Reduced graphene oxide electrodes meet lateral flow assays: A promising path to advanced point-of-care diagnostics.

Research in electrochemical detection in lateral flow assays (LFAs) has gained significant momentum in recent years. The primary impetus for this surge in interest is the pursuit of achieving lower limits of detection, especially given that LFAs are the most widely employed point-of-care biosensors. Conventionally, the strategy for merging electrochemistry and LFAs has centered on the superposition of screen-printed electrodes onto nitrocellulose substrates during LFA fabrication. Nevertheless, this approach poses substantial limitations regarding scalability. In response, we have developed a novel method for the complete integration of reduced graphene oxide (rGO) electrodes into LFA strips. We employed a CO2 laser to concurrently reduce graphene oxide and pattern nitrocellulose, exposing its backing to create connection sites impervious to sample leakage. Subsequently, rGO and nitrocellulose were juxtaposed and introduced into a roll-to-roll system using a wax printer. The exerted pressure facilitated the transfer of rGO onto the nitrocellulose. We systematically evaluated several electrochemical strategies to harness the synergy between rGO and LFAs. While certain challenges persist, our rGO transfer technology presents compelling potential for setting a new standard in electrochemical LFA fabrication.

Non-monetary narratives motivate businesses to engage with climate change.

The dominant narrative to motivate business actors to take climate actions emphasizes opportunities to increase monetary gains, linking sustainability to the financial goals of these organizations. The prevalence of monetary motivations in sustainability communication among businesses, consultancies, academics and international organizations has made this narrative a truism in the private sector. We conducted an online, real-world, large-n experiment to evaluate the comparative effectiveness of different motivations using narrative communication. We show that non-monetary narratives highlighting prosocial or achievement motivations are 55% more effective in creating responses from businesses than narratives emphasizing monetary gains. These findings are robust across most narrative and audience characteristics, including age and language. Our findings suggest that communication towards business leaders around sustainability can be multi-pronged and should incorporate prosocial and achievement motivations aside from articulating potential financial benefits.

The Cleanroom-Free, Cheap, and Rapid Fabrication of Nanoelectrodes with Low zM Limits of Detection.

Nanoscale electrodes have been a topic of intense research for many decades. Their enhanced sensitivities, born out of an improved signal-to-noise ratio as electrode dimensions decrease, make them ideal for the development of low-concentration analyte sensors. However, to date, nanoelectrode fabrication has typically required expensive equipment and exhaustive, time-consuming fabrication methods that have rendered them unsuitable for widespread use and commercialization. Herein, a method of nanoband electrode fabrication using low cost materials and equipment commonly found in research laboratories around the world is reported. The materials' cost to produce each nanoband is less than €0.01 and fabrication of a batch takes less than 1 h. The devices can be made of flexible plastics and their designs can be quickly and easily iterated. Facile methods of combining these nanobands into powerful devices, such as complete three-electrode systems, are also displayed. As a proof of concept, the electrodes are functionalized for the detection of a DNA sequence specific to SARS-CoV-2 and found to display single molecule sensitivity.

Electrochemically Controlled Hydrogels with Electrotunable Permeability and Uniaxial Actuation.

The unique properties of hydrogels enable the design of life-like soft intelligent systems. However, stimuli-responsive hydrogels still suffer from limited actuation control. Direct electronic control of electronically conductive hydrogels can solve this challenge and allow direct integration with modern electronic systems. An electrochemically controlled nanowire composite hydrogel with high in-plane conductivity that stimulates a uniaxial electrochemical osmotic expansion is demonstrated. This materials system allows precisely controlled shape-morphing at only -1 V, where capacitive charging of the hydrogel bulk leads to a large uniaxial expansion of up to 300%, caused by the ingress of ≈700 water molecules per electron-ion pair. The material retains its state when turned off, which is ideal for electrotunable membranes as the inherent coupling between the expansion and mesoporosity enables electronic control of permeability for adaptive separation, fractionation, and distribution. Used as electrochemical osmotic hydrogel actuators, they achieve an electroactive pressure of up to 0.7 MPa (1.4 MPa vs dry) and a work density of ≈150 kJ m-3 (2 MJ m-3  vs dry). This new materials system paves the way to integrate actuation, sensing, and controlled permeation into advanced soft intelligent systems.

An Artificial Miniaturized Peroxidase for Signal Amplification in Lateral Flow Immunoassays.

Signal amplification strategies are widely used for improving the sensitivity of lateral flow immunoassays (LFiAs). Herein, the artificial miniaturized peroxidase Fe(III)-MimochromeVI*a (FeMC6*a), immobilized on gold nanoparticles (AuNPs), is used as a strategy to obtain catalytic signal amplification in sandwich immunoassays on lateral flow strips. The assay scheme uses AuNPs decorated with the mini-peroxidase FeMC6*a and anti-human-IgG as a detection antibody (dAb), for the detection of human-IgG, as a model analyte. Recognition of the analyte by the capture and detection antibodies is first evidenced by the appearance of a red color in the test line (TL), due to the accumulation of AuNPs. Subsequent addition of 3,3',5,5'-tetramethylbenzidine (TMB) induces an increase of the test line color, due to the TMB being converted into an insoluble colored product, catalyzed by FeMC6*a. This work shows that FeMC6*a acts as an efficient catalyst in paper, increasing the sensitivity of an LFiA up to four times with respect to a conventional LFiA. Furthermore, FeMC6*a achieves lower limits of detection that are found in control experiments where it is replaced with horseradish peroxidase (HRP), its natural counterpart. This study represents a significant proof-of-concept for the development of more sensitive LFiAs, for different analytes, based on properly designed artificial metalloenzymes.

Laser Reduced Graphene Oxide Electrode for Pathogenic Escherichia coli Detection.

Graphene-based materials are of interest in electrochemical biosensing due to their unique properties, such as high surface areas, unique electrochemical properties, and biocompatibility. However, the scalable production of graphene electrodes remains a challenge; it is typically slow, expensive, and inefficient. Herein, we reported a simple, fast, and maskless method for large-scale, low-cost reduced graphene oxide electrode fabrication; using direct writing (laser scribing and inkjet printing) coupled with a stamp-transferring method. In this process, graphene oxide is simultaneously reduced and patterned with a laser, before being press-stamped onto polyester sheets. The transferred electrodes were characterized by SEM, XPS, Raman, and electrochemical methods. The biosensing utility of the electrodes was demonstrated by developing an electrochemical test for Escherichia coli. These biosensors exhibited a wide dynamic range (917-2.1 × 107 CFU/mL) of low limits of detection (283 CFU/mL) using just 5 µL of sample. The test was also verified in spiked artificial urine, and the sensor was integrated into a portable wireless system driven and measured by a smartphone. This work demonstrates the potential to use these biosensors for real-world, point-of-care applications. Hypothetically, the devices are suitable for the detection of other pathogenic bacteria.

Toward Next Generation Lateral Flow Assays: Integration of Nanomaterials.

Lateral flow assays (LFAs) are currently the most used point-of-care sensors for both diagnostic (e.g., pregnancy test, COVID-19 monitoring) and environmental (e.g., pesticides and bacterial monitoring) applications. Although the core of LFA technology was developed several decades ago, in recent years the integration of novel nanomaterials as signal transducers or receptor immobilization platforms has brought improved analytical capabilities. In this Review, we present how nanomaterial-based LFAs can address the inherent challenges of point-of-care (PoC) diagnostics such as sensitivity enhancement, lowering of detection limits, multiplexing, and quantification of analytes in complex samples. Specifically, we highlight the strategies that can synergistically solve the limitations of current LFAs and that have proven commercial feasibility. Finally, we discuss the barriers toward commercialization and the next generation of LFAs.

A disposable, wearable, flexible, stitched textile electrochemical biosensing platform.

Wearable sensors are a fast growing and exciting research area, the success of smart watches are a great example of the utility and demand for wearable sensing systems. The current state of the art routinely uses expensive and bulky equipment designed for long term use. There is a need for cheap and disposable wearable sensors to make single use measurements, primarily in the area of biomarker detection. Herein we report the ability to make cheap (0.22 USD/sensor), disposable, wearable sensors by stitching conductive gold coated threads into fabrics. These threads are easily functionalised with thiolate self-assembled monolayers which can be designed for the detection of a broad range of different biomarkers. This all textile sensing platform is ideally suited to be scaled up and has the added advantage of being stretchable with insignificant effect on the electrochemistry of the devices. As a proof of principle, the devices have been functionalised with a continuous glucose sensing system which was able to detect glucose in human sweat across the clinically relevant range (0.1-0.6 mM). The sensors have a sensitivity of 126 ± 14 nA/mM of glucose and a limit of detection of 301 ± 2 nM. This makes them ideally suited for biomarker detection in point-of-care sensing applications.

Woven Electroanalytical Biosensor for Nucleic Acid Amplification Tests.

Fiber-based biosensors enable a new approach in analytical diagnostic devices. The majority of textile-based biosensors, however, rely on colorimetric detection. Here a woven biosensor that integrates microfluidics structures in combination with an electroanalytical readout based on a thiol-self-assembled monolayer (SAM) for Nucleic Acid Amplification Testing, NAATs is shown. Two types of fiber-based electrodes are systematically characterized: pure gold microwires (bond wire) and off-the-shelf plasma gold-coated polyester multifilament threads to evaluate their potential to form SAMs on their surface and their electrochemical performance in woven textile. A woven electrochemical DNA (E-DNA) sensor using a SAM-based stem-loop probe-modified gold microwire is fabricated. These sensors can specifically detect unpurified, isothermally amplified genomic DNA of Staphylococcus epidermidis (10 copies/µL) by recombinase polymerase amplification (RPA). This work demonstrates that textile-based biosensors have the potential for integrating and being employed as automated, sample-to-answer analytical devices for point-of-care (POC) diagnostics.

Weaving Off-The-Shelf Yarns into Textile Micro Total Analysis Systems (µTAS).

Textile based biosensors have garnered much interest in recent years. Devices woven out of yarns have the ability to be incorporated into clothing and bandages. Most woven devices reported in the literature require yarns that are not available on an industrial scale or that require modifications which are not possible in large scale manufacturing. In this work, commercially produced yarns are taken without any modification or cleaning, and developed woven textile diagnostic devices out of them. The yarn properties that are important to their function within the device have been characterised and discussed. The wicking ability and analyte retention of Coolmax yarns, developed to wick sweat in mass produced sportswear, are determined. The electrochemistry and functionalizability of Au coated multifilament yarns are investigated with no cleaning or treatment and are found to have as good a thiolate self-assembled monolayer (SAM) coverage as cleaned Au disk electrodes. The feasibility of using these yarns is established off the shelf, with no cleaning, to make woven capillary force driven microfluidic devices and three electrode sensing devices. A proof of principle three electrode system capable of detecting clinically relevant concentrations of glucose in human sweat is reported.

Electrochemical Detection of Genomic DNA Utilizing Recombinase Polymerase Amplification and Stem-Loop Probe.

Nucleic acid tests integrated into digital point-of-care (POC) diagnostic systems have great potential for the future of health care. However, current methods of DNA amplification and detection require bulky and expensive equipment, many steps, and long process times, which complicate their integration into POC devices. We have combined an isothermal DNA amplification method, recombinase polymerase amplification, with an electrochemical stem-loop (S-L) probe DNA detection technique. By combining these methods, we have created a system that is able to specifically amplify and detect as few as 10 copies/µL Staphylococcus epidermidis DNA with a total time to result of 70-75 min.

Insulin Receptor Substrate-1 (IRS-1) and IRS-2 expression levels are associated with prognosis in non-small cell lung cancer (NSCLC).

The insulin-like growth factor-1 (IGF-1) signaling pathway has been implicated in non-small cell lung cancer (NSCLC) outcomes and resistance to targeted therapies. However, little is known regarding the molecular mechanisms by which this pathway contributes to the biology of NSCLC. The insulin receptor substrate (IRS) proteins are cytoplasmic adaptor proteins that signal downstream of the IGF-1R and determine the functional outcomes of this signaling pathway. In this study, we assessed the expression patterns of IRS-1 and IRS-2 in NSCLC to identify associations between IRS-1 and IRS-2 expression levels and survival outcomes in the two major histological subtypes of NSCLC, adenocarcinoma (ADC) and squamous cell carcinoma (SCC). High IRS-2 expression was significantly associated with decreased overall survival in adenocarcinoma (ADC) patients, whereas low IRS-1 cytoplasmic expression showed a trend toward association with decreased overall survival in squamous cell carcinoma (SCC) patients. Tumors with low IRS-1 and high IRS-2 expression were found to be associated with poor outcomes in ADC and SCC, indicating a potential role for IRS-2 in the aggressive behavior of NSCLC. Our results suggest distinct contributions of IRS-1 and IRS-2 to the biology of ADC and SCC that impact disease progression.

Functionalised microscale nanoband edge electrode (MNEE) arrays: the systematic quantitative study of hydrogels grown on nanoelectrode biosensor arrays for enhanced sensing in biological media.

Nanoelectrodes and nanoelectrode arrays show enhanced diffusion and greater faradaic current densities and signal-to-noise ratios compared to macro and microelectrodes, which can lead to enhanced sensing and detection. One example is the microsquare nanoband edge electrode (MNEE) array system, readily formed through microfabrication and whose quantitative response has been established electroanalytically. Hydrogels have been shown to have applications in drug delivery, tissue engineering, and anti-biofouling; some also have the ability to be grown electrochemically. Here, we combine these two emerging technologies to demonstrate the principles of a hydrogel-coated nanoelectrode array biosensor that is resistant to biofouling. We first electrochemically grow and analyze hydrogels on MNEE arrays. The structure of these gels is shown by imaging to be electrochemically controllable, reproducible and structurally hierarchical. This structure is determined by the MNEE array diffusion fields, consistent with the established hydrogel formation reaction, and varies in structural scale from nano (early time, near electrode growth) to micro (for isolated elements in the array) to macro (when there is array overlap) with distance from the electrode, forming a hydrogel mesh of increasing density on progression from solution to electrode. There is also increased hydrogel structural density observed at electrode corners, attributable to enhanced diffusion. The resulting hydrogel structure can be formed on (and is firmly anchored to/through) an established clinically relevant biosensing layer without compromising detection. It is also shown to be capable, through proof-of-principle model protein studies using bovine serum albumin (BSA), of preventing protein biofouling whilst enabling smaller molecules such as DNA to pass through the hydrogel matrix and be sensed. Together, this demonstrates a method for developing reproducible, quantitative electrochemical nanoelectrode biosensors able to sense selectively in real-world sample matrices through the tuning of their interfacial properties.

Correction: Impedimetric measurement of DNA-DNA hybridisation using microelectrodes with different radii for detection of methicillin resistant Staphylococcus aureus (MRSA).

Correction for 'Impedimetric measurement of DNA-DNA hybridisation using microelectrodes with different radii for detection of methicillin resistant Staphylococcus aureus (MRSA)' by Poh Quan Li et al., Analyst, 2017, 142, 1946-1952.

Impedimetric measurement of DNA-DNA hybridisation using microelectrodes with different radii for detection of methicillin resistant Staphylococcus aureus (MRSA).

Due to their electroanalytical advantages, microelectrodes are a very attractive technology for sensing and monitoring applications. One highly important application is measurement of DNA hybridisation to detect a wide range of clinically important phenomena, including single nucleotide polymorphisms (SNPs), mutations and drug resistance genes. The use of electrochemical impedance spectroscopy (EIS) for measurement of DNA hybridisation is well established for large electrodes but as yet remains relatively unexplored for microelectrodes due to difficulties associated with electrode functionalisation and impedimetric response interpretation. To shed light on this, microelectrodes were initially fabricated using photolithography and characterised electrochemically to ensure their responses matched established theory. Electrodes with different radii (50, 25, 15 and 5 µm) were then functionalised with a mixed film of 6-mercapto-1-hexanol and a thiolated single stranded DNA capture probe for a specific gene from the antibiotic resistant bacterium MRSA. The complementary oligonucleotide target from the mecA MRSA gene was hybridised with the surface tethered ssDNA probe. The EIS response was evaluated as a function of electrode radius and it was found that charge-transfer (RCT) was more significantly affected by hybridisation of the mecA gene than the non-linear resistance (RNL) which is associated with the steady state current. The discrimination of mecA hybridisation improved as electrode radius reduced with the RCT component of the response becoming increasingly dominant for smaller radii. It was possible to utilise these findings to produce a real time measurement of oligonucleotide binding where changes in RCT were evident one minute after nanomolar target addition. These data provide a systematic account of the effect of microelectrode radius on the measurement of hybridisation, providing insight into critical aspects of sensor design and implementation for the measurement of clinically important DNA sequences. The findings open up the possibility of developing rapid, sensitive DNA based measurements using microelectrodes.

Using the pimeloyl-CoA synthetase adenylation fold to synthesize fatty acid thioesters.

Biotin is an essential vitamin in plants and mammals, functioning as the carbon dioxide carrier within central lipid metabolism. Bacterial pimeloyl-CoA synthetase (BioW) acts as a highly specific substrate-selection gate, ensuring the integrity of the carbon chain in biotin synthesis. BioW catalyzes the condensation of pimelic acid (C7 dicarboxylic acid) with CoASH in an ATP-dependent manner to form pimeloyl-CoA, the first dedicated biotin building block. Multiple structures of Bacillus subtilis BioW together capture all three substrates, as well as the intermediate pimeloyl-adenylate and product pyrophosphate (PPi), indicating that the enzyme uses an internal ruler to select the correct dicarboxylic acid substrate. Both the catalytic mechanism and the surprising stability of the adenylate intermediate were rationalized through site-directed mutagenesis. Building on this understanding, BioW was engineered to synthesize high-value heptanoyl (C7) and octanoyl (C8) monocarboxylic acid-CoA and C8 dicarboxylic-CoA products, highlighting the enzyme's synthetic potential.

Differential involvement of the microtubule cytoskeleton in insulin receptor substrate 1 (IRS-1) and IRS-2 signaling to AKT determines the response to microtubule disruption in breast carcinoma cells.

The insulin receptor substrate (IRS) proteins serve as essential signaling intermediates for the activation of PI3K by both the insulin-like growth factor 1 receptor (IGF-1R) and its close family member, the insulin receptor (IR). Although IRS-1 and IRS-2 share significant homology, they regulate distinct cellular responses downstream of these receptors and play divergent roles in breast cancer. To investigate the mechanism by which signaling through IRS-1 and IRS-2 results in differential outcomes, we assessed the involvement of the microtubule cytoskeleton in IRS-dependent signaling. Treatment with drugs that either stabilize or disrupt microtubules reveal that an intact microtubule cytoskeleton contributes to IRS-2- but not IRS-1-mediated activation of AKT by IGF-1. Proximal IGF-1R signaling events, including IRS tyrosine phosphorylation and recruitment of PI3K, are not inhibited by microtubule disruption, indicating that IRS-2 requires the microtubule cytoskeleton at the level of downstream effector activation. IRS-2 colocalization with tubulin is enhanced upon Taxol-mediated microtubule stabilization, which, together with the signaling data, suggests that the microtubule cytoskeleton may facilitate access of IRS-2 to downstream effectors such as AKT. Of clinical relevance is that our data reveal that expression of IRS-2 sensitizes breast carcinoma cells to apoptosis in response to treatment with microtubule-disrupting drugs, identifying IRS-2 as a potential biomarker for the response of breast cancer patients to Vinca alkaloid drug treatment.

Ultrashort-pulse Yb3+-fiber-based laser and amplifier system producing >25-W average power.

We have demonstrated an ultrashort-pulse Yb3+-fiber laser and amplifier system that produces >400-nJ pulses at a repetition rate of 62 MHz (>25-W average power). The output pulses were recompressed to a duration of 110 fs with good pulse quality by use of a standard bulk grating-based compressor.