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Nanotechnology and Biosensors

Baeumner, Antje
Cornell University
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The Objectives of the Multistate Project are: 1. Develop new technologies for characterizing fundamental nanoscale processes 2. Construct and characterize self-assembled nanostructures 3. Develop devices and systems incorporating microfabrication and nanotechnology. Develop a framework for economic, environmental and health risk assessment for nanotechnologies applied to food, agriculture and biological systems 4. Produce education and outreach materials on nanofabrication, sensing, systems integration and application risk assessment. The main goal of the here proposed research is fits into the overall 3rd objective, specifically, the development of biosensors that can rapidly detect foodborne pathogens with limits of detection required to identify on-site contamination of food products. Based on previous biosensors that have demonstrated applicability to food-related detection due to included sample preparation, focus here will be on a novel sensing technology enabling extremely high sensitivity coupled with little hardware requirements and simple user handling.

The specific sub-objectives are: (1) Development of a miniaturized electrochemiluminescent detection unit in microfluidic housing. (2) Detection of foodborne pathogens via antibody recognition in food matrices (3) Investigating the use of photopaper for inexpensive visual readout. Model analytes will be E. coli and Cryptosporidium parvum. Initial food matrices will be apple juice and cider. Based on preliminary data and calculations, the presence of 10 cells per 10 mL sample will be feasible from a food matrix.

The intended outcome of the proposed research is a prototype of a novel biosensor for the rapid detection of foodborne pathogens. This will function as a new platform for pathogen detection with the targeted application to food safety. It will serve the challenging conditions observed with food safety detection including difficult matrices, speed, cost and simplicity, and will serve as point-of-food diagnostic device.

The major impact will be the improvement of the safety of our foods as rapid detection will become possible. This will affect our ability to test food prior to distribution and quickly identify causative agents in the unfortunate event of a foodborne outbreak. Also, routine detection can be implemented in HACCP protocols and provide early signs of potential contamination problems. These impacts are also highly relevant to food production and distribution businesses in New York, as well as people living in rural and high density urban locations of the state.

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Non-Technical Summary:
The proposed research addresses the need of ensuring a safe food supply. The contamination of our foods with pathogens and toxins causes a real threat to the safety of the consumers and increases health care costs with multi-billion dollars associated to the treatment of foodborne illnesses. The CDC estimates that 1 in 6 people in the US experience foodborne illness per year resulting in 128,000 hospitalizations and 3000 annual deaths. The most troubling fact may be that the causative agents are not identified for the majority of these cases. The detection of foodborne pathogens and toxins is complex, time consuming and costly. As much as there is a need for point-of-care diagnostics in our health care to better serve patients and reduce health care costs; there is an urgent need for point-of-food diagnostics. These would enable rapid and inexpensive identification of contamination in foods before it is distributed; enable rapid identification in the case of an outbreak and therefore protect wide-spread foodborne illnesses from occurring. Public funding is essential so that emphasis can be put toward the development of inexpensive and simple point-of-food diagnostics to protect children, the elderly and immune-compromised who are the first to suffer severe illness from foodborne pathogens. Knowledge gained by nanotechnology research needs to be translated into prototypes of devices for real-world problems, so that subsequently industrial collaborators can develop commercially viable products, create jobs and protect consumers.

For sub-objective 1, an existing electrochemical microfluidic biosensor with embedded sample preparation of the Baeumner group will be transformed to enable electrochemiluminescence (ECL) detection. The biosensor is made from plastic to enable simple and rapid prototyping as well as inexpensive mass production in the future. Here, a new electrochemical 3-electrode set up will be developed driving the ECL reaction. The signals will be monitored by a simple photodiode. A small black housing will be designed to ensure portability. The molds required for microchannel designs and the electrodes will be fabricated using the Cornell University clean room facilities. Actual prototype fabrication can be carried out in a normal biochemical wet lab. The electrode design, microfluidic channels and the photodiode set up will be optimized to enable highly sensitive ECL detection using ruthenium bipyridyl complexes.
For sub-objective 2, dendrimers tagged with the ruthenium complexes will be covalently coupled to anti-E. coli and anti-C. parvum antibodies. The organisms will be isolated from the food sample using immunomagnetic separation. The magnetic beads will be captured within the microfluidic device and cells will be quantified upon being bound by the reporter antibodies via the ECL reaction. Assay conditions will be optimized.
For sub-objective 3, sensitive photopaper will be investigated as alternative to the photodiode. Photopaper will enable semi-quantitative detection of the ECL reaction that emits light at 620 nm. Assay conditions for signal development on the photopaper will be studied.

Funding Source
Nat'l. Inst. of Food and Agriculture
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Bacterial Pathogens
Food Defense and Integrity
Sanitation and Quality Standards
Risk Assessment, Management, and Communication
Natural Toxins