Objective: The goal of proposed research is to fabricate and use highly piezoelectric microcantilever arrays for ultrasensitive, in-situ, rapid, simultaneous multiple analyte quantification in small sample volumes using electrical means with unprecedented sensitivity (10 -15 g). The proposed piezoelectric microcantilevers with antibodies specific to the target antigen immobilized at the cantilever tip will measure the presence of proteins or pathogens with femtogram (smaller than the mass of a single cell) sensitivity. This represents the ability to detect a single cell or about 1000 molecules in a small volume. We will demonstrate the application with (1) transient protein expression of a recombinant protein, alkaline phosphatase (SEAP) in Chinese Hamster Ovary cells (CHO) during growth in a bioreactor, and (2) early detection of streptococcus at ultra low concentrations , 100 - 10,000 number/ml. Approaches: The device consists of a highly piezoelectric lead magnesium niobate-lead titanate solid solution (PMN-PT) cantilever smaller than 50 um in length coupled to antibody immobilized at the cantilever tip. Binding of target antigens is detected by monitoring the resonance frequency shift. Because of the small sizes, the cantilevers will be capable of detecting a single cell or some 1000 molecules in a small volume. The resonance frequency shift transient will be used to quantify the amount of antigens present in a small volume. In the model CHO bioreactor, the SEAP expression quantification is used as an example to illustrate the proposed methodology. For pathogen detection, microcantilevers developed under this program have the potential to detect streptococcus at a concentration of 100 to 10,000 bacterial/ml, significantly enhancing detection capability. Experiments will be designed to explore lower level of detection in the range of 100 to 10,000 cells/ml. Because the proposed piezoelectric cantilever sensors use electrical signal for actuation and detection, the sensor and all necessary electronics can be organized in a compact form and easily usable in such broad ranging applications such as early disease detection and genomics-inspired proteomics. Results: It is anticipated that as a result of the proposed study, ultra sensitive, rapid, specific, multiple analyte quantification in ultra low concentrations and small sample volumes will be achieved using arrays of highly piezoelectric PMN-PT micro cantilevers of less than 50 um in length with better than10-15 g/Hz sensitivity coupled with antibodies specific to the target antigen immobilized at the cantilever tip with simple electrical means, which has wide ranging applications such early disease detection and genomics-inspired proteomics.
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