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A novel class of nanobiosensor was developed by integrating a 27-nucleotideAluIfragment of swine cytochrome b (cytb) gene to a 3-nm diameter citrate-tannate coated gold nanoparticle (GNP). The biosensor detected 0.5% and 1% pork in raw and 2.5-h autoclaved pork-beef binary admixtures in a single step without any separation or washing. The hybridization kinetics of the hybrid sensor was studied with synthetic andAluIdigested real pork targets from moderate to extreme target concentrations and a sigmoidal relationship was found. Using the kinetic curve, a convenient method for quantifying and counting target DNA copy number was developed. The accuracy of the method was over 90% and 80% for raw and autoclaved pork-beef binary admixtures in the range of 5–100% pork adulteration. The biosensor probe identified a target DNA sequence that was several-folds shorter than a typical PCR-template. This offered the detection and quantitation of potential targets in highly processed or degraded samples where PCR amplification was not possible due to template crisis. The assay was a viable alternative approach of qPCR for detecting, quantifying and counting copy number of shorter size DNA sequences to address a wide ranging biological problem in food industry, diagnostic laboratories and forensic medicine.

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A simple method for the fabrication of nano-gaps less than 50 nm by using conventional photolithography combined with patterned-size reduction techniques is presented. Silicon material is used to fabricate the nano-gap structure and gold is used for the electrode. Two chrome masks are proposed to complete this work, the first mask for the nano-gap pattern and a second mask for the electrode. The method is based on the control of the coefficients (temperature and time) with an improved pattern size resolution by thermal oxidation. With this technique, there are no principal limitations to fabricating nanostructures with different layouts down to several nanometers in dimension. In this work, the proposed method is experimentally demonstrated by preparing the nano-gaps on a Si-SiO 2 substrate down to dimensions of 50 nm. The optical characterization that is applied to check the nano-gap structure is by using the scanning electron microscope (SEM).