the quartz crystals for repeated assay use. The 16-mercaptohexadecanoic acid (MHDA) layer and adsorbed proteins on the quartz crystals can be removed by a mixture of sulphuric acid and hydrogen peroxide, known as a piranha process. This allows the crystals to be repeatedly recoated and reused. Overall, this research provides new insights into the preparation process of the quartz crystals for the specific detection of E. coli O157:H7. Conclusive results have been obtained for several tested parameters and suggestions have been raised for further studies in the optimization of the QCM for the E. coli O157:H7 detection process. With improved knowledge and recognition in the capability of the QCM as a biosensor, the QCM may soon be used in conjunction with conventional techniques for the rapid detection of E. coli O157:H7.
    Original languageEnglish
    QualificationMasters
    Publication statusUnpublished - 2007

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