Template Free Anisotropically Grown Gold Nanocluster Based Electrochemical Immunosensor for Ultralow Detection of Cardiac Troponin I.

Anisotropic gold nanostructures have fascinated with their exceptional electronic properties, henceforth exploited for the fabrication of electrochemical sensors. However, their synthesis approaches are tedious and often require a growth template. Modern lifestyle has caused an upsurge in the risk of heart attack and requires urgent medical attention. Cardiac troponin I can serve as a biomarker in identification of suspected myocardial infection (heart attack). Hence the present work demonstrates the fabrication of a sensing platform developed by assimilating anisotropic gold nanoclusters (AuNCs) with anti cTnI antibody (acTnI) for the detection of cardiac troponin I (cTnI). The uniqueness and ease of synthesis by a template-free approach provides an extra edge for the fabrication of AuNC coated electrodes. The template-free growth of anisotropic AuNCs onto the indium tin oxide (ITO) glass substrates offers high sensitivity (2.2 × 10-4 A ng-1 mL cm-2) to the developed sensor. The immunosensor was validated by spiking different concentrations of cTnI in artificial serum with negligible interference under optimized conditions. The sensor shows a wide range of detection from 0.06-100 ng/mL with an ultralow detection limit. Thus, it suggests that the template-free immunosensor can potentially be used to screen the traces of cTnI present in blood serum samples, and the AuNCs based platform holds great promise as a transduction matrix, hence it can be exploited for broader sensing applications.

Rapid sensing ofTilletia indica - Teliospore in wheat extractby apiezoelectric label free immunosensor.

'Tilletia indica', a fungal pathogen causes Karnal bunt disease in wheat. It has been renowned as a quarantine pest in more than 50 countries, therefore, urged a threat to wheat in the international market. To date, conventional methods employed to detect the disease involve the tentative identification of spores (teliospores) based on morphology. For effective and specific disease control, it is essential to get the specific protein of the analyte (teliospore) to target. In present study, a label-free immunosensor has been developed to detect Karnal bunt disease. A specifically synthesized anti-teliosporic monoclonal antibody (mAb) was immobilized on a self-assembled monolayer of 11-mercaptoundecanoic acid (11-MUA) to detect teliospore. All modified electrodes were morphologically characterized by scanning electron microscopy (SEM), atomic force microscopy(AFM), Fourier transform infra-red spectroscopy (FT-IR) techniques and analytically characterized by quartz crystal microbalance (QCM) and cyclic voltammetry (CV). The linearity range was 19 pg mL-1-10 ng mL-1, while the detection limit (LOD) was 4.4 pg mL-1 and 12.5 pg mL-1, respectively. The stability, reproducibility, and repeatability of the immunoelectrode was examined by CV, and found stable upto 18 days with negligible variation. The binding affinity (association constant (Ka)) of the developed immunoelectrode was 1.9 × 10-2 ng mL-1. The real sample has been tested in spiked wheat samples and found about 95-103 % recovery with 2.8-4.4 % relative error.

Metal Organic Framework steered electrosynthesis of anisotropic gold nanorods for specific sensing of organophosphate pesticides in vegetables collected from the field.

The uncontrolled use of organophosphate (OP) group of pesticides has led to their accumulation in food and vegetables, causing major health issues. Hence, the development of a reliable sensor is imperative for the detection of neurotoxic organophosphates (OP). In the present study, we have intertwined the interfaces of a Metal Organic Framework (MOF), MOF-directed rapid electrochemically grown gold nanorods (aAuNR), cysteamine (Cys) functionalization, and the neurotransmitter acetylcholinesterase (AChE) to fabricate a novel electrochemical bioprobe AChE/Cys/aAuNR/MOF/ITO for sensing OP pesticides with an ultra-low detection limit of 3 ng L-1 over a linear range of 30 to 600 ng L-1. Prior to sensing, in silico docking studies were employed for tracking the structural aspects of the molecular recognition of specific OP as potential inhibitors. The sensor can quantify residues of sprayed OP (chlorpyrifos, malathion, parathion, methyl parathion, ethion) in field vegetables (Abelmoschus esculentus, Solanum melongena, Capsicum annuum, Momordica charantia Linn) using a single calibration curve designed using chlorpyrifos, and the results were validated via gas chromatography-electron capture detector (GC-ECD) measurements. The inhibition rate kinetics of structurally different OP (chlorpyrifos, malathion, methyl parathion) were studied via the bioprobe and further validated using the standard Ellman method, confirming the practical applicability of the sensor for the detection of a specific group of OP. The bioprobe AChE/Cys/aAuNR/MOF/ITO offers good stability, specificity, and anti-interference properties for the detection of OP in real samples.

Rapid Electrochemical Monitoring of Bacterial Respiration for Gram-Positive and Gram-Negative Microbes: Potential Application in Antimicrobial Susceptibility Testing.

Antimicrobial resistance is a grave threat to human life. Currently used time-consuming antibiotic susceptibility test (AST) methods limit physicians in selecting proper antibiotics. Hence, we developed a rapid AST using electroanalysis with a 15 min assay time, called EAST, which is live-monitored by time-lapse microscopy video. The present work reports systematical electrochemical analysis and standardization of protocol for EAST measurement. The proposed EAST is successfully applied for Gram-positive Bacillus subtilis and Gram-negative Escherichia coli as model organisms to monitor bacterial concentration, decay kinetics in the presence of various antibiotics (ciprofloxacin, cefixime, and amoxycillin), drug efficacy, and IC50. Bacterial decay kinetics in the presence of antibiotics were validated by the colony counting method, field emission scanning electron microscopy, and atomic force microscopy image analysis. The EAST predicts the antibiotic susceptibility of bacteria within 15 min, which is a significant advantage over existing techniques that consume hours to days. The EAST was explored further by using bacteria-friendly l-lysine-functionalized cerium oxide nanoparticle coated indium tin oxide as a working electrode to observe the enhanced electron-transfer rate in the EAST. The results are very significant for future miniaturization and automation. The proposed EAST has huge potential in the development of a rapid AST device for applications in the clinical and pharmaceutical industries.

Paper based point of care immunosensor for the impedimetric detection of cardiac troponin I biomarker.

Advancements in health care monitoring demand a rapid, accurate and reliable early diagnosis of "Heart Attack" (acute myocardial infarction) with an objective to develop a cost-effective, rapid and label-free point of care diagnostic test kit for the detection of cardiac troponin I (cTnI) on paper-based multi-frequency impedimetric transducers. Paper based sensing platforms were developed by integrating carboxyl group functionalized multi-walled carbon nanotubes (MWCNT) with antibodies of cardiac troponin I (anti-cTnI) biomarker and was characterized using Electrochemical Impedance Spectroscopy (EIS). Various concentrations of cTnI with anti cTnI were studied as a function of impedance change. The suitability of the proposed immunosensor is demonstrated by spiking cTnI in blood serum samples. The limit of detection (LoD) and sensitivity of the proposed sensor was determined to be 0.05 ng/mL and 1.85 mΩ/ng/mL respectively, with a response time of ~1 min. The shelf life of the fabricated sensor was nearly 30 days. The rapid response, very low detection limit, and cost effectiveness offer a portable platform to detect cTnI in blood serum samples. The proposed immunosensor, therefore, offers an affordable healthcare diagnostic platform in resource limited areas.

Bi-enzyme functionalized electro-chemically reduced transparent graphene oxide platform for triglyceride detection.

Recently, increased attention has been drawn to application of graphene and its derivatives for construction of biosensors, since they can be used to rapidly detect the presence of bio-analytes. Present paper establishes the preparation of a unique transducer which relies on toluidine blue (TB), absorbed by electrochemically reduced graphene oxide (ERGO) transparent thin film onto the surface of the indium tin-oxide (ITO) glass electrode. The proposed TB/ERGO/ITO electrode shows excellent reversible electro-chemical properties. The novel platform has been explored to fabricate a triglyceride (TG) biosensor via co-immobilizing of lipase (LIP) and glycerol dehydrogenase (GDH) onto TB/ERGO/ITO electrode surface. The fabricated bioelectrode (LIP-GDH/TB/ERGO/ITO) directly oxidizes glycerol (produced by catalytic hydrolysis of tributyrin acting as a model TG) in the presence of GDH. The developed bioelectrode replaces unstable biological irreversible redox mediators NAD+/NADH, involved in the triglyceride breakdown reaction. NADH causes fouling on the bioelectrode surface in bi-enzymatic estimation of TG and reduces the shelf-life of biosensor. Electrochemical response studies carried out using cyclic voltammetry reveal that the fabricated electrode can detect tributyrin in the range of 50-400 mg dL-1 with high sensitivity of 29 pA mg-1 dL, low response time of 12 s, long-term stability and a low apparent Michaelis-Menten constant (Kappm) of 0.18 mM, indicating high enzyme affinity of LIP-GDH/TB/ERGO/ITO bioelectrode towards tributyrin. Furthermore, this modified bioelectrode has been explored for estimation of TG with negligible interference in human serum samples. The proposed bi-enzymatic bioelectrode for TG analysis offers an efficient and novel interface for application of graphene and its derivatives in the field of bioelectronic devices.

Biosensing Test-Bed Using Electrochemically Deposited Reduced Graphene Oxide.

The development of an efficient test-bed for biosensors requires stable surfaces, capable of interacting with the functional groups present in bioentities. This work demonstrates the formation of highly stable electrochemically reduced graphene oxide (ERGO) thin films reproducibly on indium tin oxide (ITO)-coated glass substrates using a reliable technique through 60 s chronoamperometric reduction of a colloidal suspension maintained at neutral pH containing graphene oxide in deionized water. Structural optimization and biocompatible interactions of the resulting closely packed and uniformly distributed ERGO flakes on ITO surfaces (ERGO/ITO) are characterized using various microscopic and spectroscopic tools. Lipase enzyme is immobilized on the ERGO surface in the presence of ethyl-3-[3-(dimethylamino)propyl]carbodimide and N-hydroxysuccinimide for the detection of triglyceride in a tributyrin (TBN) solution. The ERGO/ITO surfaces prepared using the current technique indicate the noticeable detection of TBN, a source of triglycerides, at a sensitivity of 37 pA mg dL(-1) cm(-2) in the linear range from 50 to 300 mg dL(-1) with a response time of 12 s. The low apparent Michaelies-Menten constant of 0.28 mM suggests high enzyme affinity to TBN. The currently developed fast, simple, highly reproducible, and reliable technique for the formation of an ERGO electrode could be routinely utilized as a test bed for the detection of clinically active bioentities.

Development of transducer matrices based upon nanostructured conducting polymer for application in biosensors.

The nanostructured polyaniline (NSPANI) and its gold nano composite (GNP) with controlled size distribution were developed using structure directing agents (SDA). The nano structure of polyaniline were investigated by UV-Visible spectroscopy, transmission electron microscopy (TEM), dynamic light scattering (DLS), fourrier transform spectroscopy (FTIR), X-ray crystallography and scanning electron microscopy(SEM) etc. These characterization techniques reveal the spherical shape of polyaniline nanoparticles and size in the range of 7-50 nm depending on the type of dopant and nature of SDA. In general, these NSCP colloidal solutions are highly stable. UV-Visible spectra show mainly two peaks at 360-430 nm and at 780-870 nm. The bathochromic shift of the UV-Visible bands as compared to bulk polyaniline, reflect high DC conductivity. TEM and DLS results demonstrate the formation of nanostructure with narrow size distribution. Due to remarkable properties of, it is used as an efficient transduction matrice for the development of highly sensitive, reproducible, stable optical cholesterol and H202 biosensors having wide range of linearity and low Km values.