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NEMS-based protein analysis

Regarding most applications for life science, as enzymatic studies, quantitative proteomics or downstream monitoring of drug discovery, require high sensitive and fast methods for the accurate determination of protein concentration. Typically, the quantity of proteins is analysed by colorimetric dye-labeled assays as (e.g. Bradford, BCA), or by means of UV absorption spectroscopy, since it is fast and covers a large concentration range from milligram down to a few hundred picogram. However, due to assay-specific limitations as interferences from chemicals prevalent buffer solutions or the lack of sensitivity for unknown mixtures, no existing method can provide both high accuracy and selectivity.

With the uprising field of nanoelectromechanical systems (NEMS), novel sensor techniques could demonstrate outstanding sensitivities for IR detection. I propose an instrument based on NEMS to enable a label-free protein quantitation combining both, UV and IR spectroscopy, improving the minimal necessary amount for detection from nanograms to single femtograms. The utilized method relies on the thermal expansion and corresponding frequency detuning of a nanomechanical resonator, made of low (tensile-) stress Silicon Nitride. In this approach, the analyte of diluted proteins will be nebulized to an aerosol and directly sampled to the resonator by using a non-diffusive limited method. Subsequently, the photothermal induced heating of the sample by absorbed UV/IR light is directly converted into a measureable frequency shift.

Compared to former studies, that successfully demonstrated such nanomechanical IR analysis down to single picograms; my developed post-treatment method, to control the tensile stress by means of oxygen plasma, enables a significant enhancement of the sensitivity by up to three orders of magnitude. In addition, the proposed project includes a major revision of the sampling efficiency to provide a quantitative, automated aerosol sampling and the implementation of a full electronic transduction with minimal influence to the sensitivity. Including a temperature-controlling element, it will be therewith possible to provide a dynamic range from femtogram to milligram concentrations. By the till now unique combination of UV and IR absorption data in one single measurement, it should be possible to identify and quantify proteins even from complex mixtures. This breakthrough would constitute a combined UV/IR NEMS protein quantification prototype with a minimum quantification range equal or better than that of state-of-the-art UV spectroscopy.



 Austrian Academy of Sciences