Nandhinee Radha Shanmugam

Ultrasensitive nanostructure sensor arrays on flexible substrates for multiplexed and simultaneous electrochemical detection of a panel of cardiac biomarkers

Multiplexed detection of protein biomarkers offers new opportunities for early diagnosis and efficient treatment of complex diseases. Cardiovascular diseases (CVDs) has the highest mortality risk in USA and Europe with 15-20 million cases being reported annually. Cardiac Troponins (T and I) are well established protein biomarkers associated with heart muscle damage and point-of-care monitoring of both these two biomarkers has significant benefits on patient care. A flexible disposable electrochemical biosensor device comprising of vertically oriented zinc oxide (ZnO) nanostructures was developed for rapid and simultaneous screening of cardiac Troponin-I (cTnI) and cardiac-Troponin-T (cTnT) in a point-of-care sensor format. The biosensors were designed by selective hydrothermal growth of ZnO nanostructures onto the working electrodes of polyimide printed circuit board platforms, resulting in the generation of high density nanostructure ZnO arrays based electrodes. The size, density and surface terminations of the nanostructures were leveraged towards achieving surface confinement of the target cTnT and cTnI molecules on to the electrode surface. Multiplexing and simultaneous detection was achieved through sensor platform design comprising of arrays of Troponin functionalized ZnO nanostructure electrodes. The sensitivity and specificity of the biosensor was characterized using two types of electrochemical techniques; electrochemical impedance spectroscopy (EIS) and Mott-Schottky analysis on the same sensor platform to demonstrate multi-configurable modes. Limit of detection of 1pg/mL in human serum was achieved for both cTnI and cTnT. Cross reactivity analysis showed the selectivity of detecting cTnT and cTnI in human serum with wide dynamic range.

Ultrasensitive and low-volume point-of-care diagnostics on flexible strips – a study with cardiac troponin biomarkers

We demonstrate a flexible, mechanically stable, and disposable electrochemical sensor platform for monitoring cardiac troponins through the detection and quantification of cardiac Troponin-T (cTnT). We designed and fabricated nanostructured zinc oxide (ZnO) sensing electrodes on flexible porous polyimide substrates. We demonstrate ultrasensitive detection is capable at very low sample volumes due to the confinement phenomenon of target species within the ZnO nanostructures leading to enhancement of biomolecular binding on the sensor electrode surface. The performance of the ZnO nanostructured sensor electrode was evaluated against gold and nanotextured ZnO electrodes. The electrochemical sensor functions on affinity based immunoassay principles whereby monoclonal antibodies for cTnT were immobilized on the sensor electrodes using thiol based chemistry. Detection of cTnT in phosphate buffered saline (PBS) and human serum (HS) buffers was achieved at low sample volumes of 20 μL using non-faradaic electrochemical impedance spectroscopy (EIS). Limit of detection (LOD) of 1E-4 ng/mL (i.e. 1 pg/mL) at 7% CV (coefficient of variation) for cTnT in HS was demonstrated on nanostructured ZnO electrodes. The mechanical integrity of the flexible biosensor platform was demonstrated with cyclic bending tests. The sensor performed within 12% CV after 100 bending cycles demonstrating the robustness of the nanostructured ZnO electrochemical sensor platform.

Electrochemical nanostructured ZnO biosensor for ultrasensitive detection of cardiac troponin-T

AIM Vertically oriented zinc oxide nanostructures based disposable diagnostic biosensor for detecting and quantifying levels of cardiac troponin-T from human serum has been developed. MATERIALS & METHODS The biosensors were designed by integrating hydrothermally grown zinc oxide nanostructures on glass and printed circuit board platforms, resulting in the generation of high-density nanostructure arrays with nanotextured zinc oxide based electrodes. The size, density and surface terminations of the nanostructures were leveraged toward achieving surface confinement of the target cTnT molecules on to the nanostructures. A combination of AC and DC spectroscopy was used to characterize the biosensor response to cTnT. RESULTS & CONCLUSION LOD of 0.1 pg/ml in human serum was achieved.

Novel Nanomonitor ultra-sensitive detection of troponin T

BACKGROUND Troponin is the preferred biomarker for diagnosing myocardial infarction. Point of care devices have not matched the sensitivity of laboratory-based methods for measuring troponin. The Nanomonitor is a novel point-of-care device that uses the change in electrical impedance that occurs when a biomarker binds to its antibody, which is then correlated to the concentration of the target biomarker. METHODS Performance characteristics of the Nanomonitor were evaluated and compared to a standard laboratory-based method. RESULTS The limit of detection of the Nanomonior for troponin T was 0.0088ng/l. Total imprecission was 2.38% and 0.85% at troponin T concentrations of 73ng/l and 1800ng/l. The functional sensitivity (10% coeffecient of variation) was 0.329ng/l. The linear regression had a slope of 0.996 (95% confidence interval, 0.991, 1.002), r=1.00, and an intercept of 15.88ng/l (95% confidence interval, -68.39ng/l, 100.15ng/l). The mean difference between the assays was -7.54ng/l, determined by Bland-Altman analysis. CONCLUSION The Nanomonitor preliminary results have favorable performance characteristics for detecting troponin T in patient blood, provide results in 15min, and are portable. More research is needed.

A novel approach for electrical tuning of nano-textured zinc oxide surfaces for ultra-sensitive troponin-T detection

We have developed a label-free, non-faradaic, electrochemical sensor for ultra-sensitive detection of a cardiac biomarker, troponin-T by utilizing the stoichiometric surface compositions of nanotextured zinc oxide (ZnO) thin films. In this study, we show how the performance of a nanotextured zinc oxide based non-faradaic biosensor is modulated by differences in the fabrication parameters of the metal oxide thin film as well as the choice of cross-linkers. Two cross-linking molecules, dithiobis succinimidyl propionate and 3-aminopropyl triethoxysilane, demonstrate significantly different binding chemistries with zinc oxide. The non-faradaic electrochemical behaviour of the sensor due to the two linkers is compared by analyzing the troponin-T dose response using electrochemical impedance spectroscopy (EIS). The sensor performance associated with both linkers is compared based on the dynamic range and limit of detection. The sensor utilizing zinc surface terminations demonstrated a wider dynamic range between the two linkers. This range extended from 26% to 54% in phosphate buffered saline and from 21% to 65% in human serum, for a concentration range from 10 fg mL−1 to 1 ng mL−1 of troponin-T. The limit of detection was found to be at 10 fg mL−1 and has potential utility in the development of point-of-care (POC) diagnostics for cardiovascular diseases. Fluorescence quantification analysis was also performed to further validate the specificity of the linker binding to the ZnO films. An ultrasensitive troponin-T biosensor can be designed by leveraging the zinc termination based surface chemistry for selective protein immobilization.

Flexible Molybdenum Electrodes towards Designing Affinity Based Protein Biosensors

Molybdenum electrode based flexible biosensor on porous polyamide substrates has been fabricated and tested for its functionality as a protein affinity based biosensor. The biosensor performance was evaluated using a key cardiac biomarker; cardiac Troponin-I (cTnI). Molybdenum is a transition metal and demonstrates electrochemical behavior upon interaction with an electrolyte. We have leveraged this property of molybdenum for designing an affinity based biosensor using electrochemical impedance spectroscopy. We have evaluated the feasibility of detection of cTnI in phosphate-buffered saline (PBS) and human serum (HS) by measuring impedance changes over a frequency window from 100 mHz to 1 MHz. Increasing changes to the measured impedance was correlated to the increased dose of cTnI molecules binding to the cTnI antibody functionalized molybdenum surface. We achieved cTnI detection limit of 10 pg/mL in PBS and 1 ng/mL in HS medium. The use of flexible substrates for designing the biosensor demonstrates promise for integration with a large-scale batch manufacturing process.

Portable nanoporous electrical biosensor for ultrasensitive detection of Troponin-T

Aim: To demonstrate the design, fabrication and testing of a portable, label-free biosensor for ultrasensitive detection of the cardiac Troponin-T (cTnT) from patient blood. Materials & methods: The biosensor is comprised of a nanoporous membrane integrated on to a microelectrode sensor platform for nanoconfinement effects. Charge perturbations due to antigen binding are recorded as impedance changes using electrochemical impedance spectroscopy. Results: The measured impedance change is used to quantitatively determine the cTnT concentration from the tested sample. We were successful in detecting and quantifying cardiac Troponin-T from a 40-patient cohort. The limit of detection was 0.01 pg/ml. Conclusion: This novel technology has promising preliminary results for rapid and sensitive detection of cTnT.

Surface modification of ZnO nanostructured electrodes with thiol and phosphonic acid moieties for biosensing applications

The analytical performance of affinity immunoassay based biosensor systems is determined by the efficiency of capture probe immobilization, which in turn depends on the stability of surface modification. In this study, we investigate the functionalization of nanostructured ZnO surfaces through two types of crosslinker molecules – DSP that favors thiol chemistry and 11-AUPA that favors phosphonic acid bonding. The type of interaction and surface composition of functionalized ZnO surfaces were evaluated using Fourier Transform Infrared Spectroscopy (FT-IR) and X-ray photoelectron spectroscopy (XPS) measurements. The crosslinker interaction with ZnO surfaces and subsequent biomolecular binding events influence the inherent electrical properties at the ZnO electrode–electrolyte interface. The changes in charge perturbations due to these events are leveraged for the sensing of target biomolecules, and cardiac troponin-I (cTnI) is investigated using Electrochemical Impedance Spectroscopy and Mott–Schottky measurements. Our results demonstrate that Zn–S formation is more sensitive for the detection of cTnI as compared to the ZnO–AUPA interactions. The dynamic range of detection is from 0.1 pg mL−1 to 100 000 pg mL−1 with a limit of detection at 0.1 pg mL−1 in human serum.

A review on ZnO-based electrical biosensors for cardiac biomarker detection

Over the past few decades zinc oxide (ZnO)-based thin films and nanostructures have shown unprecedented performance in a wide range of applications. In particular, owing to high isoelectric point, biocompatibility and other multifunctional characteristics, ZnO has extensively been studied as a transduction material for biosensor development. The fascinating properties of ZnO help retain biological activity of the immobilized biomolecule and help in achieving enhanced sensing performance. As a consequence of recent advancements in this multidisciplinary field, diagnostic biosensors are expanding beyond traditional clinical labs to point-of-care and home settings. Label-free electrical detection of biomarkers has been demonstrated using ZnO-sensing platforms. In this review we highlight the characteristics of ZnO that enable realization of its use in development of point-of-care biosensors toward disease diagnosis, in particular cardiovascular diseases.

Zinc Oxide Nanostructures as Electrochemical Biosensors on Flexible Substrates

A novel flexible electrochemical biosensor for protein biomarker detection was successfully designed and fabricated on a nanoporous polyimide membrane using zinc oxide (ZnO). Nanostructures of ZnO were grown on microelectrode platform using aqueous solution bath. Electrochemical measurements were performed using gold, ZnO seed and nanostructured electrodes to study the influence of electrode surface area on biosensing performance. Feasibility analysis of sensor platforms was evaluated using high concentrations (in ng/mL) of troponin-T. The results showed that improved performance can be obtained on nanostructured platform by careful optimization of growth conditions. This study demonstrates the development of nanostructured ZnO flexible biosensors towards ultra-sensitive protein biosensing.Copyright