This study reports on the development of a novel impedimetric immunosensor design using plant-derived antigenic glycoprotein for the detection of dengue virus (DENV) IgG antibodies. The electrochemical immunosensor platform was constructed using screen-printed carbon electrode (SPCE) modified with graphene/titanium dioxide (G/TiO2) nanocomposite to improve the electrode in terms electrochemical performance and specific surface area. A plant-derived dengue envelope domain III (EDIII) protein was used as the antigenic probe protein in this immunosensing strategy. Under optimised sensing conditions, the immunosensor demonstrated high sensitivity towards DENV IgG in a wide linear working range (62.5-2000 ng/mL), with a limit of detection of 2.81 ng/mL. The immunosensor showed high specificity for discriminating DENV IgG against antibodies of other infectious disease, including the closely related Zika virus (ZIKV). The reliability of the immunosensor in serological diagnosis was verified by challenging the immunosensor against serum samples, compared to conventional enzyme-linked immunosorbent assay (ELISA). As shown by its remarkable performance throughout the study, the devised immunosensor is proposed as a reliable and practical diagnostic tool for the serological detection of dengue in realistic applications.
The envelope glycoprotein domain III (EDIII) of dengue virus (DENV) has been recognised as the antigenic region responsible for receptor binding. In the present work, we have proposed a novel immunosensor constructed on a graphene-coated screen-printed carbon electrode (SPCE) using plant-derived EDIII as the probe antigen to target DENV IgG antibodies. The developed immunosensor demonstrated high sensitivity towards DENV IgG within a wide linear working range (125-2000 ng mL-1) under the optimised sensing conditions. The limit of detection was determined to be 22.5 ng mL-1. The immunosensor also showed high specificity towards DENV IgG, capable of differentiating DENV IgG from the antibodies of other infectious diseases including the similarly structured Zika virus (ZIKV). The ability of the immunosensor to detect dengue antibodies in serum samples was also verified by conducting tests on mouse serum samples. The proposed immunosensor was able to provide a binary (positive/negative) response towards the serum samples comparable to the conventional enzyme-linked immunosorbent assay (ELISA), indicating promising potential for realistic applications.
An efficient electrochemical impedance genosensing platform has been constructed based on graphene/zinc oxide nanocomposite produced via a facile and green approach. Highly pristine graphene was synthesised from graphite through liquid phase sonication and then mixed with zinc acetate hexahydrate for the synthesis of graphene/zinc oxide nanocomposite by solvothermal growth. The as-synthesised graphene/zinc oxide nanocomposite was characterised with scanning electron microscopy (SEM), transmission electron microscopy (TEM), Raman spectroscopy and X-ray diffractometry (XRD) to evaluate its morphology, crystallinity, composition and purity. An amino-modified single stranded DNA oligonucleotide probe synthesised based on complementary Coconut Cadang-Cadang Viroid (CCCVd) RNA sequence, was covalently bonded onto the surface of graphene/zinc oxide nanocomposite by the bio-linker 1-pyrenebutyric acid N-hydroxysuccinimide ester. The hybridisation events were monitored by electrochemical impedance spectroscopy (EIS). Under optimised sensing conditions, the single stranded CCCVd RNA oligonucleotide target could be quantified in a wide range of 1.0×10-11M to 1.0×10-6 with good linearity (R =0.9927), high sensitivity with low detection limit of 4.3×10-12M. Differential pulse voltammetry (DPV) was also performed for the estimation of nucleic acid density on the graphene/zinc oxide nanocomposite-modified sensing platform. The current work demonstrates an important advancement towards the development of a sensitive detection assay for various diseases involving RNA agents such as CCCVd in the future.
In recent years, carbon nanotubes have received widespread attention as promising carbon-based nanoelectronic devices. Due to their exceptional physical, chemical, and electrical properties, namely a high surface-to-volume ratio, their enhanced electron transfer properties, and their high thermal conductivity, carbon nanotubes can be used effectively as electrochemical sensors. The integration of carbon nanotubes with a functional group provides a good and solid support for the immobilization of enzymes. The determination of glucose levels using biosensors, particularly in the medical diagnostics and food industries, is gaining mass appeal. Glucose biosensors detect the glucose molecule by catalyzing glucose to gluconic acid and hydrogen peroxide in the presence of oxygen. This action provides high accuracy and a quick detection rate. In this paper, a single-wall carbon nanotube field-effect transistor biosensor for glucose detection is analytically modeled. In the proposed model, the glucose concentration is presented as a function of gate voltage. Subsequently, the proposed model is compared with existing experimental data. A good consensus between the model and the experimental data is reported. The simulated data demonstrate that the analytical model can be employed with an electrochemical glucose sensor to predict the behavior of the sensing mechanism in biosensors.
A novel disposable electrochemical biosensor based on immobilized calf thymus double-stranded DNA (dsDNA) on the carbon-based screen-printed electrode (SPE) is developed for rapid biorecognition of carrageenan by using methylene blue (MB) redox indicator. The biosensor protocol for the detection of carrageenan is based on the concept of competitive binding of positively charged MB to the negatively charged dsDNA and carrageenan. The decrement in the MB cathodic peak current (ipc) signal as a result of the released MB from the immobilized dsDNA, and attracted to the carrageenan can be monitored via differential pulse voltammetry (DPV). The biosensor showed high sensitivity and selectivity to carrageenan at low concentration without interference from other polyanions such as alginate, gum arabic and starch. Calibration of the biosensor with carrageenan exhibited an excellent linear dependence from 1-10 mg L-1 (R2 = 0.98) with a detection limit of 0.08 mg L-1. The DNA-based carrageenan biosensor showed satisfactory reproducibility with 5.6-6.9% (n = 3) relative standard deviations (RSD), and possessing several advantages such as simplicity, fast and direct application to real sample analysis without any prior extensive sample treatments, particularly for seaweeds and food analyses.
A potentiometric whole cell biosensor based on immobilized marine bacterium, Pseudomonas carrageenovora producing κ-carrageenase and glycosulfatase enzymes for specific and direct determination of κ-carrageenan, is described. The bacterial cells were immobilized on the self-plasticized hydrogen ion (H+)-selective acrylic membrane electrode surface to form a catalytic layer. Hydrogen ionophore I was incorporated in the poly(n-butyl acrylate) [poly(nBA)] as a pH ionophore. Catalytic decomposition of κ-carrageenan by the bienzymatic cascade reaction produced neoagarobiose, an inorganic sulfate ion and a proton. The latter was detectable by H+ ion transducer for indirect potentiometric quantification of κ-carrageenan concentration. The use of a disposable screen-printed Ag/AgCl electrode (SPE) provided no cleaning requirement and enabled κ-carrageenan detection to be carried out conveniently without cross contamination in a complex food sample. The SPE-based microbial biosensor response was found to be reproducible with high reproducibility and relative standard deviation (RSD) at 2.6% (n = 3). The whole cell biosensor demonstrated a broad dynamic linear response range to κ-carrageenan from 0.2-100 ppm in 20 mM phosphate buffer saline (PBS) at pH 7.5 with a detection limit at 0.05 ppm and a Nernstian sensitivity of 58.78±0.87 mV/decade (R2 = 0.995). The biosensor showed excellent selectivity towards κ-carrageenan compared to other types of carrageenans tested e.g. ι-carrageenan and λ-carrageenan. No pretreatment to the food sample was necessary when the developed whole cell biosensor was employed for direct assay of κ-carrageenan in dairy product.
A new sensing area for a sensor based on surface plasmon resonance (SPR) was fabricated to detect trace amounts of mercury and lead ions. The gold surface used for SPR measurements were modified with polypyrrole-chitosan (PPy-CHI) conducting polymer composite. The polymer layer was deposited on the gold surface by electrodeposition. This optical sensor was used for monitoring toxic metal ions with and without sensitivity enhancement by chitosan in water samples. The higher amounts of resonance angle unit (ΔRU) were obtained for PPy-CHI film due to a specific binding of chitosan with Pb(2+) and Hg(2+) ions. The Pb(2+) ion bind to the polymer films most strongly, and the sensor was more sensitive to Pb(2+) compared to Hg(2+). The concentrations of ions in the parts per million range produced the changes in the SPR angle minimum in the region of 0.03 to 0.07. Data analysis was done by Matlab software using Fresnel formula for multilayer system.
Due to the low titer or uneven distribution of Citrus tristeza virus (CTV) in field samples, detection of CTV by using conventional detection techniques may be difficult. Therefore, in the present work, the cadmium-telluride quantum dots (QDs) was conjugated with a specific antibody against coat protein (CP) of CTV, and the CP were immobilized on the surface of gold nanoparticles (AuNPs) to develop a specific and sensitive fluorescence resonance energy transfer (FRET)-based nanobiosensor for detecting CTV. The maximum FRET efficiency for the developed nano-biosensor was observed at 60% in AuNPs-CP/QDs-Ab ratio of 1:8.5. The designed system showed higher sensitivity and specificity over enzyme linked immunosorbent assay (ELISA) with a limit of detection of 0.13μgmL(-1) and 93% and 94% sensitivity and specificity, respectively. As designed sensor is rapid, sensitive, specific and efficient in detecting CTV, this could be envisioned for diagnostic applications, surveillance and plant certification program.
A DNA micro-optode for dengue virus detection was developed based on the sandwich hybridization strategy of DNAs on succinimide-functionalized poly(n-butyl acrylate) (poly(nBA-NAS)) microspheres. Gold nanoparticles (AuNPs) with an average diameter of ~20 nm were synthesized using a centrifugation-based method and adsorbed on the submicrometer-sized polyelectrolyte-coated poly(styrene-co-acrylic acid) (PSA) latex particles via an electrostatic method. The AuNP-latex spheres were attached to the thiolated reporter probe (rDNA) by Au-thiol binding to functionalize as an optical gold-latex-rDNA label. The one-step sandwich hybridization recognition involved a pair of a DNA probe, i.e., capture probe (pDNA), and AuNP-PSA reporter label that flanked the target DNA (complementary DNA (cDNA)). The concentration of dengue virus cDNA was optically transduced by immobilized AuNP-PSA-rDNA conjugates as the DNA micro-optode exhibited a violet hue upon the DNA sandwich hybridization reaction, which could be monitored by a fiber-optic reflectance spectrophotometer at 637 nm. The optical genosensor showed a linear reflectance response over a wide cDNA concentration range from 1.0 × 10-21 M to 1.0 × 10-12 M cDNA (R2 = 0.9807) with a limit of detection (LOD) of 1 × 10-29 M. The DNA biosensor was reusable for three consecutive applications after regeneration with mild sodium hydroxide. The sandwich-type optical biosensor was well validated with a molecular reverse transcription polymerase chain reaction (RT-PCR) technique for screening of dengue virus in clinical samples, e.g., serum, urine, and saliva from dengue virus-infected patients under informed consent.
The bovine milk allergenic protein, 'β-lactoglobulin' is one of the leading causes of milk allergic reaction. In this research, a novel label-free non-faradaic capacitive aptasensor was designed to detect β-lactoglobulin using a Laser Scribed Graphene (LSG) electrode. The graphene was directly engraved into a microgapped (~ 95 µm) capacitor-electrode pattern on a flexible polyimide (PI) film via a simple one-step CO2 laser irradiation. The novel hybrid nanoflower (NF) was synthesized using 1,1'-carbonyldiimidazole (CDI) as the organic molecule and copper (Cu) as the inorganic molecule via one-pot biomineralization by tuning the reaction time and concentration. NF was fixed on the pre-modified PI film at the triangular junction of the LSG microgap specifically for bio-capturing β-lactoglobulin. The fine-tuned CDI-Cu NF revealed the flower-like structures was viewed through field emission scanning electron microscopy. Fourier-transform infrared spectroscopy showed the interactions with PI film, CDI-Cu NF, oligoaptamer and β-lactoglobulin. The non-faradaic sensing of milk allergen β-lactoglobulin corresponds to a higher loading of oligoaptamer on 3D-structured CDI-Cu NF, with a linear range detection from 1 ag/ml to 100 fg/ml and attomolar (1 ag/ml) detection limit (S/N = 3:1). This novel CDI-Cu NF/LSG microgap aptasensor has a great potential for the detection of milk allergen with high-specificity and sensitivity.
Artikel ini menganalisis biosensor resonans plasmon permukaan (SPR) dengan lapisan grafin yang meningkatkan
kecekapan biosensor urea kerana penerapannya yang tinggi. Tatasusunan Kretschmann merupakan teknik yang paling
berkesan digunakan untuk pengujaan plasmon. Dalam kajian ini, kami menganalisis kesan ekalapisan MoS2 dengan
lapisan grafin yang didepositkan pada bahan plasmon, iaitu logam emas (Au), di dalam tatasusunan ini. Simulasi untuk
menganalisis tatasusunan ini adalah berdasarkan kepada kaedah perbezaan terhingga domain masa (FDTD). Prestasi
biosensor SPR dapat dipantau dengan menganalisis kepekaan dan lebar penuh pada separuh maksimum (FWHM) spektrum
SPR. Pengukuran diperhatikan pada panjang gelombang 670 nm dan 785 nm untuk pengesanan urea. Indeks molar dan
indeks biasan berbeza (RI) daripada 1.335 sehingga 1.342 untuk lapisan penderiaan. Keputusan menunjukkan peratus
peningkatan kepekaan biosensor Au/MoS2
/grafin berbanding biosensor Au konvensional adalah 98% dan 202% masingmasing
pada panjang gelombang 670 nm dan 785 nm. Ini menunjukkan bahawa cadangan biosensor SPR yang novel ini
adalah lebih sensitif untuk pengesanan urea.
Whole cell biosensors always face the challenge of low stability of biological components and short storage life. This paper reports the effects of poly(2-hydroxyethyl methacrylate) (pHEMA) immobilization on a whole cell fluorescence biosensor for the detection of heavy metals (Cu, Pb, Cd), and pesticides (dichlorophenoxyacetic acid (2,4-D), and chlorpyrifos). The biosensor was produced by entrapping the cyanobacterium Anabaena torulosa on a cellulose membrane, followed by applying a layer of pHEMA, and attaching it to a well. The well was then fixed to an optical probe which was connected to a fluorescence spectrophotometer and an electronic reader. The optimization of the biosensor using several factors such as amount of HEMA and drying temperature were undertaken. The detection limits of biosensor without pHEMA for Cu, Cd, Pb, 2,4-D and chlorpyrifos were 1.195, 0.027, 0.0100, 0.025 and 0.025 µg/L respectively. The presence of pHEMA increased the limits of detection to 1.410, 0.250, 0.500, 0.235 and 0.117 µg/L respectively. pHEMA is known to enhance the reproducibility of the biosensor with average relative standard deviation (RSD) of ±1.76% for all the pollutants tested, 48% better than the biosensor without pHEMA (RSD = ±3.73%). In storability test with Cu 5 µg/L, the biosensor with pHEMA performed 11.5% better than the test without pHEMA on day-10 and 5.2% better on day-25. pHEMA is therefore a good candidate to be used in whole cell biosensors as it increases reproducibility and enhances biosensor storability.
Vibrio cholerae is a Gram-negative bacterium that causes cholera, a diarrheal disease. Cholera is widespread in poor, under-developed or disaster-hit countries that have poor water sanitation. Hence, a rapid detection method for V. cholerae in the field under these resource-limited settings is required. In this paper, we describe the development of an electrochemical genosensor assay using lyophilized gold nanoparticles/latex microsphere (AuNPs-PSA) reporter label. The reporter label mixture was prepared by lyophilization of AuNPs-PSA-avidin conjugate with different types of stabilizers. The best stabilizer was 5% sorbitol, which was able to preserve the dried conjugate for up to 30 days. Three methods of DNA hybridization were compared and the one-step sandwich hybridization method was chosen as it was fastest and highly specific. The performance of the assay using the lyophilized reagents was comparable to the wet form for detection of 1aM to 1fM of linear target DNA. The assay was highly specific for V. cholerae, with a detection limit of 1fM of PCR products. The ability of the sensor is to detect LAMP products as low as 50ngµl(-1). The novel lyophilized AuNPs-PSA-avidin reporter label with electrochemical genosensor detection could facilitate the rapid on-site detection of V. cholerae.
Graphene (GR) and its derivatives are promising materials on the horizon of nanotechnology and material science and have attracted a tremendous amount of research interest in recent years. The unique atom-thick 2D structure with sp(2) hybridization and large specific surface area, high thermal conductivity, superior electron mobility, and chemical stability have made GR and its derivatives extremely attractive components for composite materials for solar energy conversion, energy storage, environmental purification, and biosensor applications. This review gives a brief introduction of GR's unique structure, band structure engineering, physical and chemical properties, and recent energy-related progress of GR-based materials in the fields of energy conversion (e.g., photocatalysis, photoelectrochemical water splitting, CO2 reduction, dye-sensitized and organic solar cells, and photosensitizers in photovoltaic devices) and energy storage (batteries, fuel cells, and supercapacitors). The vast coverage of advancements in environmental applications of GR-based materials for photocatalytic degradation of organic pollutants, gas sensing, and removal of heavy-metal ions is presented. Additionally, the use of graphene composites in the biosensing field is discussed. We conclude the review with remarks on the challenges, prospects, and further development of GR-based materials in the exciting fields of energy, environment, and bioscience.
The fabrication of an electrochemical sensor based on an iron oxide/graphene modified glassy carbon electrode (Fe3O4/rGO/GCE) and its simultaneous detection of dopamine (DA) and ascorbic acid (AA) is described here. The Fe3O4/rGO nanocomposite was synthesized via a simple, one step in-situ wet chemical method and characterized by different techniques. The presence of Fe3O4 nanoparticles on the surface of rGO sheets was confirmed by FESEM and TEM images. The electrochemical behavior of Fe3O4/rGO/GCE towards electrocatalytic oxidation of DA was investigated by cyclic voltammetry (CV) and differential pulse voltammetry (DPV) analysis. The electrochemical studies revealed that the Fe3O4/rGO/GCE dramatically increased the current response against the DA, due to the synergistic effect emerged between Fe3O4 and rGO. This implies that Fe3O4/rGO/GCE could exhibit excellent electrocatalytic activity and remarkable electron transfer kinetics towards the oxidation of DA. Moreover, the modified sensor electrode portrayed sensitivity and selectivity for simultaneous determination of AA and DA. The observed DPVs response linearly depends on AA and DA concentration in the range of 1-9 mM and 0.5-100 µM, with correlation coefficients of 0.995 and 0.996, respectively. The detection limit of (S/N = 3) was found to be 0.42 and 0.12 µM for AA and DA, respectively.
Clenbuterol (CLB) is an antibiotic and illegal growth promoter drug that has a long half-life and easily remains as residue and contaminates the animal-based food product that leads to various health problems. In this work, electrochemical immunosensor based on poly(3,4-ethylenedioxythiophene)/graphene oxide (PEDOT/GO) modified screen-printed carbon electrode (SPCE) for CLB detection was developed for antibiotic monitoring in a food product. The modification of SPCE with PEDOT/GO as a sensor platform was performed through electropolymerization, while the electrochemical assay was accomplished while using direct competitive format in which the free CLB and clenbuterol-horseradish peroxidase (CLB-HRP) in the solution will compete to form binding with the polyclonal anti-clenbuterol antibody (Ab) immobilized onto the modified electrode surface. A linear standard CLB calibration curve with R² = 0.9619 and low limit of detection (0.196 ng mL-1) was reported. Analysis of milk samples indicated that this immunosensor was able to detect CLB in real samples and the results that were obtained were comparable with enzyme-linked immunosorbent assays (ELISA).
Application of surface plasmon resonance (SPR) biosensor in detection of genetically modified organism (GMO) is demonstrated. A total of four biotinylated probes namely Tnosb, P35Sb, LECb and TSQb were successfully immobilized onto the SA chip. Results analysis indicated that the SPR system with the sensor chip immobilized with the Tnosb, P35Sb, LECb and TSQb biotinylated probes potentially detect complementary standard fragments as low as 1 nM. Biospecific interaction analysis (BIA), employing surface plasmon resonance (SPR) and biosensor technologies provide easy, rapid and automatable approach in detection of GMOs. Short assay times, label free DNA hybridization reaction and no toxic compounds are required, i.e. ethidium bromide, and the reusability of the sensor surface are some of the factors that contribute to the general advantages of the surface plasmon resonance (SPR) biosensor system in detection of GMOs.
Detection of host integrated viral oncogenes are critical for early and point-of-care molecular diagnostics of virus-induced carcinoma. However, available diagnostic approaches are incapable of combining both cost-efficient medical diagnosis and high analytical performances. To circumvent this, we have developed an improved IDE-based nanobiosensor for biorecognition of HPV-16 infected cervical cancer cells through electrochemical impedance spectroscopy. The system is fabricated by coating gold (Au) doped zinc oxide (ZnO) nanorods interfaced with HPV-16 viral DNA bioreceptors on top of the Interdigitated Electrode (IDE) chips surface. Due to the concurrently improved sensitivity and biocompatibility of the designed nanohybrid film, Au decorated ZnO-Nanorod biosensors demonstrate exceptional detection of HPV-16 E6 oncogene, the cancer biomarker for HPV infected cervical cancers. This sensor displayed high levels of sensitivity by detecting as low as 1fM of viral E6 gene target. The sensor also exhibited a stable functional life span of more than 5 weeks, good reproducibility and high discriminatory properties against HPV-16. Sensor current responses are obtained from cultured cervical cancer cells which are close to clinical cancer samples. Hence, the developed sensor is an adaptable tool with high potential for clinical diagnosis especially useful for economically challenged countries/regions.
A label -free DNAzyme amplified biosensor is found to be highly selective and sensitive towards fluorescent detection of Pb2+ ions in aqueous media. The DNAzyme complex has designed by the hybridization of the enzyme and substrate strand. In the presence of Pb2+, the DNAzyme activated and cleaved the substrate strand of RNA site (rA) into two oligonucleotide fragments. Further, the free fragment was hybridized with a complementary strand on the surface of MBs. After magnetic separation, SYBER Green I was added and readily intercalate with the dsDNA to gives a bright fluorescence signal with intensity directly proportional to the concentration of Pb2+ions. A detection limit of 5 nM in Pb2+ the detection range 0 to 500 nM was obtained. This label- free fluorescent biosensor has been successfully applied to the determination of environmental water samples. Then results open up the possibility for real-time quantitative detection of Pb2+ with convenient potential applications in the biological and environmental field. Graphical Abstract.
The development of an optical biosensor based on immobilization of 3-methyl-2-benzothiazolinone hydrazone (MBTH) in hybrid nafion/sol-gel silicate film and tyrosinase in chitosan film for the detection of phenolic compounds has been described. Tyrosinase was immobilized in chitosan film deposited on the hybrid nafion/sol-gel silicate film containing MBTH. The enzymatic oxidation product of phenolic compounds were stabilized through formation of adduct with MBTH to produce a maroon color adduct. The color intensity of adduct was found to increase proportionally with the increase of the substrate concentrations after 5min exposure. The linearity of the biosensor towards phenol, catechol and m-cresol were in the respective concentration range of 0.5-7.0, 0.5-10.0 and 1.0-13.0mg/L with detection limit of 0.18, 0.23 and 0.43mg/L, respectively. The biosensor shows a good stability for at least 3 months.