Here, we report a novel organic electrochemical transistor (OECT)-based strategy for the real-time monitoring of biofilms in flow culture. Bacteria commonly form biofilms to survive an unfavorable environment. Protected by their featured extracellular polymeric substances (EPS), biofilm-related diseases are difficult to treat using normal antibacterial treatments like antibiotics. The development of new antibiofilm agents requires real-time monitoring techniques to evaluate the response of biofilms to treatments. The OECT-based platform reported here realized the online investigation of biofilms by utilizing the intrinsic sensitivity of OECTs in non-faradic impedimetric biosensing. It achieved the capability of distinguishing different stages in the biofilm life cycle through an extended-gate design of the OECT device integrated with a microfluidic-based flow culture chip. We further explored using this system as an evaluation platform for novel antibiofilm agent development. The OECT platform successfully identified the unique effectiveness of extracted porcine gastric mucin (PGM) in treating mature biofilms in flow culture, outperforming polymyxin B.

CRISPR–Cas effectors typically rely on RNA guides to recognize target sequences. In Cas12a, the protospacer adjacent motif on DNA engages conserved protein residues, triggering target binding and nuclease activation. Here we reprogram Cas12a into a DNA-guided, RNA-targeting effector. Exploiting protospacer-adjacent motif-dependent interaction, we engineer synthetic CRISPR DNA that engages Cas12a to form a functional deoxyribonucleoprotein complex, while repurposing solely RNA as the programmable target. Structural, biophysical and biochemical analyses reveal the molecular basis of this DNA-guided, RNA-targeting configuration and support an activation pathway distinct from that of canonical RNA-guided systems. DNA-guided Cas12a enables direct RNA detection and efficient intracellular RNA knockdown, establishing a modular activation architecture for CRISPR–Cas12a and expanding the design space for programmable RNA manipulation. Synthetic DNA guides (crDNA) reprogram Cas12a nucleases for RNA targeting.

One-pot CRISPR diagnostics face a fundamental incompatibility: isothermal nucleic acid amplification enables rapid target accumulation, whereas CRISPR activation irreversibly consumes those substrates, destabilizing reaction kinetics. Here we show that reaction order can be programmed into DNA primers through thermodynamic design. Differences in primer-binding strength create two sequential amplification stages, delaying CRISPR activation until enough amplicons have accumulated without physical separation or external control. The design also introduces the protospacer adjacent motif (PAM), a short sequence required for CRISPR recognition, through the primer rather than relying on its presence in the native target, expanding target accessibility while retaining single-nucleotide discrimination. An ordinary differential equation model captures the threshold behavior and establishes a predictable framework for primer design. Building on this principle, we develop Thermodynamically Encoded Molecular Programming for One-pot diagnostics (TEMPO), which achieves attomolar sensitivity within 30 min and enables sequencing-concordant SNP genotyping and pathogen detection in a single-step microfluidic format.

With the distinctive advantages of high transconductance, low operating voltage, mixed ionic-electronic conductivities, and dynamic versatility, organic electrochemical transistor (OECT) has emerged as a promising wearable technology capable of measuring various biophysical and biochemical signals. Despite the intensive research efforts towards enhancing its wearability, challenges related to signal conversion, voltage sourcing, and manufacturing scalability are seldom addressed. Herein, we report a compact and easy-to-build integrated module that provides stable biasing from batteries while enabling current-to-voltage conversion and additional amplification of OECT responses. Given the known amplitude of target signals, transistor bias and amplification gain can be adjusted easily on site by tuning two key resistance values and ensuring sufficient battery voltage. Furthermore, the flexible OECTs in this work were fabricated through an industrial manufacturing process for flexible printed circuits (FPC), in which the polymeric channel material and device architecture were both customized to accommodate the fabrication constraints. Notably, preliminary measurements based on the battery-powered unit comprising our OECT and module demonstrate significantly amplified bio-signals compared to electrodes. The successful acquisition of on-body electrocardiogram voltages further underscores the potential of this platform to support current and future OECT interfaces. ### Competing Interest Statement The authors have declared no competing interest.

CoRPLA (CRISPR-regulated One-pot Recombinase Polymerase Loop-mediated Amplification) is an amplicon-depleted skin-temperature operated iNAAT designed for at-home testing. It uses specially designed loop primers to enhance isothermal amplification, triggering Cas12 for in-situ amplicon depletion and signal amplification. This method addresses issues like amplicon-derived aerosol contamination and complex assay formats, enabling quantitative detection with sub-attomolar sensitivity (0.5 cps/μL). CoRPLA employs a DNA hydrogel wearable tape for real-time, colorimetric readout, allowing visual differentiation of pathogen loads. It was validated with clinical samples for SARS-CoV-2, RSV, influenza A, and HPV, successfully identifying multi-level viral loads of the positive cases with results consistent with qPCR. Offering high sensitivity while eliminating false positives from aerosol contamination, CoRPLA bridges the molecular assay from benchtop to home for daily viral infections monitoring.

Investigation of the small molecule-aptamer interaction is difficult, and it usually lacks information about the conformational change of aptamers that is important for their application. Here, we present the label-free investigation of small molecule-aptamer interactions using a modularized organic electrochemical transistor (OECT) platform. Leveraging the high sensitivity of the OECT, we measured the conformational change of the aptamer encountering its ligand. This platform consists of a simply fabricated OECT device and a DNA hydrogel-based bioreceptor module. Using this device, we investigated four DNA aptamers binding to cortisol (362.46 Da) and testosterone (288.4 Da). We demonstrated the correlation between the aptamer conformational change and the OECT response. We further calculated the apparent affinities of these aptamers based on the responses. This OECT-based platform also showed potential in binding kinetic measurements. We anticipate that the presented platform could advance the application of the newly identified aptamers as a convenient assay, providing multiple metrics for characterizing small molecule-aptamer interactions.

Effective wound-infection management requires reducing bacterial load and avoiding the formation of bacterial biofilms. In this study, we characterized the structure, composition, and function of mucin extracted from the porcine stomach and demonstrated its pH responsiveness, bacterial load control, and biofilm disruption capabilities. The microstructure of the mucin extract shifts from aggregated to relaxed state and its structure changes from gel to solution as its surrounding pH increases. The deterioration of wounds, characterized, among others, by changes in pH, can be monitored by observing changes in the apparent structural features of the mucin extract, e.g., turbidity. Moreover, the extract displays membrane toxicity against lipid membranes and bacteriostatic effects on both gram-negative and gram-positive bacteria. Notably, the extract also promotes the dispersion of bacterial biofilms after 24 hand 48 h of formation. The biocompatibility, pH responsiveness, and antibiofilm properties of mucin present a novel approach for treating chronic wound infections.

Neurological injuries and disorders have a significant impact on individuals’ quality of life, often resulting in motor and sensory loss. To assess motor performance and monitor neurological disorders, non-invasive techniques such as electroencephalography (EEG) and electromyography (EMG) are commonly used. Traditionally employed wet electrodes with conductive gels are limited by lengthy skin preparation time and allergic reactions. Although dry electrodes and hydrogel-based electrodes can mitigate these issues, their applicability for long-term monitoring is limited. Dry electrodes are susceptible to motion artifacts, whereas hydrogel-based electrodes face challenges related to water-induced instability. Recently, ionogels and eutectogels derived from ionic liquids and deep eutectic solvents have gained immense popularity due to their non-volatility, ionic conductivity, thermal stability, and tunability. Eutectogels, in particular, exhibit superior biocompatibility. These characteristics make them suitable alternatives for the development of safer, robust, and reliable EEG and EMG electrodes. However, research specifically focused on their application for EEG and EMG signal acquisition remains limited. This article explores the electrode requirements and material advancements in EEG and EMG sensing, with a focus on highlighting the benefits that ionogels and eutectogels offer over conventional materials. It sheds light on the current limitations of these materials and proposes areas for further improvement in this field. The potential of these gel-based materials to achieve a seamless interface for high-quality and long-term electrophysiological signal acquisition is emphasized. Leveraging the unique properties of ionogels and eutectogels holds promise for future advancements in EEG and EMG electrode materials, leading to improved monitoring systems and enhanced patient outcomes.

Human islet amyloid polypeptide (hIAPP), an intrinsically disordered protein (IDP), plays a significant role in the pathogenesis of type 2 diabetes through its aggregation. Recent studies have suggested that certain viral protein segments exhibit amyloidogenic potential and may influence its amyloid aggregations associated with pathogenesis. However, the potential link between recurrent SARS-CoV-2 infections and the exacerbation of type 2 diabetes remains poorly understood. In this study, we explore how the amyloidogenic segments of SARS-CoV-2, specifically SK9 and FI10, influence the aggregation of hIAPP and the toxicity of the resulting conformers in a membrane environment. To investigate this, we utilized a range of biophysical techniques, including circular dichroism, nuclear magnetic resonance, atomic force microscopy, dynamic light scattering, fluorescence assays, and cell cytotoxicity assays, complemented by molecular dynamics simulations. Our results indicate that SK9 and FI10 promote hIAPP aggregation in a membrane-mimicking environment, forming distinct aggregate structures. Specifically, SK9 accelerates rapid fibril formation due to inter-chain interactions, while FI10 stabilizes oligomeric aggregates primarily through intra-chain contacts. These results reveal the differential effects of viral protein segments on amyloid formation pathways and aggregate characteristics, providing new insights into the mechanisms of amyloid aggregation for developing better therapeutic strategies against amyloid-associated diseases, particularly diabetes.

The biosafety and sustainability of inorganic perovskites or organic piezopolymers is a major concern in the field of wearable piezoelectric sensors. Naturally occurring, silk fibroin (SF), is a promising alternative for the realization of organic piezoelectric devices due to its excellent biocompatibility and tunable material properties. Nevertheless, its scope for practical sensing applications is limited by the weak innate piezoelectricity of 1 pC N-1. This research aims to improve the piezoelectricity of pristine SF by intrinsically growing non-toxic metal-free perovskite (MFP) to achieve organic and bendable silk fibroin-metal free perovskite (SF-MFP) composite films. The increase in MFP loading has an influence on the resulting morphology and crystallinity of SF-MFP films. In comparison to pristine SF, the films with the highest perovskite loading exhibit an enhanced normal piezoelectric response of 4.6 pm V-1. The poled SF-MFP sensors display an appreciable sensitivity of 0.61 +/- 0.05 V N-1 with a fast response time and outstanding stability arising from a synergistic combination of mechanically robust SF and piezoelectric MFP. Furthermore, these sensors can detect various joint bending and muscle movements in human subjects, proving their suitability for wearable bioelectronics. This research demonstrates the potential of biomaterials and organic perovskites for the development of biocompatible and sustainable wearable piezoelectric solutions.

Polymerase chain reaction (PCR) with probe-based melting curve techniques has been widely used in multiplex genotyping for early diagnosis of genetic diseases and common cancers. However, traditional designs face limitations due to the high modification cost of sensing probes and complex background fluorescence signals. Herein, we introduce the Universal Melting Probes (Uni-Melt) system, a closed-tube PCR assay for multiplex genotyping that incorporates barcoded PCR amplification and universal hybridization probes for melting curve analysis. Uni-Melt can detect multiple genotypes using a single set of universal fluorophore-modified probes and unmodified target-specific mediator strands, which was verified by detecting five different human papillomavirus (HPV) genotypes utilizing two fluorescent channels with the lowest detectable concentration at 102 copies/μL. In addition, Uni-Melt can differentiate multiple mutation sites including single nucleotide polymorphisms (SNPs), as demonstrated in the SLC25A13 gene. Overall, Uni-Melt is a closed-tube, sensitive, and specific system for multiplexed genotyping compatible with commonly employed quantitative polymerase chain reaction (qPCR) thermocyclers. Compared to the traditional probe-based melting curve analysis, Uni-Melt can reduce the assay cost associated with multiple probe modifications and decrease the complexity of the reaction without sacrificing sensitivity and specificity, which makes it a practically useful tool for clinical screening.

This review highlights important biomarkers in blood and dermal fluids, i.e. sweat and interstitial fluid, and envisions non-invasive monitoring of dermal fluids in critical health concerns and drug therapy suitable for remote ambulatory monitoring.

Continuous electrocardiography (ECG) monitoring is a necessity to prevent the sudden occurrence of cardiac events and premature death. Wearable bioelectronics provides the opportunity to realize noninvasive 12-lead ambulatory monitoring for cardiovascular disease (CVD) detection and diagnosis. This work introduces a 12-lead ECG patch that eliminates the issues associated with compliance, skin damage, and user-discomfort encountered in conventional ECG sensors. The patch is safe, stretchable, and skin-adherent with an adaptable design that allows for user-friendly ECG monitoring. These can be attributed to the unique 3-bridge patch structure, embedded liquid metal circuitry, and the wet-adhesive polyacrylamide-polydopamine-tannic acid (PAM-PDA-TA) hydrogel. The hydrogel displays improved wet adhesiveness and biosafety for long-term usage on deforming skin. The patch can acquire clean and consistent ECG signals with a higher signal-to-noise (SNR) ratio compared to commercial electrodes during both static and dynamic monitoring. The outstanding performance of the patch ascertains its potential for daily continuous ECG monitoring and remote diagnosis.

Etaq-PCR, a portable real-time polymerase chain reaction platform, has been developed based on an electroactive hydrolysis probe design and immobilization-free electrochemical readout. The Etaq-PCR platform integrates cost effective commercial instruments and thermally stable cycling into a minimal device enabling real-time monitoring of PCR amplification. No complex electrode surface treatment or micro-fabrication is involved in the construction of the Etaq-PCR platform. Thus, one can easily construct a portable PCR platform via our design. Etaq-PCR achieved sufficient sensitivity, selectivity, and rapidness to meet the WHO ASSURED criteria (affordable, sensitive, specific, user-friendly, rapid, and robust, equipment-free, and deliverable) for point-of-care diagnosis systems. It showed good tolerance to clinical sample matrix as it displayed 100% accuracy in a validation experiment testing a drug resistance gene of nontyphoidal Salmonella in 25 raw clinical isolated samples in comparison with the gold standard qPCR. All in all, Etaq-PCR is a portable, deployable, and cost-effective PCR platform suitable for clinical applications.

Pooling multiple samples prior to real-time reverse-transcription polymerase chain reaction (RT-PCR) analysis has been proposed as a strategy to minimize expenses and boost test throughput during the COVID-19 pandemic. Nevertheless, the traditional pooling approach cannot be effectively deployed in high-prevalence settings due to the need for secondary tests in the case of a positive pool. In this study, we present a pooling test platform with high adaptability and simplicity that allows sample-specific detection of multiple-tagged samples in a single run without the need for retesting. This was accomplished by labeling distinct samples with predefined ID-Primers and identifying tagged pooled samples using one-step RT-PCR followed by melting curve analysis with rationally designed universal fluorescence- and quencher-tagged oligo probes. Using magnetic beads (MBs), nucleic acid targets from different individuals can be tagged and extracted concurrently and then pooled before RT, eliminating the need for extra RNA extraction and separate RT and enzyme digestion steps in the recently developed barcoding strategies. Pools of six samples (positive and negative) were successfully identified by melting temperature values under two fluorescent channels, with a detection sensitivity of 5 copies/μL. We validated the reproducibility of this assay by running it on 40 clinical samples with a hypothetical infection rate of 15%. In addition, to aid the scenario of large-scale pooling tests, we constructed a melting curve autoreadout system (MCARS) for statistical analysis of melting curve plots to eliminate error-prone manual result readout. Our results suggest that this strategy could be a simple and adaptable tool for alleviating existing bottlenecks in diagnostic pooling testing.

Wearable Electronics In article 2300326, I-Ming Hsing and co-workers develop a wearable adhesive 12-lead ECG patch for ambulatory monitoring, with an innovative and user-friendly design that can seamlessly interface with human skin. The unique combination of hydrogel, elastomer, and liquid metal imparts biocompatibility, stretchability, and wet adhesiveness. It can accurately measure stable ECG signals during diverse physical activities by eliminating motion and sweat artifacts.

The necessity for the large-scale screening of viral pathogens has been amply demonstrated during the COVID-19 pandemic. During this time, SARS-CoV-2 nucleic acid pooled testing, such as Dorfman-based group testing, was widely adopted in response to the sudden increased demand for detection. However, the current approach still necessitates the individual retesting of positive pools. Here, we established an efficient method termed the fragment-length identification of pooled nucleic acid samples (FLIPNAS), where all subsamples (n = 8) can be uniquely labelled and tested in a single-time detection among pools of samples. We used a novel and simple design of unique primers (UPs) to generate amplicons of unique lengths after reverse transcription and polymerase chain reaction to reach this aim. As a result, the unique lengths of the amplicons can be recognized and traced back to the corresponding UPs and specific samples. Our results demonstrated that FLIPNAS could recognize one to eight positive subsamples in a single test without retesting positive pools. The system also showed sufficient sensitivity for the mass monitoring of SARS-CoV-2 and no cross-reactivity against three common respiratory diseases. Moreover, the FLIPNAS results of 40 samples with a positive ratio of 7.8% were in 100% agreement with their individual detection results using the gold standard. Collectively, this study shows that the efficiency of nucleic acid pooling detection can be further improved by FLIPNAS, which can speed up testing and mitigate the urgent demand for resources.

Rapid point-of-care diagnostics, essential in settings such as airport on-site testing and home-based screening, displayed important implications for infectious disease control during the SARS-CoV-2 outbreak. However, the deployment of simple and sensitive assays in real-life scenarios still faces the concern of aerosol contamination. Here, we report an amplicon-depleting CRISPR-based one-pot loop-mediated isothermal amplification (CoLAMP) assay for point-of-care diagnosis of SARS-CoV-2 RNA. In this work, AapCas12b sgRNA is designed to recognize the activator sequence sited in the loop region of the LAMP product, which is crucial for exponential amplification. By destroying the aerosol-prone amplifiable products at the end of each amplification reaction, our design can significantly reduce the amplicons contamination that causes false positive results in point-of-care diagnostics. For at-home self-testing, we designed a low-cost sample-to-result device for fluorescence-based visual interpretation. As well, a commercial portable electrochemical platform was deployed as a proof-of-concept of ready-to-use point-of-care diagnostic systems. The field deployable CoLAMP assay can detect as low as 0.5 copies/μL of SARS-CoV-2 RNA in clinical nasopharyngeal swab samples within 40 min without the need for specialists for its operation.

Internal ion-gated organic electrochemical transistor (IGT) demonstrates volume-dependent transconductance with the unprecedented advantages of high speed and self-(de)doping capability among ion-based transistors. The novel characteristics have albeit rendered IGT a promising platform for integrated bioelectronics, its potential in high-frequency applications has yet been fully harnessed. Moreover, a study from a material's point of view is especially needed for this recently emerged platform as the necessity of maintaining the internal ion reservoir has posed difficulties in processing hydrated poly(3,4-ethylenedioxythiophene) doped with poly(styrene sulphonate) (PEDOT:PSS) with electronically favorable morphologies. Herein, a comprehensive investigation of the structural and functional properties of ion-embedded PEDOT:PSS modified by different annealing temperatures is performed and correlated to the IGT performance. A short-time high-temperature annealing treatment is found effective in facilitating the formation of compact microstructures without significantly influencing film hydration. The structural improvement enhances the film's conductivity and hole mobility, with the corresponding IGTs exhibiting higher gain, higher conductance, and high cut-off frequency consistently in a batch. This study also successfully demonstrates the first use of electrochemical transistors like IGTs in high-frequency applications through proof-of-concept experiments simulating fluid estimation in 50 kHz bioimpedance analysis. This work contributes to the development of high-performance IGTs for extensive biological applications.

Wearable devices offer a revolutionary approach to ambulatory electrocardiography (ECG) for cardiovascular disease diagnosis. Herein, an adhesive, highly stretchable and conformal (ASC) patch for long‐term ambulatory ECG monitoring is presented. The ASC patch is designed with a “three‐bridge” structure to provide inhomogeneous strain distribution during stretching. Meanwhile, the electrical stability is achieved by stretchable liquid metal interconnects at the domain experiencing larger strain. Moreover, the long‐term usability can be attained by an adhesive layer made of polydopamine/polyacrylamide glycerol–water hydrogel, maintaining good adhesiveness and stretchability for more than two weeks. Altogether, the patch can successfully measure stable high‐quality ECG signals in relaxed, stretched, or underwater conditions. Enhanced mechanical and electrical properties exhibited by our ASC patch confer superior skin‐device interface on either dry or wet conditions than that of current electrodes. The hydrogel‐based patch displays conformal deformation along with the stretched skin. This work provides a scalable prototype for multilead wearable ECG devices and promises new opportunities for medical applications in soft electronics.