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Updated: Aug 9, 2026
Fabrication of Electrochemical-DNA Biosensors for the Reagentless Detection of Nucleic Acids, Proteins and Small Molecules
Published on: June 1, 2011
Development of an electrochemical DNA-based biosensor for the detection of the cardiovascular
Stephanie L Morais1, Júlia M C S Magalhães2, Valentina F Domingues1
1REQUIMTE/LAQV, ISEP, Polytechnic of Porto, Rua Dr. António Bernardino de Almeida, 4249-015, Porto, Portugal.
Insights
A novel DNA biosensor detects the CYP2C9*3 genetic polymorphism, crucial for warfarin dosing. This rapid, low-cost device aids in preventing adverse drug reactions like hemorrhage or thromboembolism in cardiovascular patients.
Area of Science:
- Biomedical Engineering
- Pharmacogenomics
- Analytical Chemistry
Background:
- Cardiovascular diseases are leading causes of death, with warfarin commonly prescribed as an anticoagulant.
- Warfarin exhibits significant dose variability, increasing risks of bleeding or clotting due to factors like CYP2C9*3 genetic polymorphism.
- Accurate, real-time detection of genetic variations influencing drug metabolism is essential for personalized medicine.
Purpose of the Study:
- To develop a selective, rapid, and cost-effective electrochemical biosensor for detecting the CYP2C9*3 genetic polymorphism.
- To enable pre-prescription warfarin dosing adjustments based on individual genetic profiles.
- To mitigate risks associated with warfarin therapy, such as hemorrhage and thromboembolism.
Main Methods:
- Designed specific DNA probes for wild-type (A) and variant (C) CYP2C9*3 alleles.
- Immobilized DNA capture probes on screen-printed gold electrodes using a self-assembled monolayer.
- Employed a sandwich assay format with enzymatic amplification for enhanced electrochemical detection via chronoamperometry.
Main Results:
- Achieved a limit of detection of 42 pM for both target DNA sequences.
- Demonstrated successful discrimination between synthetic DNA targets and denatured genomic DNA.
- Validated the biosensor's ability to identify non-variant (A/A) and heterozygous (A/C) CYP2C9*3 genotypes.
Conclusions:
- The developed disposable DNA-based biosensor offers a promising tool for rapid CYP2C9*3 genotyping.
- This technology can facilitate personalized warfarin therapy, improving patient safety and treatment efficacy.
- The biosensor's selectivity and sensitivity support its clinical application in pharmacogenetic testing.
Abstract:
Cardiovascular diseases are among the major causes of mortality and morbidity. Warfarin is often prescribed for these disorders, an anticoagulant with inter and intra-dosage variability dose required to achieve the target international normalized ratio. Warfarin presents a narrow therapeutic index, and due to its variability, it can often be associated with the risk of hemorrhage, or in other patients, thromboembolism. Single-nucleotide polymorphisms are included in the causes that contribute to this variability. The Cytochrome P450 (CYP) 2C9*3 genetic polymorphism modifies its enzymatic activity, and hence warfarin's plasmatic concentration. Thus, the need for a selective, rapid, low-cost, and real-time detection device is crucial before prescribing warfarin. In this work, a disposable electrochemical DNA-based biosensor capable of detecting CYP2C9*3 polymorphism was developed. By analyzing genomic databases, two specific 78 base pairs DNA probes; one with the wild-type adenine (Target-A) and another with the cytosine (Target-C) single-nucleotide genetic variation were designed. The biosensor implied the immobilization on screen-printed gold electrodes of a self-assembled monolayer composed by mercaptohexanol and a linear CYP2C9*3 DNA-capture probe. To improve the selectivity and avoid secondary structures a sandwich format of the CYP2C9*3 allele was designed using complementary fluorescein isothiocyanate-labeled signaling DNA probe and enzymatic amplification of the electrochemical signal. Chronoamperometric measurements were performed at a range of 0.015-1.00 nM for both DNA targets achieving limit of detection of 42 p.m. The developed DNA-based biosensor was able to discriminate between the two synthetic target DNA targets, as well as the targeted denatured genomic DNA, extracted from volunteers genotyped as non-variant homozygous (A/A) and heterozygous (A/C) of the CYP2C9*3 polymorphism.
