Designing and developing biosensors for evaluation of meat adulteration in cooked sausages.

Abstract

Introduction: Meat products as a rich source of protein, has particular importance in the diet. So it is necessary to develop an accurate method for quality control and to detect related adulteration. Aim: The design and manufacture of electrochemical and SPR-based biosensors to identify and determine target oligonucleotides and their application in real samples and compare the results with PCR results.Methode: A unique probe using donkey sequence was designed and its species specificity was confirmed bioinformatically. A novel species-specific electrochemical DNA probe (locked nucleic acid, LNA) was synthesized commercially and implemented in a construction of DNA-based electrochemical biosensor for sensitive, convenient and selective detection of donkey adulteration. The electrochemical behavior of the fabricated genosensor was studied by linear sweep, square wave, differential pulse voltammetry and electrochemical impedance spectroscopy techniques. To design the SPR-based biosensor, Stre@GNSs were synthesized via physicochemical adsorption of streptavidin on GNSs surface and characterized by ultraviolet and visible spectrophotometer (UV–Vis) and transmission electron microscope (TEM). Then, the direct detection assay was compared to sandwich format enhanced with gold nanostars. Results: The electrochemical biosensor under the optimal hybridization conditions, presented an excellent sensitivity toward donkey marker with LLOQ as low as 148 pM and relative standard deviation of 0.1581%, in simple hybridization system. The SPR-based biosensor showed target DNA could be detected with the lower limit of quantification (LLOQ) of 1.0 nM with a relative standard deviation (RSD, n=3) of 0.85%, in sandwich format assay. Discussion and Conclusion: QRT-PCR analysis was performed for DNA extracted from the sample and the results of this method were compared with biosensors results. Genosensor and SPR-based biosensor were both able to detect different percentages of donkey meat in sausages prepared from bovine-donkey binary mixtures. These biosensors were more sensitive than qRT-PCR for detection donkey meat adulteration.