Proteins aggregates are supramolecular ensembles capable to elicit significant biological responses and alter molecular reactions and quantitative, high throughput monitoring of the aggregation process is needed in high impact fields such as pharmacy and medicine. Very recent results show the possibility of ultrasensitive and conformation specific detection of proteins using sensors with Molecular Imprinted Polymers (MIP’s). Formed in the presence of target analyte, these polymers retain the memory of analyte’s shape.
We propose to take advantage of the heightened and specific recognition properties of MIP’s to build an integrated sensing platform for the simultaneous, specific detection of monomer, dimer and a higher protein aggregate. This device will circumvent the need for prior separation of protein species and will be employed to monitor the aggregation of two proteins, lyzosyme and calcitonin. The innovative platform we envisage consists in nanoarrays with layers of MIP’s specific for monomer, dimer and a higher aggregate, deposited on the tips of gold nanopillars. Detection is based on the principle of impedance restoration when the “holes” in the polymer layer are filled by target protein species.
Very sensitive detection of lyzosyme and calcitonin is anticipated and feasability of MIP micro arrays for monitoring the aggregation process will be assessed through a forced protein degradation study and rigorous validation against a reference method. The project takes a multidisciplinary approach to shed light on the kinetics of orientation of protein species at MIP “holes” and investigate non-specific adsorption, with additional knowledge to be gained through Atomic Force Microscopy, Electrochemical Methods and Surface Plasmon Resonance studies using state-of-the-art equipment available at host institution. Many answers to questions regarding aggregates themselves, their differences in shape and properties are expected to be provided in this way
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