Os(II/III) complex supports pH-insensitive electrochemical DNA-based sensing with superior operational stability than the benchmark methylene blue reporter.

DNA-based electrochemical sensors use redox reporters to transduce affinity events into electrical currents. Ideally, such reporters must be electrochemically reversible, chemically stable for thousands of redox cycles, and tolerant to changing chemical environments. Here we report the first use of an Os(II/III) complex in DNA-based sensors, which undergoes pH-insensitive electron transfer with 35% better operational stability relative to the benchmark methylene blue, making it a promising reporter for continuous molecular monitoring applications where pH fluctuates with time.

Optimization of Vancomycin Aptamer Sequence Length Increases the Sensitivity of Electrochemical, Aptamer-Based Sensors In Vivo.

The measurement of serum vancomycin levels at the clinic is critical to optimizing dosing given the narrow therapeutic window of this antibiotic. Current approaches to quantitate serum vancomycin levels are based on immunoassays, which are multistep methods requiring extensive processing of patient samples. As an alternative, vancomycin-binding electrochemical, aptamer-based sensors (E-ABs) were developed to simplify the workflow of vancomycin monitoring. E-ABs enable the instantaneous measurement of serum vancomycin concentrations without the need for sample dilution or other processing steps. However, the originally reported vancomycin-binding E-ABs had a dissociation constant of 45 µM, which is approximately 1 order of magnitude higher than the recommended trough concentrations of vancomycin measured in patients. This limited sensitivity hinders the ability of E-ABs to accurately support vancomycin monitoring. To overcome this problem, here we sought to optimize the length of the vancomycin-binding aptamer sequence to enable a broader dynamic range in the E-AB platform. Our results demonstrate, via isothermal calorimetry and E-AB calibrations in undiluted serum, that superior affinity and near-equal sensor gain in vitro can be achieved using a one-base-pair-longer aptamer than the truncated sequence originally reported. We validate the impact of the improved binding affinity in vivo by monitoring vancomycin levels in the brain cortex of live mice following intravenous administration. While the original sequence fails to resolve vancomycin concentrations from baseline noise (SNR = 1.03), our newly reported sequence provides an SNR of 1.62 at the same dose.

Antibody-Invertase Fusion Protein Enables Quantitative Detection of SARS-CoV-2 Antibodies Using Widely Available Glucometers.

Rapid diagnostics that can accurately inform patients of disease risk and protection are critical to mitigating the spread of the current COVID-19 pandemic and future infectious disease outbreaks. To be effective, such diagnostics must rely on simple, cost-effective, and widely available equipment and should be compatible with existing telehealth infrastructure to facilitate data access and remote care. Commercial glucometers are an established detection technology that can overcome the cost, time, and trained personnel requirements of current benchtop-based antibody serology assays when paired with reporter molecules that catalyze glucose conversion. To this end, we developed an enzymatic reporter that, when bound to disease-specific patient antibodies, produces glucose in proportion to the level of antibodies present in the patient sample. Although a straightforward concept, the coupling of enzymatic reporters to secondary antibodies or antigens often results in low yields, indeterminant stoichiometry, reduced target binding, and poor catalytic efficiency. Our enzymatic reporter is a novel fusion protein that comprises an antihuman immunoglobulin G (IgG) antibody genetically fused to two invertase molecules. The resulting fusion protein retains the binding affinity and catalytic activity of the constituent proteins and serves as an accurate reporter for immunoassays. Using this fusion, we demonstrate quantitative glucometer-based measurement of anti-SARS-CoV-2 spike protein antibodies in blinded clinical sample training sets. Our results demonstrate the ability to detect SARS-CoV-2-specific IgGs in patient serum with precise agreement to benchmark commercial immunoassays. Because our fusion protein binds all human IgG isotypes, it represents a versatile tool for detection of disease-specific antibodies in a broad range of biomedical applications.

A self-powered skin-patch electrochromic biosensor.

One of the limitations of many skin-patch wearable sensors today is their dependence on silicon-based electronics, increasing their complexity and unit cost. Self-powered sensors, in combination with electrochromic materials, allow simplifying the construction of these devices, leading to powerful analytical tools that remove the need for external detection systems. This work describes the construction, by screen-printing, of a self-powered electrochromic device that can be adapted for the determination of metabolites in sweat by the naked eye in the form of a 3 × 15 mm colour bar. The device comprises a lactate oxidase and osmium-polymer -based anode connected to a coplanar 3 × 15 mm Prussian Blue, PB, cathode printed over a transparent poly(3,4-ethylenedioxythiophene) polystyrene sulfonate, PEDOT:PSS electrode. An ion-gel composed of Poly(vinylidene fluoride-co-hexafluoropropylene), PVDF-co-HFP, a gelling agent, and ionic liquid 1-Ethyl-3-methylimidazolium trifluoromethanesulfonate, EMIM-Tf, effectively separates the cathode display from the biosensing anode, protecting it from the sample. Despite its cathodic electrochromism, the PEDOT:PSS has a transmission above 90% and does not mask the Prussian Blue colour change because the cathode does not operate below 0 V vs Ag/AgCl at any time. The sensor displays lactate concentrations in the range of 0-10 mM over the length of the electrochromic display, which has a contrast ratio of 1.43. Although full response takes up to 24 min, 85% of the colour change is displayed within 10 min.

Rapid prototyping of electrochemical lateral flow devices: stencilled electrodes.

A straightforward and very cost effective method is proposed to prototype electrodes using pressure sensitive adhesives (PSA) and a simple cutting technique. Two cutting methods, namely blade cutting and CO2 laser ablation, are compared and their respective merits are discussed. The proposed method consists of turning the protective liner on the adhesive into a stencil to apply screen-printing pastes. After the electrodes have been printed, the liner is removed and the PSA can be used as a backing material for standard lateral flow membranes. We present the fabrication of band electrodes down to 250 µm wide, and their characterization using microscopy techniques and cyclic voltammetry. The prototyping approach presented here facilitates the development of new electrochemical devices even if very limited fabrication resources are available. Here we demonstrate the fabrication of a simple lateral-flow device capable of determining glucose in blood. The prototyping approach presented here is highly suitable for the development of novel electroanalytical tools.