A biomolecule-free electrochemical sensing approach based on a novel electrode modification technique: Detection of ultra-low concentration of Δ9-tetrahydrocannabinol in saliva by turning a sample analyte into a sensor analyte.

Cannabis is currently one of the most consumed drugs in many countries. Δ9-tetrahydrocannabinol (THC) is the principal psychoactive component of this drug and is present in saliva after consumption. This paper reports a novel biomolecule-free electrochemical approach to detect an ultra-low level of THC in saliva using modified electrodes with molecules of the same analyte (THC) that are detected later via square wave voltammetry. The results from this research revealed that the electrodeposition of THC on the working electrode (sensor analyte) could highly enhance the limit of detection by improving the affinity of the THC molecules present in the sample (sample analyte) to the sensing electrode surface. Detailed descriptions about the optimization of the sensor and its performance in simple media, such as PBS, and complex media, such as simulated and real saliva, are provided. This novel and yet simple electrochemical-based sensing strategy allowed for a low limit of detection of 1.6 ng/mL THC in simulated and real saliva, distinguishing concentrations ranging from 2 to 25 ng/mL, making this technology viable for a real-world application such as roadside testing.

Nanovehicles for Plant Modifications towards Pest- and Disease-Resistance Traits.

In agriculture, plant transformation is a versatile platform for crop improvement with the aim of increased pest resistance and an improved nutrient profile. Nanotechnology can overcome several challenges that face conventional methods of gene delivery. Specifically, nanomaterials offer an optimal platform for biomolecule delivery with unique physiochemical properties as well as the ability to traverse the challenging barrier of the plant cell wall. We review the potential of diverse nanovehicles for biomolecule delivery in plant systems to obtain desired genetic traits. The efficacy of nanoparticles against pests or pathogens is also explored, as well as the interaction of nanovehicles with plant organelles, with due consideration of the effects and toxic profile of nanoparticles.

Noninvasive Label-Free Detection of Cortisol and Lactate Using Graphene Embedded Screen-Printed Electrode.

ABSTRACT: A sensitive and specific immunosensor for the detection of the hormones cortisol and lactate in human or animal biological fluids, such as sweat and saliva, was devised using the label-free electrochemical chronoamperometric technique. By using these fluids instead of blood, the biosensor becomes noninvasive and is less stressful to the end user, who may be a small child or a farm animal. Electroreduced graphene oxide (e-RGO) was used as a synergistic platform for signal amplification and template for bioconjugation for the sensing mechanism on a screen-printed electrode. The cortisol and lactate antibodies were bioconjugated to the e-RGO using covalent carbodiimide chemistry. Label-free electrochemical chronoamperometric detection was used to analyze the response to the desired biomolecules over the wide detection range. A detection limit of 0.1 ng mL-1 for cortisol and 0.1 mM for lactate was established and a correlation between concentration and current was observed. A portable, handheld potentiostat assembled with Bluetooth communication and battery operation enables the developed system for point-of-care applications. A sandwich-like structure containing the sensing mechanisms as a prototype was designed to secure the biosensor to skin and use capillary action to draw sweat or other fluids toward the sensing mechanism. Overall, the immunosensor shows remarkable specificity, sensitivity as well as the noninvasive and point-of-care capabilities and allows the biosensor to be used as a versatile sensing platform in both developed and developing countries.

A novel approach for amine derivatization of MoS2 nanosheets and their application toward label-free immunosensor.

The application of molybdenum disulfide (MoS2) nanosheets has assumed great significance in the design of next-generation biosensors. The immobilization of biomolecules on MoS2 nanosheets has generally been achieved via hydrophobic interactions or through other complicated surface modifications. In this work, we report a novel strategy for electrochemically assisted amine derivatization of MoS2 nanosheets. This newly proposed approach helped to immobilize the MoS2 nanosheets with antibodies via facile EDC/NHS {N-(3-dimethylaminopropyl)-N-(ethylcarbodiimide/N-hydroxysuccinimide)} cross-linking chemistry. The MoS2 nanosheets were first exfoliated and then electrochemically modified with 2-aminobenzylamine. Through a subsequent bioconjugation of the above amine-derivatized MoS2 nanosheets with anti-prostate-specific antigen (PSA) antibodies, an immunosensing device was realized for the detection of the 'prostate specific antigen'. The application of the proposed immunosensor was characterized with a low detection limit (10-3 ng/mL) over a very wide quantitation range (10-3 to 200 ng/mL).

Exploration of two-dimensional bio-functionalized phosphorene nanosheets (black phosphorous) for label free haptoglobin electro-immunosensing applications.

We report on the development of an antibody-functionalized interface based on electrochemically active liquid-exfoliated two-dimensional phosphorene (Ph) nanosheets-also known as black phosphorous nanosheets-for the label-free electrochemical immunosensing of a haptoglobin (Hp) biomarker, a clinical marker of severe inflammation. The electrodeposition has been achieved over the screen-printed electrode (SPE) using liquid-assisted ultrasonically exfoliated black phosphorus nanosheets. Subsequently, Ph-SPEs bioconjugated with Hp antibodies (Ab), using electrostatic interactions via a poly-L-lysine linker for biointerface development. Electrochemical analysis demonstrates that the Ab-modified Ph-SPEs (Ab@Ph-SPE) exhibit enhanced electroconducting behavior as compared to the pristine electrodes. This Ab-functionalized phosphorene-based electrochemical immunosensor platform has demonstrated remarkable sensitivity and specificity, having a dynamic linear response range from 0.01-10 mg ml-1 for Hp in standard and serum samples with a low detection limit (∼0.011 mg ml-1) using the label-free electrochemical technique. The sensor electrodes were also studied with other closely relative interferents to investigate cross reactivity and specificity. This strategy opens up avenues to POC (point-of-care) and on-farm livestock disease monitoring technologies for multiplexed diagnosis in complex biological samples such as serum. The technique is simple in fabrication and provides an analytical response in less than 60 s.

A comprehensive review on nano-molybdenum disulfide/DNA interfaces as emerging biosensing platforms.

The development of nucleic acid-based portable platforms for the real-time analysis of diseases has attracted considerable scientific and commercial interest. Recently, 2D layered molybdenum sulfide (2D MoS2 from here on) nanosheets have shown great potential for the development of next-generation platforms for efficient signal transduction. Through combination with DNA as a biorecognition medium, MoS2 nanostructures have opened new opportunities to design and construct highly sensitive, specific, and commercially viable sensing devices. The use of specific short ssDNA sequences like aptamers has been proven to bind well with the unique transduction properties of 2D MoS2 nanosheets to realize aptasensing devices. Such sensors can be operated on the principles of fluorescence, electro-cheumuluminescence, and electrochemistry with many advantageous features (e.g., robust biointerfacing through various conjugation chemistries, facile sensor assembly, high stability with regard to temperature/pH, and high affinity to target). This review encompasses the state of the art information on various design tactics and working principles of MoS2/DNA sensor technology which is emerging as one of the most sought-after and valuable fields with the advent of nucleic acid inspired devices. To help achieve a new milestone in biosensing applications, great potential of this emerging technique is described further with regard to sensitivity, specificity, operational convenience, and versatility.

A highly efficient 2D exfoliated metal dichalcogenide for the on-farm rapid monitoring of non-esterified fatty acids.

Herein, we report on the development of an electrochemically active liquid exfoliated 2D MoS2 nanosheet based biointerface for the on-farm monitoring of non-esterified fatty acid (NEFA) biomarkers.

Liquid exfoliation of 2D MoS2 nanosheets and their utilization as a label-free electrochemical immunoassay for subclinical ketosis.

We report the step-by-step fabrication of a 2D MoS2 nanostructure-based disposable electrochemical immunosensor to detect ß-hydroxybutyrate (ßHBA), a novel subclinical ketosis biomarker. The MoS2 nanosheets were exfoliated in the liquid phase by ultrasonication, and then followed by deposition on gold colloid modified screen-printed electrodes (Au-SPE). The MoS2-modified electrodes were thoroughly characterized by physical, electrochemical as well as spectroscopic techniques, and the obtained results indicate the successful and irreversible electrodeposition of MoS2 nanosheets. These MoS2-modified disposable electrodes were subsequently bioconjugated with anti-ßHBA antibodies and then employed for the label-free immunosensing of the ßHBA biomarker to detect subclinical ketosis. A simple electrochemical Differential Pulse Voltammetry (DPV) technique based immunodetection was realized for the sensing of varying concentrations of ßHBA antigen. The bioassay demonstrated remarkable sensitivity and specificity having a dynamic linear response range of 0.7 mM to 10 mM for ßHBA in standard antigen solutions, spiked serum, blood, milk and clinical samples, with linear results being obtained with R2 ∼ 0.9923. The sensor electrodes were also studied with other relative interferents to investigate cross reactivity and non-specificity. These electrodes showed a linear, specific, reproducible and stable response towards the ßHBA antigen over a wide range of concentrations.

Recent advancement in biosensors technology for animal and livestock health management.

The term biosensors encompasses devices that have the potential to quantify physiological, immunological and behavioural responses of livestock and multiple animal species. Novel biosensing methodologies offer highly specialised monitoring devices for the specific measurement of individual and multiple parameters covering an animal's physiology as well as monitoring of an animal's environment. These devices are not only highly specific and sensitive for the parameters being analysed, but they are also reliable and easy to use, and can accelerate the monitoring process. Novel biosensors in livestock management provide significant benefits and applications in disease detection and isolation, health monitoring and detection of reproductive cycles, as well as monitoring physiological wellbeing of the animal via analysis of the animal's environment. With the development of integrated systems and the Internet of Things, the continuously monitoring devices are expected to become affordable. The data generated from integrated livestock monitoring is anticipated to assist farmers and the agricultural industry to improve animal productivity in the future. The data is expected to reduce the impact of the livestock industry on the environment, while at the same time driving the new wave towards the improvements of viable farming techniques. This review focusses on the emerging technological advancements in monitoring of livestock health for detailed, precise information on productivity, as well as physiology and well-being. Biosensors will contribute to the 4th revolution in agriculture by incorporating innovative technologies into cost-effective diagnostic methods that can mitigate the potentially catastrophic effects of infectious outbreaks in farmed animals.

Graphene quantum dot modified screen printed immunosensor for the determination of parathion.

The widespread use of pesticides has immense effect on increased crop productions. However, they are also responsible for posing detrimental health hazards and/or for contaminating the environment with chemical residues. A routine and an on-field detection of pesticide residues in different food, water, and soil samples has become a need of the hour for which biosensors can offer a viable alternative. The present work reports a functionalized graphene quantum dot (GQD) based screen printed electrochemical immunosensor for the detection of parathion. The application of GQDs has permitted the realization of a sensitive, robust, and reproducible sensor unlike those carried out earlier for the similar purposes. This immunosensor exhibited a dynamic linear response for parathion within the range of 0.01-106 ng/L with a very low detection limit of 46 pg/L. According to the analysis of potential interferences, the proposed sensor was specifically detecting parathion even in the presence of its metabolite, paraoxon. The investigations of the proposed sensing approach with respect to stability, response reproducibility, and regeneration have fully supported its potential practical applicability.

Graphene-based multiplexed disposable electrochemical biosensor for rapid on-farm monitoring of NEFA and ßHBA dairy biomarkers.

Elevated circulating concentrations of non-esterified fatty acids (NEFA) and beta hydroxy butyrate (ßHBA) in biological fluids are recognized as critical biomarkers for early diagnosis of negative energy balance (NEB) in dairy cows. Herein, we report the development of a cost-effective, bio-friendly and electrochemically active dual screen printed electrode (SPE) sensor platform composed of electro-reduced graphene oxide nanosheets (E-rGO) modified with specific antibodies against NEFA and ßHBA. The chemically synthesized graphene oxide (GO) was reduced directly on the screen printed electrode (SPE) surface via a green electrochemical approach without using toxic chemicals. The E-rGO was characterized using various analytical techniques, like XPS, SEM, TEM, AFM, UV-Vis, Raman spectroscopy, FTIR, and XRD, to get insight into its properties. Electrochemical analysis demonstrates that the E-rGO-modified SPE electrodes exhibit enhanced and durable redox properties as compared to the pristine graphite and GO electrodes. Target specificity is accomplished through immobilization of specific antibodies against NEFA and ßHBA over the nanostructure-modified surface of the SPE, which only interacts with its counterpart NEFA and ßHBA only. The antibodies retain their characteristic immuno-complex formation property upon immobilization and exhibit changes to amperometric signals upon interaction with various concentrations of NEFA and ßHBA in standard, spiked blood and real clinical samples. The DPV signals resulted from the developed immunosensor platform exhibited a good correlation (R2∼ 0.99 for both NEFA and ßHBA) for a wide range of target concentrations from 0.1 mM to 10 mM. The proposed immunosensor design not only provides a rapid analytical response time (≥1 min), but also simplicity in fabrication and instrumentation, which may provide a promising approach for on-farm diagnostics of ketosis and metabolic disorders associated with NEB.

A label-free electrochemical immunosensor for the detection of cardiac marker using graphene quantum dots (GQDs).

A label-free immunosensor based on electrochemical impedance spectroscopy has been developed for the sensitive detection of a cardiac biomarker myoglobin (cMyo). Hydrothermally synthesized graphene quantum dots (GQDs) have been used as an immobilized template on screen printed electrodes for the construction of an impedimetric sensor platform. The GQDs-modified electrode was conjugated with highly specific anti-myoglobin antibodies to develop the desired immunosensor. The values of charge transfer resistance (Rct) were monitored as a function of varying antigen concentration. The Rct value of the immunosensor showed a linear increase (from 0.20 to 0.31kΩ) in the range of 0.01-100ng/mL cMyo. The specific detection of cMyo was also made in the presence of other competing proteins. The limit of detection for the proposed immunosensor was estimated as 0.01ng/mL which is comparable to the standard ELISA techniques.

A New Electrolytic Synthesis Method for Few-Layered MoS2 Nanosheets and Their Robust Biointerfacing with Reduced Antibodies.

We report an efficient method for the synthesis of few-layered MoS2 nanosheets and demonstrate their application in the label-free detection of the prostate-specific antigen (PSA) cancer marker. As a novel strategy, the electro-dissolution of molybdenum metal sheets in the presence of Na(+) and S(2-) ions led to the formation of Na(+) intercalated MoS2. Further exfoliation by ultrasonication yielded the desired formation of few-layered MoS2 nanosheets. After comprehensive characterization, the synthesized MoS2 nanosheets were channeled in a field-effect transistor (FET) microdevice. Chemically reduced anti-PSA antibodies were immobilized on the MoS2 channel above the FET microdevice to construct a specific PSA immunosensor. The antibodies were deliberately reduced to expose the hinge-region disulfide bonds. This approach offered a robust and site-directed immunosensing device through biointerfacing of the sulfhydryl groups (-SH) in the reduced antibody with the surface S atoms of MoS2. This device was validated as an effective immunosensor with a low detection limit (10(-5) ng/mL) over a wide linear detection range (10(-5) to 75 ng/mL).

Graphene modified screen printed immunosensor for highly sensitive detection of parathion.

Due to indiscriminate use of pesticides, there is a growing need to develop sensors that can sensitively detect the trace amount of pesticides in food and water samples. Parathion, identified as an acetylcholinesterase inhibitor, had been one of the most widely used pesticides throughout the world. Symptoms of its poisoning are found to be diverse enough to include nausea, vomiting, diarrhea, muscle cramping/twitching, and shortness of breath. In this work, a graphene based impedimetric immunosensor has been fabricated and employed for highly sensitive and specific detection of parathion. The fabrication proceeded through the modification of the screen-printed carbon electrodes (SPE) with graphene sheets, followed by their functionalization with 2-aminobenzyl amine (2-ABA) via an electrochemical reaction. These amine functionalized graphene electrodes were then bio-interfaced with the anti-parathion antibodies. In the impedimetric mode, this biosensor detected parathion in a broad linear range, i.e. 0.1-1000ng/L with a very low limit of detection (52pg/L). It also showed high selectivity towards parathion in the presence of malathion, paraoxon, and fenitrothion. The viability of this biosensor was demonstrated by detecting parathion in real samples (e.g., tomato and carrot) and through cross-calibration against HPLC.

One step in-situ synthesis of amine functionalized graphene for immunosensing of cardiac marker cTnI.

2-Aminobenzyl amine (2-ABA) functionalized graphene is proposed for the ultrasensitive immunosensing of Cardiac Troponin I (cTnI). 2-ABA was electrochemically polymerized on the graphene decorated interdigitated electrode to obtain the amine functionalized graphene (f-GN). The f-GN electrode was then modified with monoclonal anti-cTnI antibodies via Schiff reaction based chemistry. Detailed characteristics of the processes involved and the finally developed antibody conjugated f-GN interdigitated electrode have been studied. The above micro-device was used in a drain source configuration for the sensing of cTnI. A wide dynamic linear range of antigen detection (0.01-1ng/mL) is achieved with the limit of detection of 0.01ng/mL. The utility of the proposed sensing technique is demonstrated by successfully testing the antigen concentration in spiked serum samples.

Biofunctionalized rebar graphene (f-RG) for label-free detection of cardiac marker troponin I.

One-step microwave-assisted unscrolling of carbon nanotubes to form functionalized rebar graphene (f-RG) is reported. The well-characterized f-RG on an interdigitated electrode biochip in a FET configuration showed enhanced electronic properties, as demonstrated with I-V characteristics. The developed device was biofunctionalized with specific anti-cTnI antibodies exhibiting a shift of threshold voltage from -2.15 V to -0.5 V and decrease in electron mobility from 3.609 × 10(4) to 8.877 × 10(3) cm(2) V(-1) s(-1). The new sensing strategy holds great promise for its applicability in diagnostics exhibiting high sensitivity (∼ 1 pg/mL) and specificity toward cardiac marker (cTnI).

Phosphinic acid functionalized carbon nanotubes for sensitive and selective sensing of chromium(VI).

Single-walled carbon nanotubes (SWCNTs) have been functionalized with a phosphinic acid derivative 'bis(2,4,4-trimethylpentyl) phosphinic acid' (PA/d). It has been achieved by treating the chlorinated SWCNTs with PA/d at 80°C. Successful functionalization and different nanomaterial properties have been investigated by UV-vis-NIR, FTIR, Raman spectroscopy, AFM and FE-SEM. PA/d conjugated SWCNTs (CNT-PA) are dispersible in some common organic solvents, e.g. CH2Cl2, DMF, CHCl3, and THF. The 'CNT-PA' complex was spin-casted on boron doped silicon wafer. Thus fabricated sensing electrode is demonstrated for sensitive and selective electrochemical sensing of chromium(VI) ions. A linear response is obtained over a wide range of Cr(VI) concentration (0.01-10 ppb). The sensor's sensitivity and the limit of detection are observed to be 35 ± 4 nA/ppb and 0.01 ppb, respectively. The practical utility of the proposed sensor is demonstrated by determining the Cr(VI) concentration in an industrial effluent sample and an underground water sample.

Graphene-gated biochip for the detection of cardiac marker Troponin I.

We report lithium ion intercalation mediated efficient exfoliation of graphite to form monolithic graphene sheets which have subsequently been investigated for the development of highly sensitive label-free electrochemical detection platform for cardiac biomarker, Troponin I (cTnI). The spectroscopic and morphological analysis demonstrated the formation of defect free graphene sheets which were successfully employed to fabricate an inter-digited microdevice in a drain-source configuration on a silicon biochip. The graphene gated biochip functionalized with anti-cTnI antibodies used in label free detection of cTnI which exhibited an excellent sensitivity in the picogram range (~1 pg mL(-1)) for cTnI without the use of any enzymatic amplification that promises its potential applicability for bio-molecular detection in clinical diagnosis.

Bio-functionalized graphene-graphene oxide nanocomposite based electrochemical immunosensing.

We report a novel in-situ electrochemical synthesis approach for the formation of functionalized graphene-graphene oxide (fG-GO) nanocomposite on screen-printed electrodes (SPE). Electrochemically controlled nanocomposite film formation was studied by transmission electron microscopy (TEM) and Raman spectroscopy. Further insight into the nanocomposite has been accomplished by the Fourier transformed infrared spectroscopy (FTIR), thermal gravimetric analysis (TGA) and X-ray diffraction (XRD) spectroscopy. Configured as a highly responsive screen-printed immunosensor, the fG-GO nanocomposite on SPE exhibits electrical and chemical synergies of the nano-hybrid functional construct by combining good electronic properties of functionalized graphene (fG) and the facile chemical functionality of graphene oxide (GO) for compatible bio-interface development using specific anti-diuron antibody. The enhanced electrical properties of nanocomposite biofilm demonstrated a significant increase in electrochemical signal response in a competitive inhibition immunoassay format for diuron detection, promising its potential applicability for ultra-sensitive detection of range of target analytes.