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ISEP - DM – Engenharia Biomédica

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http://hdl.handle.net/10400.22/18076

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Browsing ISEP - DM – Engenharia Biomédica by advisor "Domingues, Valentina Maria Fernandes"

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  • Detection of Polymorphisms Associated to Cardiovascular Pharmacogenetic Using Electrochemical Genosensor
    Publication . Marshaha, Aya; Domingues, Valentina Maria Fernandes
    Cardiovascular diseases (CVD) are one of the leading causes of death worldwide. To prevent CVD events and further loss of life anticoagulants, like warfarin, are prescribed and administered to patients. Nevertheless, warfarin presents a strict therapeutic window which can lead to other CVD complications or treatment prevention. Studies have shown that genetic determinants, such as single-nucleotide polymorphisms (SNP), can alter the function of enzymes involved in vitamin K’s and warfarin’s metabolism and action, causing interindividual differences in the drug response. In this thesis, electrochemical genosensors for the detection of the vitamin K epoxide reductase complex (VKORC1) gene polymorphisms was developed and optimized. This device detects the electrochemical signal of the hybridization reaction between two complementary DNA sequences. Analyzing public databases, two DNA target probes (52 bp) with the A and G nucleotide variants were selected and designed. The construction of the genosensor was obtained in various steps (i) Sensorial phase: creation of the thiolated DNA and mercapto-hexanol mixed self-assembled monolayer (SAM) on the screen-printed gold electrode (SPGE) surface; (ii) Promotion of the DNA hybridization reaction in a sandwich format (to increase the selectivity) and (iii) Electrochemical detection of the hybridization reaction by evaluating the reduction reaction of tetramethylbenzidine/peroxide (TMB/H2O2) substrate. After optimizing all of the analytical parameters, the calibration curves for both sequences were determined. A linear correlation between the analytical signal (electrochemical current) and the corresponding DNA target concentration were obtained in the 0.50 and 1.00 nM range. Therefore, the developed electrochemical genosensor is a promising and low-cost analytical tool to determine and discriminate an individual’s genotype and predict the adequate warfarin dose
    2021Master thesis Open access Show more