Electrochemical DNA Biosensor Research Articles

A novel and ultrasensitive label-free electrochemical DNA biosensor for Trichomonas vaginalis detection based on a nanostructured film of poly(ortho-aminophenol)

Trichomoniasis, as a major public health concern, is the world’s most common sexually transmissible disease. This infection is caused by a protozoan parasite called Trichomonas vaginalis (TV) that can lead to reproductive and genital area problems. Accordingly, it is of great significance to expand new methods for increasing the sensitivity of TV detection. In this study, a novel label-free electrochemical DNA biosensor for TV quantitation was constructed based on an electropolymerized poly(ortho-aminophenol) thin film simultaneously acting as a transducer as well as a redox indicator. The redox-active polymeric film was utilized to covalent immobilization of a specific thiolated DNA probe. The hybridization process was then monitored by differential pulse voltammetry. The proposed biosensor detected a synthetic TV target sequence with a calibration sensitivity of − 0.0113 µA (log (concentration/mol L−1))−1, a linear concentration range of 1.0 × 10−20 to 1.0 × 10−12 mol L−1, and a detection limit of 3.9 × 10−21 mol L−1. It also showed an ability to differentiate between the complementary sequence and the base-mismatched and non-complementary sequences with a nice selectivity. Moreover, the designed biosensor was able to detect the TV genome with a calibration sensitivity of − 0.0774 µA (log (concentration/ng µL−1))−1, a linear range of 0.55–64 ng mL−1 and a detection limit of 1.0 pg µL−1.

A new strategy to design label-free electrochemical biosensor for ultrasensitive diagnosis of CYFRA 21–1 as a biomarker for detection of non-small cell lung cancer

Lung cancer is one of the most dangerous cancers with high mortality rate among other cancers therefore, early detection of this cancer is very important. Many studies have been reported in ways of diagnostic lung cancer early. According to reports, one of the most important biomarkers to detect lung cancer is Cytokeratin 19 fragment 21–1 (CYFRA21-1), which is significantly related to non-small cell lung cancer, in particular, squamous cell carcinoma. Thus, finding a new method for the early diagnosis of CYFRA 21–1 (DNA target probe) is essential. In the present report, we design a novel label-free electrochemical DNA-biosensor related to the signal of guanine oxidation. The proposed DNA biosensor is fabricated by a modified glassy carbon electrode (GCE) with reduced-graphene oxide (rGO), poly pyrrole (PPy), silver nanoparticles (AgNPs) and single-strand DNA (ssDNA as capture probe) GCE/rGO/PPy/AgNPs/ssDNA. The differential pulse voltammetry (DPV) and cyclic voltammetry (CV) techniques are used to verify the hybridization process between capture and target probes. Electrochemical impedance spectroscopy (EIS), energy diffraction X-ray (EDX) and field-emission scanning microscopy (FE-SEM) techniques are applied to the characterization of different modified GCE surfaces as well as X-ray diffraction (XRD) for graphene oxide synthesis. The XRD pattern of the synthesized GO that its diffraction peak appears at 10.2. The applied CV and DPV for the guanine oxidation are determined under optimal conditions. The label-free DNA biosensor showed a great result for the determination of CYFRA21-1 with a wide linear range from two consecutive linear relationships of peak current and CYFRA21-1 concentration were found (1.0 × 10−14 - 1.0 × 10−10 M, R2 = 0.9936 and 1.0 × 10−9 - 1.0 × 10−6 M, R2 = 0.9955). Proposed electrochemical biosensor displayed low detection limit (2.4 fM).

Comparison of two pyrolytic graphite representatives in the construction of hybrid electrochemical DNA biosensors for monitoring DNA damage

• Fabrication of hybrid electrochemical DNA biosensors via electrostatic adsorption. • In-depth optimization process of the most crucial biosensor parameters. • Large potential window allowing dsDNA cathodic/anodic processes at single electrode. • First direct monitoring of DNA damage via cathodic signals at a carbon electrode. Electrochemical DNA biosensors represent a variety of versatile analytical tools successfully used in many important applications. Their utilization in the area of monitoring DNA damage is considered as the prominent field that can provide us with an improved general knowledge regarding DNA damaging events occurring in vivo by an unorthodox overall insight employing the various electrochemical methods. Herein, we present development, optimization process, and subsequent analytical testing of the novel hybrid electrochemical DNA biosensors based on the two representatives of the pyrolytic graphite – “basal-plane” pyrolytic graphite (BPPG) and “edge-plane” pyrolytic graphite (EPPG) – and low-molecular-weight double-stranded DNA (dsDNA). Closer resolution of the nature of the oxidation/reduction signals of the dsDNA, as well as the most important optimization of parameters securing satisfying operational stability of the proposed hybrid biosensors, is investigated. Subsequent analytical testing of the novel hybrid biosensors and the evaluation of their advantages/disadvantages in terms of monitoring DNA damage caused by an UV light irradiation employing various voltammetric methods (square wave (SWV), linear sweep (LSV), or cyclic (CV) voltammetry) and electrochemical impedance spectroscopy (EIS) is also presented.

Recent Progresses in Electrochemical DNA Biosensors for MicroRNA Detection.

MicroRNAs (miRNAs), as the small, non-coding, evolutionary conserved, and post-transcriptional gene regulators of the genome, have been highly associated with various diseases such as cancers, viral infections, and cardiovascular diseases. Several techniques have been established to detect miRNAs, including northern blotting, real-time polymerase chain reaction (RT-PCR), and fluorescent microarray platform. However, it remains a significant challenge to develop sensitive, accurate, rapid, and cost-effective methods to detect miRNAs due to their short size, high similarity, and low abundance. The electrochemical biosensors exhibit tremendous potential in miRNA detection because they satisfy feature integration, portability, massproduction, short response time, and minimal sample consumption. This article reviewed the working principles and signal amplification strategies of electrochemical DNA biosensors summarized the recent improvements. With the development of DNA nanotechnology, nanomaterials and biotechnology, electrochemical DNA biosensors of high sensitivity and specificity for microRNA detection will shortlybe commercially accessible.

Mapping the DNA Damaging Effects of Polypyridyl Copper Complexes with DNA Electrochemical Biosensors.

Several classes of copper complexes are known to induce oxidative DNA damage that mediates cell death. These compounds are potentially useful anticancer agents and detailed investigation can reveal the mode of DNA interaction, binding strength, and type of oxidative lesion formed. We recently reported the development of a DNA electrochemical biosensor employed to quantify the DNA cleavage activity of the well-studied [Cu(phen)2]2+ chemical nuclease. However, to validate the broader compatibility of this sensor for use with more diverse—and biologically compatible—copper complexes, and to probe its use from a drug discovery perspective, analysis involving new compound libraries is required. Here, we report on the DNA binding and quantitative cleavage activity of the [Cu(TPMA)(N,N)]2+ class (where TPMA = tris-2-pyridylmethylamine) using a DNA electrochemical biosensor. TPMA is a tripodal copper caging ligand, while N,N represents a bidentate planar phenanthrene ligand capable of enhancing DNA interactions through intercalation. All complexes exhibited electroactivity and interact with DNA through partial (or semi-) intercalation but predominantly through electrostatic attraction. Although TPMA provides excellent solution stability, the bulky ligand enforces a non-planar geometry on the complex, which sterically impedes full interaction. [Cu(TPMA)(phen)]2+ and [Cu(TPMA)(DPQ)]2+ cleaved 39% and 48% of the DNA strands from the biosensor surface, respectively, while complexes [Cu(TPMA)(bipy)]2+ and [Cu(TPMA)(PD)]2+ exhibit comparatively moderate nuclease efficacy (ca. 26%). Comparing the nuclease activities of [Cu(TPMA)(phen)] 2+ and [Cu(phen)2]2+ (ca. 23%) confirms the presence of TPMA significantly enhances chemical nuclease activity. Therefore, the use of this DNA electrochemical biosensor is compatible with copper(II) polypyridyl complexes and reveals TPMA complexes as a promising class of DNA damaging agent with tuneable activity due to coordinated ancillary phenanthrene ligands.

Strategies for the preparation of non-amplified and amplified genomic dengue gene samples for electrochemical DNA biosensing applications.

The application of electrochemical DNA biosensors in real genomic sample detection is challenging due to the existence of complex structures and low genomic concentrations, resulting in inconsistent and low current signals. This work highlights strategies for the treatment of non-amplified and amplified genomic dengue virus gene samples based on real samples before they can be used directly in our DNA electrochemical sensing system, using methylene blue (MB) as a redox indicator. The main steps in this study for preparing non-amplified cDNA were cDNA conversion, heat denaturation, and sonication. To prepare amplified cDNA dengue virus genomic samples using an RT-PCR approach, we optimized a few parameters, such as the annealing temperature, sonication time, and reverse to forward (R/F) primer concentration ratio. We discovered that the generated methylene blue (MB) signals during the electrochemical sensing of non-amplified and amplified samples differ due to the different MB binding affinities based on the sequence length and base composition. The findings show that our developed electrochemical DNA biosensor successfully discriminates MB current signals in the presence and absence of the target genomic dengue virus, indicating that both samples were successfully treated. This work also provides interesting information about the critical factors in the preparation of genomic gene samples for developing miniaturized PCR-based electrochemical sensing applications in the future. We also discuss the limitations and provide suggestions related to using redox-indicator-based electrochemical biosensors to detect real genomic nucleic acid genes.

Interaction of the Anticancer Drug Abiraterone with dsDNA

The electroanalytical characteristics of double-stranded DNA (dsDNA) and the complex of dsDNA and the antitumor drug abiraterone acetate (AA) were studied by differential pulse voltammetry. The effect of abiraterone acetate on dsDNA was shown, which was registered by alteration the intensity of electrochemical oxidation of purine heterocyclic bases guanine and adenine using screen printed electrodes modified with functionalized carbon nanotubes. The binding constants (Kb) of the [dsDNA-AA] complex for guanine and adenine were 1.63×104 M-1 and 1.93×104 M-1, respectively. The electrochemical coefficients of the toxic effect were calculated as the ratio of the intensity of the electrochemical oxidation signals of guanine and adenine, in the presence of abiraterone acetate to the intensity of the electrooxidation signals of these nucleobases without drug (%). At concentrations of abiraterone acetate exceeding 60 μM, a decrease in the currents of electrochemical oxidation of guanine and adenine by 50% or more is recorded. Based on the analysis of electrochemical parameters and values ​​of binding constants, an assumption was made about the mechanism of interaction of abiraterone acetate with DNA, mainly due to the formation of hydrogen bonds with the minor groove. An electrochemical DNA biosensor was first used to study the mechanism of interaction of the anticancer drug abiraterone acetate with dsDNA.

A signal-on electrochemical DNA biosensor based on exonuclease III-assisted recycling amplification.

DNA electrochemical detection technology has attracted tremendous interest in recent years. However, a facile and sensitive method for the detection of the disease indicators or genes is still waiting. Herein, we constructed a signal-on electrochemical platform for detecting the manganese superoxide dismutase (MnSOD) gene by incorporating a redox electrochemical signal probe (methylene blue) and exonuclease III-assisted target recycling signal amplification strategy. The sensor was prepared by self-assembly of a capture DNA probe of thiol-modified on GCE with gold electrodeposition. In the presence of target DNA, the exonuclease III can cleave the duplexes formed by the target DNA and the redox-labeled hairpin probes, release the target DNA and produce a residual sequence. The target DNA can continue to hybridize with the hairpin probe for the next cycle of amplification. The residual sequence hybridized with the surface-immobilized capture probes on AuNPs-modified GCE to generate a significantly amplified redox current. In particular, the redox current value of the resultant sensor showed a linear relationship with MnSOD gene concentration in the range of 1-104 pM with the detection limit as low as 0.3 pM. Furthermore, the sensor has excellent specificity and can distinguish single-base mismatch from perfectly matched target DNA. The sensor is fast in operation, and simple in design for detecting different DNA sequences or DNA identification by selecting the appropriate probe sequence, thus shedding light on a good promising application when encountering disease outbreaks or for the early clinical diagnosis of gene-related diseases.

Label-based and label-free electrochemical DNA biosensors for the detection of viruses: A review

Label-based and label-free electrochemical DNA biosensors for the detection of viruses: A review

Electrochemical Resistive DNA Biosensor for the Detection of HPV Type 16

In this work, a low-cost and rapid electrochemical resistive DNA biosensor based on the current relaxation method is described. A DNA probe, complementary to the specific human papillomavirus type 16 (HPV-16) sequence, was immobilized onto a screen-printed gold electrode. DNA hybridization was detected by applying a potential step of 30 mV to the system, composed of an external capacitor and the modified electrode DNA/gold, for 750 µs and then relaxed back to the OCP, at which point the voltage and current discharging curves are registered for 25 ms. From the discharging curves, the potential and current relaxation were evaluated, and by using Ohm’s law, the charge transfer resistance through the DNA-modified electrode was calculated. The presence of a complementary sequence was detected by the change in resistance when the ssDNA is transformed in dsDNA due to the hybridization event. The target DNA concentration was detected in the range of 5 to 20 nM. The results showed a good fit to the regression equation ∆Rtotal (Ω) = 2.99 × [DNA] + 81.55, and a detection limit of 2.39 nM was obtained. As the sensing approach uses a direct current, the electronic architecture of the biosensor is simple and allows for the separation of faradic and nonfaradaic contributions. The simple electrochemical resistive biosensor reported here is a good candidate for the point-of-care diagnosis of HPV at a low cost and in a short detection time.

Ultrafast Electrochemical DNA Biosensor Based on Electrical Potential-Assisted Hybridization Using Differential Pulse Voltammetry (DPV)

A simple electrochemical DNA (E-DNA) biosensor for rapid and specific determination of DNA was developed based on electrical potential-assisted hybridization. The E-DNA biosensor was constructed by a capture probe immobilized on the gold electrode. An external alternating voltage was applied to the electrode interface during DNA hybridization between the capture probe with its target chain and a ferrocene-labeled peptide nucleic acid (Fc-PNA). The PNA was used to minimize the electrostatic repulsion between nucleic acids due to its neutral backbone. The hybridized DNA was examined by differential pulse voltammetry (DPV) and electrochemical impedance spectroscopic (EIS). Compared with the passive hybridization strategy that generally takes several hours, this E-DNA biosensor took the advantage of controllable potential to accelerate nucleic acid hybridization and significantly shorten it to less than 90 s, thereby achieving rapid detection. The electrochemical signal showed a good linear relationship with the target DNA from 10 pM to 10 nM. In addition, this E-DNA biosensor had good specificity that was capable of distinguishing mismatched sequences. The developed biosensor is a promising diagnostic for the rapid and accurate determination of DNA.

Distinct Detection of Ganoderma Boninense On Metal Oxides-Gold Nanoparticle Composite Deposited Interdigitated Electrode DNA sensor

Oil palms suffer severe losses due to Ganoderma boninense infection that causes Basal Stem Rot (BSR). The available detection measuring the severity of BSR disease have not proved satisfactory output. Due to the influence of oil palm industry in country’s economy, effective and efficient means of diagnostic measure is mandatory. Among the available diagnostic tools, biosensors were redeemed to yield the most rapid and selective results. To overcome the current issues, herein Interdigitated Electrode (IDE) electrochemical DNA biosensor to detect Ganoderma boninense was successfully designed and fabricated by thermal deposition. Lift-off photolithography fabrication process was applied followed by the surface chemical functionalization via seed deposition. Zinc Oxide (ZnO) and Titanium Dioxide (TiO2) were overlaid and the functionalized metal oxides IDE surfaces were used to detect DNA sequence complementation from Ganoderma boninense. Furthermore, gold nanoparticles were doped to increase the surface to volume ratio and enhance biocompatibility. Characterizations were made by validating the sensor’s topology characteristics and electrical characteristics. From the results recorded, it has been justified that IDE with ZnO doped with gold nanoparticles surface serves as an excellent DNA sensor for the detection of Ganoderma boninense with a remarkable current of 290 nA and 176 nA for immobilization and hybridization respectively.

A DNA functionalized advanced electrochemical biosensor for identification of the foodborne pathogen Salmonella enterica serovar Typhi in real samples

An efficient platform for the detection of Salmonella enterica serovar Typhi (S. Typhi) is essential for early-stage diagnosis of typhoid to prevent and contain outbreaks. Here, we fabricated an electrochemical DNA biosensor for selective identification of S. Typhi in real samples. The biosensor has been fabricated by immobilizing an amine labelled S. Typhi specific single-strand capture probe on the surface of gold nanoparticles (AuNP) and poly cysteine (P-Cys) modified screen-printed electrode. Differential pulse voltammetry (DPV) of anthraquinone-2-sulfonic acid monohydrate sodium salt (AQMS) as a signal indicator was monitored to detect S. Typhi by hybridization of target DNA with the probe DNA. The fabricated biosensor shows a detection range of 1 × 10−6 to 1 × 10−22 molL−1 with a LOD of 6.8 × 10−25 molL−1 in S. Typhi complementary linear target and 1.8 × 105 to 1.8 CFUml−1 with a LOD of 1 CFUml−1 in a real S. Typhi sample. The biosensor shows excellent discrimination ability to some bases mismatched and different bacterial cultures (same and distant genera). The most beneficial points of the proposed DNA biosensor are the lower limit of detection and the ability to reuse the biosensor more than 6 to 7 times. In addition, the practicability of the biosensor was investigated via detecting S. Typhi in blood, poultry feces, egg, and milk whereby excellent recoveries ranging from 96.54 to 103.47% were demonstrated indicating that this biosensor might be the most promising diagnostic tool for monitoring S. Typhi in clinical and food samples.

Progress of Wearable and Flexible Electrochemical Biosensors With the Aid of Conductive Nanomaterials.

Conductive nanomaterials have recently gained a lot of interest due to their excellent physical, chemical, and electrical properties, as well as their numerous nanoscale morphologies, which enable them to be fabricated into a wide range of modern chemical and biological sensors. This study focuses mainly on current applications based on conductive nanostructured materials. They are the key elements in preparing wearable electrochemical Biosensors, including electrochemical immunosensors and DNA biosensors. Conductive nanomaterials such as carbon (Carbon Nanotubes, Graphene), metals and conductive polymers, which provide a large effective surface area, fast electron transfer rate and high electrical conductivity, are summarized in detail. Conductive polymer nanocomposites in combination with carbon and metal nanoparticles have also been addressed to increase sensor performance. In conclusion, a section on current challenges and opportunities in this growing field is forecasted at the end.

Further insight into the possible influence of electrode blocking agents on the stem-loop based electrochemical DNA sensor parameters

Understanding the electrochemical and interfacial behavior of the biosensor’s recognition layer is crucial for developing devices dedicated to real-life applications. Commonly used binary monolayers with ssDNA as a receptor are also built with 6-mercapto-1-hexanol as an electrode blocking agent (EBA). However, still, poorly reproducible results of such biosensors are reported, especially in the presence of electroactive redox probes freely accessible in measurement buffer. Herein we present investigations on the application of different electrode blocking agents in electrochemical DNA biosensors based on the methylene blue labeled stem-loop-based probe architecture. Electrochemical and working parameters of prepared biosensor were strongly dependent on EBA type used: (i) partial negative charge originated from –OH group in classical mercapto-derivatives of alkanols, 6-mercapto-1-hexanol (MCH) and 2-mercaptoethanol (MCE), (ii) positively charged head group, cysteamine hydrochloride (CHC) (iii) neutral molecule showing two resonance structures, sodium diethyldithiocarbamate, (DEDTC), (iv) differently oriented at the electrode surface aromatic rings, perpendicular to the electrode surface, 2-phenylethanethiol (2-PhET) and significantly tilted, 1,3-benzenedithiol (BDT). Presented investigations show the strong dependence between resonance structures and registered currents. Moreover, because of the improved biosensor’s working parameters and significantly reduced unspecific protein adsorption, DEDTC shows high potential applicability as a new electrode blocking agent during biosensor’s electrochemical sensing layers preparation.

Electrochemical DNA biosensor based on gold nanostars for detection of Escherichia coli O157:H7 from river water

Electrochemical DNA biosensor based on gold nanostars for detection of Escherichia coli O157:H7 from river water

A DNA Electrochemical Sensor via Terminal Protection of Small-Molecule-Linked DNA for Highly Sensitive Protein Detection.

The qualitative and quantitative determination of marker protein is of great significance in the life sciences and in medicine. Here, we developed an electrochemical DNA biosensor for protein detection based on DNA self-assembly and the terminal protecting effects of small-molecule-linked DNA. This strategy is demonstrated using the small molecule biotin and its receptor protein streptavidin (SA). We immobilized DNA with a designed structure and sequence on the surface of the gold electrode, and we named it M1-Biotin DNA. M1-Biotin DNA selectively combines with SA to generate M1-Biotin-SA DNA and protects M1-Biotin DNA from digestion by EXO III; therefore, M1-Biotin DNA remains intact on the electrode surface. M1-Biotin-SA DNA was modified with methylene blue (MB); the MB reporter molecule is located near the surface of the gold electrode, which generates a substantial electrochemical signal during the detection of SA. Through this strategy, we can exploit the presence or absence of an electrochemical signal to provide qualitative target protein determination as well as the strength of the electrochemical signal to quantitatively analyze the target protein concentration. This strategy has been proven to be used for the quantitative analysis of the interaction between biotin and streptavidin (SA). Under optimal conditions, the detection limit of the proposed biosensor is as low as 18.8 pM, and the linear range is from 0.5 nM to 5 μM, showing high sensitivity. The detection ability of this DNA biosensor in complex serum samples has also been studied. At the same time, we detected the folate receptor (FR) to confirm that this strategy can be used to detect other proteins. Therefore, this electrochemical DNA biosensor provides a sensitive, low-cost, and fast target protein detection platform, which may provide a reliable and powerful tool for early disease diagnosis.

Nucleic acid-based electrochemical biosensor: Recent advances in probe immobilization and signal amplification strategies.

With the increasing importance of accurate and early disease diagnosis and the development of personalized medicine, DNA-based electrochemical biosensor has attracted broad scientific and clinical interests in the past decades due to its unique hybridization specificity, fast response time, and potential for miniaturization. In order to achieve high detection sensitivity, the design of DNA electrochemical biosensors depends critically on the improvement of the accessibility of target molecules and the enhancement of signal readout. Here, we summarize the recent advances in DNA probe immobilization and signal amplification strategies with a special focus on DNA nanostructure-supported DNA probe immobilization method, which provides the opportunity to rationally control the distance between probes and keep them in upright confirmation, as well as the contribution of functional nanomaterials in enhancing the signal amplification. The next challenge of biosensors will be the fabrication of point-of-care devices for clinical testing. The advancement of multidisciplinary areas, including nanofabrication, material science, and biochemistry, has exhibited profound promise in achieving such portable sensing devices. This article is categorized under: Diagnostic Tools > Biosensing Diagnostic Tools > Diagnostic Nanodevices Biology-Inspired Nanomaterials > Nucleic Acid-Based Structures.

Potentialities of core@shell nanomaterials for biosensor technologies

This featured article is based on the author's experience and personal overview of the synthesis, characterization, high performances and ultra-responsive utilization of core@shell nanomaterials (CSNs) in the biosensing field. Over the last few years, biosensor-related research using CSNs has progressed significantly, but, recently, CSNs with specific shape and size-tunable dependent physical and chemical properties have been found to be utilized for fabrication of chemosensors and biosensors with substantial attention. Following that, nano and microsensors made from stable, susceptible CSNs are used in diagnostics, enzymatic glucose determination, electrochemical DNA biosensors to detect pathogenic bacteria, and integrated wearable healthcare electronics as sensors. Nowadays, the utilization of plasmonic CSNs as probes for the colorimetric detection of various analytes and antibody conjugated CSNs for naked-eye immunosensing have also received considerable importance. Hence, this featured article compiles the up-to-date data of the state-of-the-art development on CSNs based biosensors and discusses their future aspects and challenges.

Electrochemical DNA Biosensor Based on thiolated-MWCNTs modified glassy carbon electrode for Determination of Protein and Hemoglobin in Urine at Exercise Training Levels

Electrochemical DNA Biosensor Based on thiolated-MWCNTs modified glassy carbon electrode for Determination of Protein and Hemoglobin in Urine at Exercise Training Levels