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A path model was utilized to determine the relationship of demographic, family system, adolescent perceptions of parental behavior, and youth characteristics to adolescent substance abuse. Self-report questionnaire data were collected from 214 high school students at two high schools in a southwestern state. Results showed direct positive relationships of adolescent reports of parental substance use and adolescent reports of parental support with adolescent reports of substance use. Indirect relationships were found for school, gender of the adolescent, family hardiness, family coherence and parental substance use to perceived parental support to adolescent substance use. Implications are presented for prevention and intervention at multiple levels of the family system in relation to adolescent substance use. [ABSTRACT FROM AUTHOR]

Data from 77 adolescents in farm and ranch families were used to examine the relationship of demographic variables, family stressor events, and family coping strategies to adolescent adaptation. Results indicated that adolescent age and family transitions were positively related to individual stress. Males reported less family stress than did females. Seeking spiritual support was negatively related to family stress, while the perceived impact of the farm crisis was positively related to family stress. Family support was positively related, and family substance use issues were negatively related, to adolescent satisfaction with family life. The implications of these findings are discussed. [ABSTRACT FROM AUTHOR]

Using self-report questionnaire data from high school students (N = 253), the relation between adolescents' perceptions of family characteristics and adolescent substance use patterns were examined. Results indicated that adolescents' perception of maternal substance use, family hardiness, and age of the adolescent were significant predictors of adolescent substance use. Implications are presented. [ABSTRACT FROM AUTHOR]

The purpose of this study was to examine adolescent perceptions of family system characteristics and parental behaviors as predictors of adolescent substance use. Self-report questionnaire data were collected from a sample of 489 high school students. Bivariate correlations and multiple regression analysis were used to examine the research hypotheses. Results indicated that the frequency of parental substance use was positively related to adolescent substance use, while family bonding and parental support were negatively related. The results provide support for considering both family system characteristics and parenting behaviors in relation to adolescent substance use. [ABSTRACT FROM AUTHOR]

Iron deficiency anemia (IDA) is a condition characterized by lower-than-average iron (Fe) levels in the body, affecting a substantial number of young children and pregnant women globally. Existing diagnostic methods for IDA rely on invasive analysis of stored Fe in ferritin from blood samples, posing challenges, especially for toddlers and young children. To address this issue, saliva has been proposed as a non-invasive sample matrix for IDA diagnosis. However, conventional Fe analysis techniques often necessitate complex and costly instrumentation. This study presents the first non-invasive, saliva-based preliminary screening test for IDA using a nitrocellulose lateral flow system. In this study, we introduce a novel approach using the ferroin reaction with bathophenanthroline (Bphen) and ferrous (Fe 2+ ) ions to quantify Fe levels in saliva. Our methodology involves a capillary flow-driven microfluidic device integrated into a lateral flow system utilizing nitrocellulose membranes. Here, we present the first instance of saliva on a nitrocellulose substrate to detect salivary Fe levels. The optimized system yielded a linear response over the 1-200 ppm range in buffer solution, with a limit of detection (LoD) of 5.6 ppm. Furthermore, the system demonstrated a linear response in pooled saliva samples across the 1-1000 ppm range, with a LoD of 55.1 ppm. These results underscore the potential of our capillary flow-driven microfluidic device as a viable non-invasive diagnostic tool for IDA, particularly in remote and resource-limited settings.

Integrating electrochemistry into capillary-flow driven immunoassay devices provides unique opportunities for quantitative point-of-care testing. Although custom electrodes can be inexpensive and are tunable, they require skilled fabrication. Here, we report the incorporation of a commercial electrode into a capillary-flow driven immunoassay (iceCaDI) device for a single end-user step sandwich electrochemical enzyme-linked immunosorbent assay (ELISA). The iceCaDI device is a pump-free portable microfluidic device with an integrated commercial screen-printed electrode and flow driven by capillary action. The iceCaDI device is composed of alternating polyester transparency film and double-sided adhesive film layers that are patterned with a laser cutter. This platform was designed to address known limitations of laminated device fabrication methods and operation. First, we developed a foldable laminated device fabrication using hinges for easy assembly and precise alignment. Second, reagent dispersing was achieved by incorporating a 1 mm wide arrow-shaped notch in the middle of the channel that trapped an air bubble and formed a baffle that facilitated reagent spreading to cover the detection area. Third, small vent holes were added to the top layer of the channels to prevent air bubbles from blocking flow. Finally, we fabricated a CRP immunosensor with a detection range of 0.625 to 10.0 μg mL -1 as a proof-of-concept to demonstrate an automatically driven sandwich electrochemical ELISA using the iceCaDI device. Three concentrations of CRP were successfully measured under flow conditions within 8 min. Our proposed device is a promising approach and a step forward in the development of point-of-care (POC) devices for techniques that traditionally require multiple user steps.

Multiplexed analysis in medical diagnostics is widely accepted as a more thorough and complete method compared to single-analyte detection. While analytical methods like polymerase chain reaction and enzyme-linked immunosorbent assay (ELISA) exist for multiplexed detection of biomarkers, they remain time-consuming and expensive. Lateral flow assays (LFAs) are an attractive option for point-of-care testing, and examples of multiplexed LFAs exist. However, these devices are limited by spatial resolution of test lines, large sample volume requirements, cross-reactivity, and poor sensitivity. Recent work has developed capillary-flow microfluidic ELISA platforms as a more sensitive alternative to LFAs; however, multiplexed detection on these types of devices has yet to be demonstrated. In the aftermath of the initial SARS-CoV-2 pandemic, the need for rapid, sensitive point-of-care devices has become ever clearer. Moving forward, devices that can distinguish between diseases with similar presenting symptoms would be the ideal home diagnostic. Here, the first example of a multiplexed capillary-flow immunoassay device for the simultaneous detection of multiple biomarkers is reported. From a single sample addition step, the reagents and washing steps required for two simultaneous ELISAs are delivered to spatially separated test strips. Visual results can be obtained in <15 min, and images captured with a smartphone can be analyzed for quantitative data. This device was used to distinguish between and quantify H1N1 hemagglutinin (HA) and SARS-CoV-2 nucleocapsid protein (N-protein). Using this device, analytical detection limits of 840 and 133 pg/mL were obtained for hemagglutinin and nucleocapsid protein, respectively. The presence of one target in the device did not increase the signal on the other test line, indicating no cross-reactivity between the assays. Additionally, simultaneous detection of both N-protein and HA was performed as well as simultaneous detection of N-protein and human C-reactive protein (CRP). Elevated levels of CRP in a patient infected with SARS-CoV-2 have been shown to correlate with more severe outcomes and a greater risk of death as well. To further expand on the simultaneous detection of two biomarkers, CRP and N-protein were detected simultaneously, and the presence of SARS-CoV-2 N-protein did not interfere with the detection of CRP when both targets were present in the sample.

Microfluidic devices have emerged as advantageous tools for detecting environmental contaminants due to their portability, ease of use, cost-effectiveness, and rapid response capabilities. These devices have wide-ranging applications in environmental monitoring of air, water, and soil matrices, and have also been applied to agricultural monitoring. Although several previous reviews have explored microfluidic devices' utility, this paper presents an up-to-date account of the latest advancements in this field for environmental monitoring, looking back at the past five years. In this review, we discuss devices for prominent contaminants such as heavy metals, pesticides, nutrients, microorganisms, per- and polyfluoroalkyl substances (PFAS), etc. We cover numerous detection methods (electrochemical, colorimetric, fluorescent, etc. ) and critically assess the current state of microfluidic devices for environmental monitoring, highlighting both their successes and limitations. Moreover, we propose potential strategies to mitigate these limitations and offer valuable insights into future research and development directions.

An individual's therapeutic drug exposure level is directly linked to corresponding clinical effects. Rapid, sensitive, inexpensive, portable and reliable devices are needed for diagnosis related to drug exposure, treatment, and prognosis of diseases. Electrochemical sensors are useful for drug monitoring due to their high sensitivity and fast response time. Also, they can be combined with portable signal read-out devices for point-of-care applications. In recent years, nanomaterials such as carbon-based, carbon-metal nanocomposites, noble nanomaterials have been widely used to modify electrode surfaces due to their outstanding features including catalytic abilities, conductivity, chemical stability, biocompatibility for development of electrochemical sensors. This review paper presents the most recent advances about nanomaterials-based electrochemical sensors including the use of green assessment approach for detection of drugs including anticancer, antiviral, anti-inflammatory, and antibiotics covering the period from 2019 to 2023. The sensor characteristics such as analyte interactions, fabrication, sensitivity, and selectivity are also discussed. In addition, the current challenges and potential future directions of the field are highlighted., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Heart failure (HF) is an emerging epidemic and remains a major clinical and public health problem. Advances in the healthcare management of HF may lead to lower morbidity and mortality rates but require diagnostics to guide the process. Current diagnostics/prognostics approaches rely on expensive equipment, centralized facilities and trained personnel, marginalizing healthcare access in developing countries and rural communities. These issues have led researchers to focus on developing portable and affordable diagnostics that can be deployed at the point-of-care (POC). Typically, HF biomarkers are measured in blood not saliva. Recently, our team correlated concentrations of salivary Galectin-3 (Gal-3) to outcomes in patients with HF. We have developed an analytical device which consists of an immunoassay based on a screen-printed carbon electrode (SPCE) to quantify Gal-3 levels in saliva samples. Using 10 μL of saliva, the proposed electrochemical immunoassay achieved a concentration dependent signal response in the clinically relevant range with a limit of detection of 9.66 ng/mL. In addition, the storage stability of the modified electrode was investigated, and only a 10.9% loss in current response over a 35-day period. The results of the immunoassay on the modified SPCEs suggest validity as a POC biosensor system for the management of HF., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests.

The field of formulation and stabilization of protein therapeutics has become rather extensive. However, most of the focus has been on stabilization of the final drug product. Yet, proteins experience stress and degradation through the manufacturing process, starting with fermentaition. This review describes how formulation principles can be applied to stabilize biopharmaceutical proteins during bioprocessing and manufacturing, considering each unit operation involved in prepration of the drug substance. In addition, the impact of the container on stabilty is discussed as well., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 American Pharmacists Association. Published by Elsevier Inc. All rights reserved.)

Cu(I)-catalyzed 1,3-dipolar cycloaddition (CuAAC), also known as click chemistry, has been demonstrated to be highly robust while providing versatile surface chemistry. One specific application is biosensor fabrication. Recently, we developed thermoplastic electrodes (TPEs) as an alternative to traditional carbon composite electrodes in terms of cost, performance, and robustness. However, their applications in biosensing are currently limited due to a lack of facile methods for electrode modification. Here, we demonstrate the feasibility of using CuAAC following the diazonium grafting of TPEs to take advantage of two powerful technologies for developing a customizable and versatile biosensing platform. After a stepwise characterization of the electrode modification procedures was performed, electrodes were modified with model affinity reagents. Streptavidin and streptavidin-conjugated IgG antibodies were successfully immobilized on the TPE surface, as confirmed by electrochemical impedance spectroscopy and X-ray photoelectron spectroscopy.

Over the last few years, the SARS-CoV-2 pandemic has made the need for rapid, affordable diagnostics more compelling than ever. While traditional laboratory diagnostics like PCR and well-plate ELISA are sensitive and specific, they can be costly and take hours to complete. Diagnostic tests that can be used at the point-of-care or at home, like lateral flow assays (LFAs) are a simple, rapid alternative, but many commercially available LFAs have been criticized for their lack of sensitivity compared to laboratory methods like well-plate ELISAs. The Capillary-Driven Immunoassay (CaDI) device described in this work uses microfluidic channels and capillary action to passively automate the steps of a traditional well-plate ELISA for visual read out. This work builds on prior capillary-flow devices by further simplifying operation and use of colorimetric detection. Upon adding sample, an enzyme-conjugated secondary antibody, wash steps, and substrate are sequentially delivered to test and control lines on a nitrocellulose strip generating a colorimetric response. The end user can visually detect SARS-CoV-2 antigen in 15-20 min by naked eye, or results can be quantified using a smartphone and software such as ImageJ. An analytical detection limit of 83 PFU/mL for SARS-CoV-2 was determined for virus in buffer, and 222 PFU/mL for virus spiked into nasal swabs using image analysis, similar to the LODs determined by traditional well-plate ELISA. Additionally, a visual detection limit of 100 PFU/mL was determined in contrived nasal swab samples by polling 20 untrained end-users. While the CaDI device was used for detecting clinically relevant levels of SARS-CoV-2 in this study, the CaDI device can be easily adapted to other immunoassay applications by changing the reagents and antibodies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

This work focuses on a systematic method to produce Ag, Cu, and Ag/Cu metallic nanoparticles (MNPs) in situ assisted with ultrasound on cellulose paper. By tuning the concentration of AgNO 3 and CuSO 4 salt precursors and ultrasound time, combined with a fixed concentration of ascorbic acid (AA) as a reducing agent, it was possible to control the size, morphology, and polydispersity of the resulting MNPs on cellulose papers. Notably, high yield and low polydispersity of MNPs and bimetallic nanoparticles are achieved by increasing the sonication time on paper samples pre-treated with salt precursors before reduction with AA. Moreover, mechanical analysis on paper samples presenting well-dispersed and distributed MNPs showed slightly decreasing values of Young's modulus compared to neat papers. The strain at break is substantially improved in papers containing solely Ag or Cu MNPs. The latter suggests that the elastic/plastic transition and deformation of papers are tuned by cellulose and MNPs interfacial interaction, as indicated by mechanical analysis. The proposed method provides insights into each factor affecting the sonochemistry in situ synthesis of MNPs on cellulose papers. In addition, it offers a straightforward alternative to scale up the production of MNPs on paper, ensuring an eco-friendly method., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

To protect the body from external pathogens, the intestines have sophisticated epithelial and mucosal barriers. Disruptions to barrier integrity are associated with a variety of disorders such as irritable bowel disease, Crohn's disease, and celiac disease. One critical component of all barriers are collagens in the extracellular matrix. While the importance of the intestinal barrier is established, current models lack the ability to represent the complex biology that occurs at these barriers. For the current study a microfluidic device model was modified to determine the effectiveness of collagen breakdown to cause barrier disruption. Bacterial collagenase was added for 48 h to the luminal channel of a dual flow microfluidic device to examine changes in intestinal barrier integrity. Tissues exhibited dose-dependent alterations in immunoreactive collagen-1 and claudin-1, and coincident disruption of the epithelial monolayer barrier as indicated by goblet cell morphologies. This ex vivo model system offers promise for further studies exploring factors that affect gut barrier integrity and potential downstream consequences that cannot be studied in current models.

The human microbiome influences the efficacy and safety of a wide variety of commonly prescribed drugs. Designing precision medicine approaches that incorporate microbial metabolism would require strain- and molecule-resolved, scalable computational modeling. Here, we extend our previous resource of genome-scale metabolic reconstructions of human gut microorganisms with a greatly expanded version. AGORA2 (assembly of gut organisms through reconstruction and analysis, version 2) accounts for 7,302 strains, includes strain-resolved drug degradation and biotransformation capabilities for 98 drugs, and was extensively curated based on comparative genomics and literature searches. The microbial reconstructions performed very well against three independently assembled experimental datasets with an accuracy of 0.72 to 0.84, surpassing other reconstruction resources and predicted known microbial drug transformations with an accuracy of 0.81. We demonstrate that AGORA2 enables personalized, strain-resolved modeling by predicting the drug conversion potential of the gut microbiomes from 616 patients with colorectal cancer and controls, which greatly varied between individuals and correlated with age, sex, body mass index and disease stages. AGORA2 serves as a knowledge base for the human microbiome and paves the way to personalized, predictive analysis of host-microbiome metabolic interactions., (© 2023. The Author(s).)

Thermoplastic carbon electrodes (TPEs) are an alternative form of carbon composite electrodes that have shown excellent electrochemical performance with applications in biological sensing. However, little has been done to apply TPEs to environmental sensing, specifically heavy metal analysis. The work here focuses on lead analysis and based on their electrochemical properties, TPEs are expected to outperform other carbon composite materials; however, despite testing multiple formulations, TPEs showed inferior performance. Detailed electrode characterization was conducted to examine the cause for poor lead sensing behavior. X-Ray photoelectron spectroscopy (XPS) was used to analyze the surface functional groups, indicating that acidic and alkaline functional groups impact lead electrodeposition. Further, scanning electron microscopy (SEM) and electrochemical characterization demonstrated that both the binder and graphite can influence the surface morphology, electroactive area, and electron kinetics.

Motivation: The importance and rate of development of genome-scale metabolic models have been growing for the last few years, increasing the demand for software solutions that automate several steps of this process. However, since TRIAGE's release, software development for the automatic integration of transport reactions into models has stalled., Results: Here, we present the Transport Systems Tracker (TranSyT). Unlike other transport systems annotation software, TranSyT does not rely on manual curation to expand its internal database, which is derived from highly curated records retrieved from the Transporters Classification Database and complemented with information from other data sources. TranSyT compiles information regarding transporter families and proteins, and derives reactions into its internal database, making it available for rapid annotation of complete genomes. All transport reactions have GPR associations and can be exported with identifiers from four different metabolite databases. TranSyT is currently available as a plugin for merlin v4.0 and an app for KBase., Availability and Implementation: TranSyT web service: https://transyt.bio.di.uminho.pt/; GitHub for the tool: https://github.com/BioSystemsUM/transyt; GitHub with examples and instructions to run TranSyT: https://github.com/ecunha1996/transyt&#95;paper., (Published by Oxford University Press 2023.)

Tonic immobility is a passive antipredator strategy employed late in the predation sequence that may decrease individual mortality in prey animals. Here, we investigate how energetic state and genetic predisposition influence antipredator decision-making in green lacewing larvae, Chrysoperla plorabunda (Fitch), using simulated predatory encounters. We demonstrate that tonic immobility is a plastic response influenced by energetic resource limitation. Larvae exposed to 1 or 2 days of food deprivation initiate tonic immobility more often and with less physical provocation than individuals fed ad libitum. Recently molted individuals exposed to food deprivation, the individuals most energetically challenged, engage in tonic immobility at a higher rate than any other group. We also find that variation in antipredator strategy between individuals is partly the result of within-population genetic variation. We estimate the propensity to enter tonic immobility to have a broad-sense heritability of 0.502. Taken together our results suggest that larval lacewings under energetic stress are more likely to engage in tonic immobility. Yet, energetic state does not explain all within-population variation, as individuals can have a genetic predisposition for tonic immobility., (© The Author(s) 2023. Published by Oxford University Press on behalf of Entomological Society of America.)

A capillary-driven microfluidic sequential flow device, designed for eventual at-home or doctor's office use, was developed to perform an enzyme-linked immunosorbent assay (ELISA) for serology assays. Serology assays that detect SARS-CoV-2 antibodies can be used to determine prior infection, immunity status, and/or individual vaccination status and are typically run using well-plate ELISAs in centralized laboratories, but in this format SARs-CoV-2 serology tests are too expensive and/or slow for most situations. Instead, a point-of-need device that can be used at home or in doctor's offices for COVID-19 serology testing would provide critical information for managing infections and determining immune status. Lateral flow assays are common and easy to use, but lack the sensitivity needed to reliably detect SARS-CoV-2 antibodies in clinical samples. This work describes a microfluidic sequential flow device that is as simple to use as a lateral flow assay, but as sensitive as a well-plate ELISA through sequential delivery of reagents to the detection area using only capillary flow. The device utilizes a network of microfluidic channels made of transparency film and double-sided adhesive combined with paper pumps to drive flow. The geometry of the channels and storage pads enables automated sequential washing and reagent addition steps with two simple end-user steps. An enzyme label and colorimetric substrate produce an amplified, visible signal for increased sensitivity, while the integrated washing steps decrease false positives and increase reproducibility. Naked-eye detection can be used for qualitative results or a smartphone camera for quantitative analysis. The device detected antibodies at 2.8 ng mL -1 from whole blood, while a well-plate ELISA using the same capture and detection antibodies could detect 1.2 ng mL -1 . The performance of the capillary-driven immunoassay (CaDI) system developed here was confirmed by demonstrating SARS-CoV-2 antibody detection, and we believe that the device represents a fundamental step forward in equipment-free point-of-care technology.

The potential reach of point-of-care (POC) diagnostics into daily routines for exposure to reactive oxygen species (ROS) and Cu in aerosolized particulate matter (PM) demands that microfluidic paper-based analytical devices (μPADs) take into consideration the simple detection of these toxic PM components. Here, we propose μPADs with a dual-detection system for simultaneous ROS and Cu(II) detection. For colorimetric ROS detection, the glutathione (GSH) assay with a folding design to delay the reaction yielded complete ROS and GSH oxidation, and improved homogeneity of color development relative to using the lateral flow pattern. For electrochemical Cu(II) determination, 1,10-phenanthroline/Nafion modified graphene screen-printed electrodes showed ability to detect Cu(II) down to pg level being low enough to be applied to PM analysis. No intra- and inter-interference affecting both systems were found. The proposed μPADs obtained LODs for 1,4-naphthoquinone (1,4-NQ), used as the ROS representative, and Cu(II) of 8.3 ng and 3.6 pg, respectively and linear working ranges of 20 to 500 ng for ROS and 1 × 10 -2 to 2 × 10 2 ng for Cu(II). Recovery of the method was between 81.4 and 108.3% for ROS and 80.5-105.3% for Cu(II). Finally, the sensors were utilized for simultaneous ROS and Cu(II) determination in PM samples and the results statistically agreed with those using the conventional methods at 95% confidence., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

A significant bottleneck exists for mass-production of ion-selective electrodes despite recent developments in manufacturing technologies. Here, we present a fully-automated system for large-scale production of ISEs. Three materials, including polyvinyl chloride, polyethylene terephthalate and polyimide, were used as substrates for fabricating ion-selective electrodes (ISEs) using stencil printing, screen-printing and laser engraving, respectively. We compared sensitivities of the ISEs to determine the best material for the fabrication process of the ISEs. The electrode surfaces were modified with various carbon nanomaterials including multi-walled carbon nanotubes, graphene, carbon black, and their mixed suspensions as the intermediate layer to enhance sensitivities of the electrodes. An automated 3D-printed robot was used for the drop-cast procedure during ISE fabrication to eliminate manual steps. The sensor array was optimized, and the detection limits were 10 -5  M, 10 -5  M and 10 -4  M for detection of K + , Na + and Ca 2+ ions, respectively. The sensor array integrated with a portable wireless potentiometer was used to detect K + , Na + and Ca 2+ in real urine and simulated sweat samples and results obtained were in agreement with ICP-OES with good recoveries. The developed sensing platform offers low-cost detection of electrolytes for point-of-care applications., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Human exposure to heavy metals is a concerning global problem because of its detrimental effect on our health and ecosystem. Assessing the levels of these metals is cost- and labor-intensive and nonuser friendly because current analysis approaches typically rely on heavy instrumentations like inductively coupled plasma-mass spectrometry, which is only possible in centralized labs. Hence, simple economical detection methods are in high demand in developing countries and areas with insufficient infrastructure, professional experts, and appropriate environmental treatment. Several microfluidic paper-based analytical devices have been reported as promising alternatives to conventional testing methods for on-site heavy metal detection. Paper-based microfluidics are advantageous because of their simple fabrication, biodegradability, low cost, and ability to operate without pumps. However, typical assay times for current platforms are slow, and they typically rely on pipetting a fixed volume into the assay cards. This adds complexity in actual field scenarios. Here, we report a novel, inexpensive, and straightforward capillary-driven microfluidic device combined with paper for rapid and user-friendly detection of Ni(II), Cu(II), and Fe(III) in water. A colorimetric approach was adopted to quantify these metals. The device was able to produce a homogeneous color signal within 8 s of sample insertion. The limit of detection and limit of quantification were calculated to be 2 and 6.67 ppm for nickel, 0.3 and 1 ppm for Cu, and 1.1 and 3.67 ppm for Fe, respectively. The majority (>90%) of the collected samples showed recovery in the 80-110% range with acceptable accuracy and precision (<15% RSD) for a colorimetric device. This technique can be beneficial for rapidly assessing heavy metal exposure in drinking and surface water at drastically reduced assay time and is the first example of capillary flow-driven microfluidic devices as a transport medium for heavy metal detection.

Microbial genome annotation is the process of identifying structural and functional elements in DNA sequences and subsequently attaching biological information to those elements. DRAM is a tool developed to annotate bacterial, archaeal, and viral genomes derived from pure cultures or metagenomes. DRAM goes beyond traditional annotation tools by distilling multiple gene annotations to genome level summaries of functional potential. Despite these benefits, a downside of DRAM is the requirement of large computational resources, which limits its accessibility. Further, it did not integrate with downstream metabolic modeling tools that require genome annotation. To alleviate these constraints, DRAM and the viral counterpart, DRAM-v, are now available and integrated with the freely accessible KBase cyberinfrastructure. With kb&#95;DRAM users can generate DRAM annotations and functional summaries from microbial or viral genomes in a point-and-click interface, as well as generate genome-scale metabolic models from DRAM annotations., Availability and Implementation: For kb&#95;DRAM users, the kb&#95;DRAM apps on KBase can be found in the catalog at https://narrative.kbase.us/#catalog/modules/kb&#95;DRAM. For kb&#95;DRAM users, a tutorial workflow with all documentation is available at https://narrative.kbase.us/narrative/129480. For kb&#95;DRAM developers, software is available at https://github.com/shafferm/kb&#95;DRAM., (© The Author(s) 2023. Published by Oxford University Press.)

Competing Interests: Conflict of interest statement. None declared.

Rapid species radiations provide insight into the process of speciation and diversification. The radiation of Chrysoperla carnea-group lacewings seems to be driven, at least in part, by their species-specific pre-mating vibrational duets. We associated genetic markers from across the genome with courtship song period in the offspring of a laboratory cross between Chrysoperla plorabunda and Chrysoperla adamsi, two species primarily differentiated by their mating songs. Two genomic regions were strongly associated with the song period phenotype. Large regions of chromosomes one and two were associated with song phenotype, as fewer recombination events occurred on these chromosomes relative to the other autosomes. Candidate genes were identified by functional annotation of proteins from the C. carnea reference genome. The majority of genes that are associated with vibrational courtship signals in other insects were found within QTL for lacewing song phenotype. Together these findings suggest that decreased recombination may be acting to keep together loci important to reproductive isolation between these species. Using wild-caught individuals from both species, we identified signals of genomic divergence across the genome. We identified several candidate genes both in song-associated regions and near divergence outliers including nonA, fruitless, paralytic, period, and doublesex. Together these findings bring us one step closer to identifying the genomic basis of a mating song trait critical to the maintenance of species boundaries in green lacewings., (© 2022 The Authors. Insect Molecular Biology published by John Wiley & Sons Ltd on behalf of Royal Entomological Society.)

3D printing technologies are an attractive for fabricating electrochemical sensors due to their ease of operation, freedom of design, fast prototyping, low waste, and low cost. We report the fabrication of a simple 3D-printed electrochemical sensing device for non-enzymatic detection of creatinine, an important indicator of renal function. To create the 3D-printed electrodes (3DE), carbon black/polylactic acid (CB/PLA) composite filament was used. The 3DE was activated using 0.5 M NaOH via amperometry prior to use to improve electrochemical performance. To give selectivity for creatinine, the activated 3DE was modified with a copper oxide nanoparticle-ionic liquid/reduced graphene oxide (CuO-IL/rGO) composite. The modified 3DE was characterized using microscopy and electrochemistry. Cyclic voltammetry and amperometry were used to evaluate sensor performance. The modified 3DE provided electrocatalytic activity towards creatinine without enzymes. Under optimal conditions, the modified 3DE directly coupled with a portable smartphone potentiostat exhibited the linear detection range of 0.5-35.0 mM, and the limit of detection was 37.3 μM, which is sufficient for detecting creatinine in human urine samples. Furthermore, the other physiological compounds present in human urine were not detected on the modified 3DE. Therefore, the modified 3DE could be a tool for effective creatinine screening in the urine., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The COVID-19 pandemic has significantly increased the development of the development of point-of-care (POC) diagnostic tools because they can serve as useful tools for detecting and controlling spread of the disease. Most current methods require sophisticated laboratory instruments and specialists to provide reliable, cost-effective, specific, and sensitive POC testing for COVID-19 diagnosis. Here, a smartphone-assisted Sensit Smart potentiostat (PalmSens) was integrated with a paper-based electrochemical sensor to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). A disposable paper-based device was fabricated, and the working electrode directly modified with a pyrrolidinyl peptide nucleic acid (acpcPNA) as the biological recognition element to capture the target complementary DNA (cDNA). In the presence of the target cDNA, hybridization with acpcPNA probe blocks the redox conversion of a redox reporter, leading to a decrease in electrochemical response correlating to SARS-CoV-2 concentration. Under optimal conditions, a linear range from 0.1 to 200 nM and a detection limit of 1.0 pM were obtained. The PNA-based electrochemical paper-based analytical device (PNA-based ePAD) offers high specificity toward SARS-CoV-2 N gene because of the highly selective PNA-DNA binding. The developed sensor was used for amplification-free SARS-CoV-2 detection in 10 nasopharyngeal swab samples (7 SARS-CoV-2 positive and 3 SARS-CoV-2 negative), giving a 100% agreement result with RT-PCR., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

There has been a long-standing interest in point-of-care (POC) diagnostics as a tool to improve patient care because it can provide rapid, actionable results near the patient. Some of the successful examples of POC testing include lateral flow assays, urine dipsticks, and glucometers. Unfortunately, POC analysis is somewhat limited by the ability to manufacture simple devices to selectively measure disease specific biomarkers and the need for invasive biological sampling. Next generation POCs are being developed that make use of microfluidic devices to detect biomarkers in biological fluids in a non-invasive manner, addressing the above-mentioned limitations. Microfluidic devices are desirable because they can provide the ability to perform additional sample processing steps not available in existing commercial diagnostics. As a result, they can provide more sensitive and selective analysis. While most POC methods make use of blood or urine as a sample matrix, there has been a growing push to use saliva as a diagnostic medium. Saliva represents an ideal non-invasive biofluid for detecting biomarkers because it is readily available in large quantities and analyte levels reflect those in blood. However, using saliva in microfluidic devices for POC diagnostics is a relatively new and an emerging field. The overarching aim of this review is to provide an update on recent literature focused on the use of saliva as a biological sample matrix in microfluidic devices. We will first cover the characteristics of saliva as a sample medium and then review microfluidic devices that are developed for the analysis of salivary biomarkers., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Uncultivated Bacteria and Archaea account for the vast majority of species on Earth, but obtaining their genomes directly from the environment, using shotgun sequencing, has only become possible recently. To realize the hope of capturing Earth's microbial genetic complement and to facilitate the investigation of the functional roles of specific lineages in a given ecosystem, technologies that accelerate the recovery of high-quality genomes are necessary. We present a series of analysis steps and data products for the extraction of high-quality metagenome-assembled genomes (MAGs) from microbiomes using the U.S. Department of Energy Systems Biology Knowledgebase (KBase) platform ( http://www.kbase.us/ ). Overall, these steps take about a day to obtain extracted genomes when starting from smaller environmental shotgun read libraries, or up to about a week from larger libraries. In KBase, the process is end-to-end, allowing a user to go from the initial sequencing reads all the way through to MAGs, which can then be analyzed with other KBase capabilities such as phylogenetic placement, functional assignment, metabolic modeling, pangenome functional profiling, RNA-Seq and others. While portions of such capabilities are available individually from other resources, the combination of the intuitive usability, data interoperability and integration of tools in a freely available computational resource makes KBase a powerful platform for obtaining MAGs from microbiomes. While this workflow offers tools for each of the key steps in the genome extraction process, it also provides a scaffold that can be easily extended with additional MAG recovery and analysis tools, via the KBase software development kit (SDK)., (© 2022. Springer Nature Limited.)

While food safety issues are attracting public concern due to their detrimental effects on human health, monitoring livestock health is urgently needed to diagnose animal diseases at an early stage by applying proper treatments, controlling, and preventing outbreaks, particularly in resource- limited countries. In addition, unhealthy farms are not only a threat to livestock but also to human lives. The available diagnostic techniques for the detection of key health threats within both the food and livestock sectors require labor-intensive and time-consuming experimental procedures and sophisticated and expensive instruments. To tackle this issue, optical biosensing strategies have been incorporated into point-of-care (POC) systems, offering real-time monitoring, field-deployable, and low-cost devices, which help make on-the-spot decisions. This review aims to discuss the recent cutting-edge research on POC optical biosensing platforms for on-farm diagnosis of animal diseases and on-site detection of animal-derived food-borne contaminants, including pathogens, antibiotics, and mycotoxins. Moreover, this review briefly presents the basic knowledge of various types of optical biosensors and their development using various recent strategies, including nanomaterial combinations, to enhance their performance in POC tests. This review is expected to help scientists to understand the evolution and challenges in the development of point-of-care biosensors for the food and livestock industry, benefiting global healthcare., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Small, single-layer microfluidic paper-based analytical devices (µPADs) offer potential for a range of point-of-care applications; however, they have been limited to low flow rates. Here, we investigate the role of laser cutting paper channels in maximizing flow rate in small profile devices with limited fluid volumes. We demonstrate that branching, laser-cut grooves can provide a 59.23-73.98% improvement in flow rate over a single cut, and a 435% increase over paper alone. These design considerations can be applied to more complex microfluidic devices with the aim of increasing the flow rate, and could be used in stand-alone channels for self-pumping., Supplementary Information: The online version contains supplementary material available at 10.1007/s10404-023-02679-8., Competing Interests: Conflict of interestThere are no financial or non-financial conflicts to declare., (© The Author(s) 2023.)

The COVID-19 pandemic focused attention on a pressing need for fast, accurate, and low-cost diagnostic tests. This work presents an electrochemical capillary driven immunoassay (eCaDI) developed to detect SARS-CoV-2 nucleocapsid (N) protein. The low-cost flow device is made of polyethylene terephthalate (PET) and adhesive films. Upon addition of a sample, reagents and washes are sequentially delivered to an integrated screen-printed carbon electrode for detection, thus automating a full sandwich immunoassay with a single end-user step. The modified electrodes are sensitive and selective for SARS-CoV-2 N protein and stable for over 7 weeks. The eCaDI was tested with influenza A and Sindbis virus and proved to be selective. The eCaDI was also successfully applied to detect nine different SARS-CoV-2 variants, including Omicron., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

Here, we developed a microfluidic paper device by combining ion-selective electrodes (ISE) and a distance-based paper device (dPAD) for simultaneous potentiometric and colorimetric detection of urine electrolytes including K + , Na + and Cl - . The working and reference electrode zones were coated with polystyrene as a non-ionic polymer to improve hydrophobic properties on the paper surface for fabrication of K + -ISE and Na + -ISE. The layer of polymer coating was optimized to enhance the sensitivity of the ISEs. Under optimized conditions, the electrode surfaces were modified with carbon black to improve the electrochemical characteristics of the ISEs. The ISEs showed good performance with sensitivities of 54.14 ± 3.94 mV per decade and 55.08 ± 1.15 mV per decade for K + and Na + within the linear concentration range 0.100 mM-100 mM K + and 5 mM-1 M Na + , respectively. The limits of detection (LOD) were 0.05 mM and 1.36 mM for K + and Na + , respectively. The linear working range of Cl - was 0.50 to 50 mM and the LOD and limit of quantification (LOQ) were found to be 0.16 ± 0.05 mM (3SD) and 0.53 ± 0.05 mM (10SD), respectively. The dual-mode ISE-dPAD was validated in human urine and recoveries were obtained as 90-108%, 94-105%, and 90-96% for K + , Na + , and Cl - , respectively, showing successful application of the developed device in a complex matrix. The ISE-dPAD has advantages including low-cost ($ 0.33 per test), eco-friendly, portability, simple operation, the need of low sample volume (100 μL), and simultaneous analysis on a single device.

Biogenic amines are an important and widely studied class of molecules due to their link to the physiological processes of food-related illnesses and histamine poisoning. Electrochemiluminescent (ECL) detection offers an inexpensive and portable analytical method of detection for biogenic amines when coupled with recent advancements in low-cost carbon-based electrodes and a smartphone camera. In this work, a mobile phone camera was evaluated against a piece of conventional instrumentation, the charge-coupled device, for the detection of ECL from the reaction of biogenic amines with the luminescent compound tris(2,2'-bipyridyl)ruthenium(II). Assisted by a 3D-printed light-tight housing, the mobile phone achieved limits of detection of 127, 425 and 421 μM for spermidine, putrescine, and histamine, respectively. The mobile phone's analytical figures of merit were lesser than the CCD camera but were still within the range to detect contamination. In an exploration of real-world samples, the mobile phone was able to determine the contents of amines in skim milk on par with that of a CCD camera.

At home electrolyte analysis is of growing interest due to the importance for early diagnosis for various diseases. This work presents the first device that integrates a potentiometric ion-selective electrode (ISE) with distance-based colorimetric detection for the simultaneous analysis of K + and Cl - ions at the point of care (POC). This hybrid sensing device was designed in a two-dimensional configuration using a plastic transparency sheet containing a stencil printed K + -ISE and a paper substrate for distance-based colorimetric detection of Cl - . K + quantification was performed using a low-cost (<$25) lab-made Wi-Fi supported potentiometer with a custom smartphone application while Cl - ions were quantified with an instrument-free distance-based paper analytical device (dPAD). The total analysis time is 5 min once the sample is added. The K + -ISE showed a sensitivity of 55.89 ± 2.52 mV/decade from 0.1 to 100 mM with a limit of detection (LOD) of 0.01 mM. The linear working range for Cl - detection was 0.5-50 mM whereas the LOD was 0.16 ± 0.05 mM. The proof-of-concept application of the sensing hybrid device was demonstrated in human urine and artificial sweat samples containing K + and Cl - ions at physiologically relevant ranges. The recoveries were found to be 88-108% for K + and 90-104% for Cl - , showing the potential use of the proposed device for low-cost routine analysis of biological samples at POC., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

We report automated fabrication of solid-contact sodium-selective (Na + -ISEs) and potassium-selective electrodes (K + -ISEs) using a 3D printed liquid handling robot controlled with Internet of Things (IoT) technology. The printing system is affordable and can be customized for the use with micropipettes for applications such as drop-casting, biological assays, sample preparation, rinsing, cell culture, and online analyte monitoring using multi-well plates. The robot is more compact (25 × 30 × 35 cm) and user-friendly than commercially available systems and does not require mechatronic experience. For fabrication of ion-selective electrodes, a carbon black intermediate layer and ion-selective membrane were successively drop-cast on the surface of stencil-printed carbon electrode using the dispensing robot. The 3D-printed robot increased ISE robustness while decreasing the modification time by eliminating manual steps. The Na + -ISEs and K + -ISEs were characterized for their potentiometric responses using a custom-made, low-cost (<$25) multi-channel smartphone-based potentiometer capable of signal processing and wireless data transmission. The electrodes showed Nernstian responses of 58.2 ± 2.6 mV decade -1 and 56.1 ± 0.7 mV decade -1 for Na + and K + , respectively with an LOD of 1.0 × 10 -5  M. We successfully applied the ISEs for multiplexed detection of Na + and K + in urine and artificial sweat samples at clinically relevant concentration ranges. The 3D-printed pipetting robot cost $100 and will pave the way for more accessible mass production of ISEs for those who cannot afford the expensive commercial robots., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Analysis of the genes retained in the minimized Mycoplasma JCVI-Syn3A genome established that systems that repair or preempt metabolite damage are essential to life. Several genes known to have such functions were identified and experimentally validated, including 5-formyltetrahydrofolate cycloligase, coenzyme A (CoA) disulfide reductase, and certain hydrolases. Furthermore, we discovered that an enigmatic YqeK hydrolase domain fused to NadD has a novel proofreading function in NAD synthesis and could double as a MutT-like sanitizing enzyme for the nucleotide pool. Finally, we combined metabolomics and cheminformatics approaches to extend the core metabolic map of JCVI-Syn3A to include promiscuous enzymatic reactions and spontaneous side reactions. This extension revealed that several key metabolite damage control systems remain to be identified in JCVI-Syn3A, such as that for methylglyoxal. IMPORTANCE Metabolite damage and repair mechanisms are being increasingly recognized. We present here compelling genetic and biochemical evidence for the universal importance of these mechanisms by demonstrating that stripping a genome down to its barest essentials leaves metabolite damage control systems in place. Furthermore, our metabolomic and cheminformatic results point to the existence of a network of metabolite damage and damage control reactions that extends far beyond the corners of it that have been characterized so far. In sum, there can be little room left to doubt that metabolite damage and the systems that counter it are mainstream metabolic processes that cannot be separated from life itself.

Eight-hundred thousand to one trillion prokaryotic species may inhabit our planet. Yet, fewer than two-hundred thousand prokaryotic species have been described. This uncharted fraction of microbial diversity, and its undisclosed coding potential, is known as the "microbial dark matter" (MDM). Next-generation sequencing has allowed to collect a massive amount of genome sequence data, leading to unprecedented advances in the field of genomics. Still, harnessing new functional information from the genomes of uncultured prokaryotes is often limited by standard classification methods. These methods often rely on sequence similarity searches against reference genomes from cultured species. This hinders the discovery of unique genetic elements that are missing from the cultivated realm. It also contributes to the accumulation of prokaryotic gene products of unknown function among public sequence data repositories, highlighting the need for new approaches for sequencing data analysis and classification. Increasing evidence indicates that these proteins of unknown function might be a treasure trove of biotechnological potential. Here, we outline the challenges, opportunities, and the potential hidden within the functional dark matter (FDM) of prokaryotes. We also discuss the pitfalls surrounding molecular and computational approaches currently used to probe these uncharted waters, and discuss future opportunities for research and applications., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Authors. Published by Elsevier B.V.)

Environmental challenges and development require plants to reallocate resources between primary and specialized metabolites to survive. Genome-scale metabolic models, which map carbon flux through metabolic pathways, are a valuable tool in the study of tradeoffs that arise at this interface. Due to annotation gaps, models that characterize all the enzymatic steps in individual specialized pathways and their linkages to each other and to central carbon metabolism are difficult to construct. Recent studies have successfully curated subsystems of specialized metabolism and characterized the interfaces where flux is diverted to the precursors of glucosinolates, terpenes, and anthocyanins. Although advances in metabolite profiling can help to constrain models at this interface, quantitative analysis remains challenging because of the different timescales on which specialized metabolites from constitutive and reactive pathways accumulate., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this article., (Copyright © 2022. Published by Elsevier Ltd.)

It has been shown that short-chain fatty acids (SCFAs) produced by the gut microbiome are of importance to host tissue health; however, measuring such compounds in biological samples is often limited to using hours to days old fecal and blood plasma samples. Organ-on-a-chip models have been created to simplify the complexity but struggle to reproduce the full biology of the gut specifically. We recently reported a tissue-in-a-chip gut model that incorporates gut explanted tissue into a microfluidic device. The system maintains a biologically relevant oxygen gradient and tissue ex vivo for days at a time, but minimal characterization of biological activity was reported. Herein, we use 1 H-NMR to analyze the SCFA content of tissue media effluents from gut explants cultured in the recently developed microfluidic organotypic device (MOD). 1 H-NMR can identify key SCFAs in the complex samples with minimal sample preparation. Our findings show that maintaining physiologically relevant oxygen conditions, something often missing from many other culture systems, significantly impacts the SCFA profile. Additionally, we noted the changes in SCFAs with culture time and potential variability between SCFA levels in male and female mouse tissue explants cultured in the MOD system based on 1 H-NMR spectral profiles.

Urinary tract infections (UTIs) are one of the most common infections across the world and can lead to serious complications such as sepsis if not treated in a timely manner. Uropathogenic Escherichia coli account for 75% of all UTIs. Early diagnosis is crucial to help control UTIs, but current culturing methods are expensive and time-consuming and lack sensitivity. The existing point-of-care methods fall short because they rely on indirect detection from elevated nitrates in urine rather than detecting the actual bacteria causing the infection. Magnetophoresis is a powerful method used to separate and/or isolate cells of interest from complex matrices for analysis. However, magnetophoresis typically requires complex and expensive instrumentation to control flow in microfluidic devices. Coupling magnetophoresis with microfluidic paper-based analytical devices (μPADs) enables pump-free flow control and simple and low-cost operation. Early magnetophoresis μPADs showed detection limits competitive with traditional methods but higher than targets for clinical use. Here, we demonstrate magnetophoresis using hybrid μPADs that rely on capillary action in hydrophilic polyethylene terephthalate channels combined with paper pumps. We were able to detect E. coli with a calculated limit of detection of 2.40 × 10 2 colony-forming units per mL.

Among lateral flow immunoassay (LFIA) platforms, enzyme-based LFIAs provide signal amplification to improve sensitivity. However, most enzyme-based LFIAs require multiple timed steps, complicating their utility in point-of-care testing (POCT). Here, we report a microfluidic interface for LFIAs that automates sample, buffer, and reagent addition, greatly simplifying operation while achieving the high analytical stringency associated with more complex assays. The microfluidic interface also maintains the low cost and small footprint of standard LFIAs. The platform is fabricated from a combination of polyester film, double-sided adhesive tape, and nitrocellulose, and fits in the palm of your hand. All reagents are dried on the nitrocellulose to facilitate sequential reagent delivery, and the sample is used as the wash buffer to minimize steps. After the sample addition, a user simply waits 15 min for a colorimetric result. This manuscript discusses the development and optimization of the channel geometry to achieve a simple step enzyme amplified immunoassay. As a proof-of-concept target, we selected lipoarabinomannan (LAM), a WHO identified urinary biomarker of active tuberculosis, to demonstrate the device feasibility and reliability. The results revealed that the device successfully detected LAM in phosphate buffer (PBS) as well as spiked urine samples within 15 min after sample loading. The minimum concentration of color change was achieved at 25 ng mL -1 .

All nine presently known European species of the Apertochrysa prasina group (formerly the Pseudomallada prasinus group) are diagnosed. Separate keys for the identification of living and preserved specimens of each sex are presented. For males, the furwing trait (increased numbers of setae on costal crossveins) is often crucial for identification, especially in preserved specimens. In some cases, females of the newly recognized prasinoid species (Ap1, Ap2, Ap3) are impossible to distinguish without live coloration. This is largely why the three new species are difficult to assign to one of the many old (often female) type specimens of former prasinoid species or varieties, which had been synonymized with Chrysopa prasina in the past.

A microfluidic paper-based thermoplastic electrode (TPE) array has been developed for point-of-care detection of Na + and K + ions using a custom-made portable potentiometer. TPEs were fabricated using polystyrene as the binder and two different types of graphite to compare the electrode performance. The newly designed TPE array embedded in a polymethyl methacrylate chip consists of two working electrodes modified with carbon black nanomaterial and an ion-selective membrane, and an all-solid-state reference electrode modified with Ag/AgCl ink and poly(butyl methacrylate-co-methyl methacrylate) membrane via drop-casting. Ion-selective membrane compositions and conditioning steps were optimized. Under optimized conditions, ion-selective TPEs demonstrated fast response time (4 s) and good stability. The TPE array demonstrated a Nernstian behavior for K + with a sensitivity of 59.2 ± 0.2 mV decade -1 and near-Nernstian response for Na + with a sensitivity of 54.0 ± 1.1 mV decade -1 in the range 10 -1  - 10 -4  M and 1 -  10 -3  M, respectively. The detection limits were 1 × 10 -5  M and 1 × 10 -4  M for K + and Na + , respectively. In addition, a K + and Na + selective microfluidic paper-based analytical device (µPAD) was applied to artificial serum analysis and found in good agreement with average recoveries of 101.3% and 99.7%, respectively, suggesting that the developed ISE array is suitable for detection of sodium and potassium in complex matrix., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Point-of-care (POC) methods currently available for detecting SARS-CoV-2 infections still lack accuracy. Here, we report the development of a highly sensitive electrochemical immunoassay capable of quantitatively detecting the presence of the SARS-CoV-2 virus in patient nasopharyngeal samples using stencil-printed carbon electrodes (SPCEs) functionalized with capture antibodies targeting the SARS-CoV-2 nucleocapsid protein (N protein). Samples are added to the electrode surface, followed by horseradish peroxidase (HRP)-conjugated detection antibodies also targeting the SARS-CoV-2 N protein. The concentration of the virus in samples is quantified using chronoamperometry in the presence of 3,3'5,5'-tetramethylbenzidine. Limits of detection equivalent to less than 50 plaque forming units/mL (PFU/mL) were determined with virus sample volumes of 20 μL. No cross-reactivity was detected with the influenza virus and other coronavirus N proteins. Patient nasopharyngeal samples were tested as part of a proof-of-concept clinical study where samples were also tested using the gold-standard real-time quantitative polymerase chain reaction (RT-qPCR) method. Preliminary results from a data set of 22 samples demonstrated a clinical specificity of 100% ( n = 9 negative samples according to RT-qPCR) and a clinical sensitivity of 70% for samples with RT-PCR cycle threshold (Ct) values under 30 ( n = 10) and 100% for samples with Ct values under 25 ( n = 5), which complies with the World Health Organization (WHO) criteria for POC COVID-19 diagnostic tests. Our functionalized SPCEs were also validated against standard plaque assays, and very good agreement was found between both methods ( R 2 = 0.9993, n = 6), suggesting that our assay could be used to assess patient infectivity. The assay currently takes 70 min from sampling to results.