An integrated magnetic separation enzyme-linked colorimetric sensing platform for field detection of Escherichia coli O157: H7 in food.

An intelligent colorimetric sensing platform integrated with in situ immunomagnetic separation function was developed for ultrasensitive detection of Escherichia coli O157: H7 (E. coli O157: H7) in food. Captured antibody modified magnetic nanoparticles (cMNPs) and detection antibody/horseradish peroxidase (HRP) co-functionalized AuNPs (dHAuNPs) were firstly synthesized for targeted enrichment and colorimetric assay of E. coli O157: H7, in which remarkable signal amplification was realized by loading large amounts of HRP on the surface of AuNPs. Coupling with the optical collimation attachments and embedded magnetic separation module, a highly integrated optical device was constructed, by which in situ magnetic separation and high-quality imaging of 96-well microplates containing E. coli O157: H7 was achieved with a smartphone. The concentration of E. coli O157: H7 could be achieved in one-step by performing digital image colorimetric analysis of the obtained image with a custom-designed app. This biosensor possesses high sensitivity (1.63 CFU/mL), short detecting time (3 h), and good anti-interference performance even in real-sample testing. Overall, the developed method is expected to be a novel field detection platform for foodborne pathogens in water and food as well as for the diagnosis of infections due to its portability, ease of operation, and high feasibility.

Oriented Antibody Coupling to an Antifouling Polymer Using Glycan Remodeling for Biosensing by Particle Motion.

Biosensors based on immobilized antibodies require molecular strategies that (i) couple the antibodies in a stable fashion while maintaining the conformation and functionality, (ii) give outward orientation of the paratope regions of the antibodies for good accessibility to analyte molecules in the biofluid, and (iii) surround the antibodies by antibiofouling molecules. Here, we demonstrate a method to achieve oriented coupling of antibodies to an antifouling poly(l-lysine)-grafted-poly(ethylene glycol) (PLL-g-PEG) substrate, using glycan remodeling to create antibody-DNA conjugates. The coupling, orientation, and functionality of the antibodies were studied using two analysis methods with single-molecule resolution, namely single-molecule localization microscopy and continuous biosensing by particle motion. The biosensing functionality of the glycan-remodeled antibodies was demonstrated in a sandwich immunosensor for procalcitonin. The results show that glycan-remodeled antibodies enable oriented immobilization and biosensing functionality with low nonspecific binding on antifouling polymer substrates.

All-in-one platform: Versatile, Easy, and User-friendly System (VEUS) based on automated and expert-independent antibody immobilization and immunoassay by utilizing customized movement of magnetic particles.

The ELISA is the most worldwide method for immunoassay. However, the ELISA is losing ground due to low reproducibility of manual experimental processes in both R&D and IVD areas. An automated platform is a good solution, but there are still limitations owning to extremely high cost and requiring large space to set up especially for a small size laboratory. Here, we present a novel all-in-one platform called "VEUS" settable on the laboratory table that offers comprehensive automation of the entire multiplex immunoassay process by exploiting antibody conjugated magnetic particles, quality control and then immunoanalytical reaction, thereby enhancing detection sensitivity and high reproducibility. As a proof of concept, the system exhibits a sensitive LOD of 0.6 and 3.1 pg mL-1 within 1 h run, comparable precision that of molecular diagnostic systems based on PCR method, enabling rapid multiplex diagnosis of Influenza A, Influenza B, and COVID-19 viruses with similar symptoms. Through automation by the all-in-one system, it can be used by novice users, something innovative for immunoassays, relying heavily on user experience. Furthermore, it can contribute to streamline entire immunoassay processes of diverse biomarkers with high reproducibility and convenience in laboratories.

An immobilized antibody-based affinity grid strategy for on-grid purification of target proteins enables high-resolution cryo-EM.

In cryo-electron microscopy (cryo-EM), sample preparation poses a critical bottleneck, particularly for rare or fragile macromolecular assemblies and those suffering from denaturation and particle orientation distribution issues related to air-water interface. In this study, we develop and characterize an immobilized antibody-based affinity grid (IAAG) strategy based on the high-affinity PA tag/NZ-1 antibody epitope tag system. We employ Pyr-NHS as a linker to immobilize NZ-1 Fab on the graphene oxide or carbon-covered grid surface. Our results demonstrate that the IAAG grid effectively enriches PA-tagged target proteins and overcomes preferred orientation issues. Furthermore, we demonstrate the utility of our IAAG strategy for on-grid purification of low-abundance target complexes from cell lysates, enabling atomic resolution cryo-EM. This approach greatly streamlines the purification process, reduces the need for large quantities of biological samples, and addresses common challenges encountered in cryo-EM sample preparation. Collectively, our IAAG strategy provides an efficient and robust means for combined sample purification and vitrification, feasible for high-resolution cryo-EM. This approach holds potential for broader applicability in both cryo-EM and cryo-electron tomography (cryo-ET).

Versatile conductometric biosensors for rapid and selective detection of inflammatory and cardiac biomarkers in saliva.

Biosensors have become promising alternatives to the conventional methods in early identification of diseases. However, translation of biosensors from lab to commercial products have challenges such as complex sensor fabrications and complicated detection, and inadequate sensitivity and selectivity. Here, we introduce simple and low-cost fabricated conductometric sensors based on high resistivity silicon wafers (HR-Si) which can be adopted to functionalise with both natural and synthetic antibodies in detecting five biomarkers including interleukin-6, C reactive protein, cardiac troponin I, brain natriuretic peptide, and N terminal-probrain natriuretic peptide. All five biomarkers show selective and rapid (10 min sample incubation and <1 min of reading time) detection in both media of phosphate buffer saline and saliva with the detection limits lower than that of reported healthy levels in saliva. This work highlights the versatility of HR-Si sensors in functionalisation of both natural and synthetic antibodies in sensitive and selective biomarker detection. As these miniaturised conductometric biosensors can be easily modified with on-demand biomaterials to detect corresponding target biomarkers, they enable a new category of compact point-of-care medical devices.

Development of a label-free photoelectrochemical immunosensor for novel astrovirus detection.

Since 2017, an infectious goose gout disease characterized by urate precipitation in viscera, mainly caused by novel goose astrovirus (GoAstV) infection, has emerged in the main goose-producing region of China. The current challenge in managing goose gout disease is largely due to the absence of a rapid and efficient detection method for the GoAstV pathogen. Notably, the potential application of immunosensors in detecting GoAstV has not yet been explored. Herein, a label-free PEC immunosensor was fabricated by using purchased TiO2 as the photoactive material and antibody against GoAstV P2 proteins as the specific recognition element. First, we successfully expressed the capsid spike domain P2 protein of ORF2 from GoAstV CHSH01 by using the pET prokaryotic expression system. Meanwhile, the polyclonal antibody against GoAstV capsid P2 protein was produced by purified protein. To our knowledge, this is the first establishment and preliminary application of the label-free photoelectrochemical immunosensor method in the detection of AstV. The PEC immunosensor had a linear range of 1.83 fg mL-1 to 3.02 ng mL-1, and the limit of detection (LOD) was as low as 0.61 fg mL-1. This immunosensor exhibited high sensitivity, great specificity, and good stability in detecting GoAstV P2 proteins. To evaluate the practical application of the immunosensor in real-world sample detection, allantoic fluid from goose embryos was collected as test samples. The results indicated that of the eight positive samples, one false negative result was detected, while both negative samples were accurately detected, suggesting that the constructed PEC immunosensor had good applicability and practical application value, providing a platform for the qualitative detection of GoAstV.

Ultrasensitive electrochemical immunosensor for the detection of C-reactive protein antigen.

Cardiovascular disease is one of the leading causes of premature death worldwide, and the determination of C-reactive protein (CRP) from human serum is of vital importance for the diagnosis of the disease. For this study, we have developed an electrochemical immunosensor based on onion-like carbon@polyacrylonitrile (OLC-PAN) for the detection of CRP antigens. This was accomplished by immobilizing CRP antibodies on a modified glassy carbon electrode (GCE). Several electrochemical techniques such as cyclic voltammetry (CV), square wave voltammetry (SWV), and electrochemical impedance spectroscopy (EIS) were employed to evaluate the electrochemical detection of the CRP antigen. This ultrasensitive method for CRP antigen detection exhibited a very good logarithmic plot from -4.52 to -12.05 g mL-1 and a limit of detection (LOD) of 0.9 fg mL-1. The high selectivity, sensitivity, and stability of the developed electrochemical immunosensor would facilitate miniaturization for point-of-care applications and the efficient diagnosis of cardiovascular diseases.

A sandwich-type photoelectrochemical biosensor based on Ru(bpy)32+ sensitized In2S3 for aflatoxin B1 detection.

Aflatoxin B1 (AFB1), classified as a class I carcinogen, is a widespread mycotoxin that poses a serious threat to public health and economic development, and the food safety problems caused by AFB1 have aroused worldwide concern. The development of accurate and sensitive methods for the detection of AFB1 is significant for food safety monitoring. In this work, a sandwich-type photoelectrochemical (PEC) biosensor for AFB1 detection was constructed on the basis of an aptamer-antibody structure. A good photocurrent response was obtained due to the sensitization of In2S3 by Ru(bpy)32+. In addition, this sandwich-type sensor constructed by modification with the antibody, target detector, and aptamer layer by layer attenuated the migration hindering effect of photogenerated carriers caused by the double antibody structure. The aptamer and antibody synergistically recognized and captured the target analyte, resulting in more reliable PEC response signals. CdSe@CdS QDs-Apt were modified as a signal-off probe onto the sensor platform to quantitatively detect AFB1 with a "signal-off" response, which enhanced the sensitivity of the sensor. The PEC biosensor showed a linear response range from 10-12 to 10-6 g mL-1 with a detection limit of 0.023 pg mL-1, providing a feasible approach for the quantitative detection of AFB1 in food samples.

Rapid and High-Density Antibody Immobilization Using Electropolymerization of Pyrrole for Highly Sensitive Immunoassay.

Polypyrrole (Ppy) is a biologically compatible polymer that is used as a matrix, in which drugs and enzymes can be incorporated by doping. Here, we suggest an inventive application of Ppy as a biorecognition film encapsulated with an antibody (Ab) as an alternative strategy for the on-site multistep functionalization of thiol-based self-assembled monolayers. The fabrication steps of the recognition films were followed by dropping pyrrole and Ab mixed solutions onto the electrode and obtaining a thin film by direct current electropolymerization. The efficiency of Ab immobilization was studied by using fluorescence microscopy and electrochemical (EC) methods. Finally, the Ab density was increased and immobilized in 1 min, and the sensing performance as an EC immunosensor was demonstrated using α-fetoprotein with a limit of detection of 3.13 pg/mL and sensing range from 1 pg/mL to 100 ng/mL. This study demonstrates the potential for electrochemical functionalization of biomolecules with high affinity and rapidity.

A graphene oxide-assisted protein immobilization paper-tip immunosensor with smartphone and naked eye readout for the detection of okadaic acid.

BACKGROUND: Okadaic acid (OA), as a diarrhetic shellfish poisoning, can increase the risk of acute carcinogenic or teratogenic effects for the ingestion of OA contaminated shellfish. At present, much effort has been made to graft immunoassay onto a paper substrate to make paper-based sensors for rapid and simple detection of shellfish toxin. However, the complicated washing steps and low protein fixation efficiency on the paper substrate need to be further addressed. RESULTS: A novel paper-tip immunosensor for detecting OA was developed combined with smartphone and naked eye readout. The trapezoid paper tip was consisted of quantitative and qualitative detection zones. To improve the OA antigen immobilization efficiency on the paper substrate, graphene oxide (GO)-assisted protein immobilization method was introduced. Meanwhile, Au nanoparticles composite probe combined with the lateral flow washing was developed to simplify the washing step. The OA antigen-immobilized zone, as the detection zone Ⅰ, was used for quantitative assay by smartphone imaging. The paper-tip front, as the detection zone Ⅱ, which could qualitatively differentiate OA pollution level within 45 min using the naked eye. The competitive immunoassay on the paper tip exhibited a wide linear range for detecting OA (0.02-50 ng∙mL-1) with low detection limit of 0.02 ng∙mL-1. The recovery of OA in spiked shellfish samples was in the range of 90.3 %-113.%. SIGNIFICANCE: These results demonstrated that the proposed paper-tip immunosensor could provide a simple, low-cost and high-sensitivity test for OA detection without the need for additional large-scale equipment or expertise. We anticipate that this paper-tip immunosensor will be a flexible and versatile tool for on-site detecting the pollution of marine products.

Laser-printed paper ELISA and hydroxyapatite immobilization for colorimetric congenital anomalies screening in saliva.

BACKGROUND: Alpha-fetoprotein (AFP) is a fetal protein that can indicate congenital anomalies such as Down syndrome and spinal canal blockage when detected at abnormal levels in pregnant women. Current AFP detection methods rely on invasive blood or serum samples, which require sophisticated equipment. From the many solutions proposed, colorimetric paper-based assays excel in point-of-care settings. The concept of paper-based ELISA (p-ELISA) enhances traditional methods, aligning with the ASSURED criteria for diagnostics in resource-limited regions. Despite success in microfluidic paper-based assay devices, laser printing remains underexplored for p-ELISA. Additionally, modifying the paper surface provides an additional layer of sensitivity enhancement. RESULTS: In this study, we developed a novel laser-printed paper-based ELISA (LP-pELISA) for rapid, sensitive, and noninvasive detection of AFP in saliva samples. The LP-pELISA platform was fabricated by printing hydrophobic barriers on filter paper using a laser printer, followed by depositing hydroxyapatite (HAp) as an immobilization material for the antibodies. The colorimetric detection was achieved using AuNPs functionalized with anti-AFP antibodies and silver nitrate enhancement. The LP-pELISA exhibited a linear response for AFP detection in both buffer and saliva samples over a range of 1.0-800 ng mL-1, with a limit of detection (LOD) reaching 1.0 ng mL-1. The assay also demonstrated good selectivity, repeatability, reproducibility, and stability. The LP-pELISA was further validated by testing spiked human saliva samples, showing its potential for point-of-care diagnosis of congenital disabilities. SIGNIFICANCE: The LP-pELISA is a noninvasive platform showcasing simplicity, cost-effectiveness, and user-friendliness, utilizing laser printing, hydroxyapatite modification, and saliva samples to efficiently detect AFP. Beyond its application for AFP, this method's versatility extends to other biomarkers, positioning it as a catalyst for the evolution of paper-based biosensors. The LP-pELISA holds promise as a transformative tool for point-of-care diagnostics, fostering advancements in healthcare with its innovative technology.

Versatile Metal-Organic Framework Incorporating Ag2S for Constructing a Photoelectrochemical Immunosensor for Two Breast Cancer Markers.

Breast cancer poses the significance of early diagnosis and treatment. Here, we developed an innovative photoelectrochemical (PEC) immunosensor characterized by high-level dual photocurrent signals and exceptional sensitivity. The PEC sensor, denoted as MIL&Ag2S, was constructed by incorporating Ag2S into a metal-organic framework of MIL-101(Cr). This composite not only enhanced electron-hole separation and conductivity but also yielded robust and stable dual photocurrent signals. Through the implementation of signal switching, we achieved the combined detection of cancer antigen 15-3 (CA15-3) and carcinoembryonic antigen (CEA) with outstanding stability, reproducibility, and specificity. The results revealed a linear range for CEA detection spanning 0.01-32 ng/mL, with a remarkably low detection limit of 0.0023 ng/mL. Similarly, for CA15-3 detection, the linear range extended from 0.1 to 320 U/mL, with a low detection limit of 0.014 U/mL. The proposed strategy introduces new avenues for the development of highly efficient, cost-effective, and user-friendly PEC sensors. Furthermore, it holds promising prospects for early clinical diagnosis, contributing to potential breakthroughs in medical detection and ultimately improving patient outcomes.

Nitrocellulose/acrylic resin coated screen-printed carbon electrode to construct a capacitive immunosensor for anti-BSA.

The capacitive immunosensor, known for its label-free simplicity, has great potential for point-of-care diagnostics. However, the interaction between insulation and recognition layers on the sensing electrode greatly affects its performance. This study introduces a pioneering dual-layer strategy, implementing a novel combination of acrylic resin (AR) and nitrocellulose (NC) coatings on screen-printed carbon electrodes (SPCEs). This innovative approach not only enhances the dielectric properties of the capacitive sensor but also streamlines the immobilization of recognizing elements. Particularly noteworthy is the superior reliability and insulation offered by the AR coating, surpassing the limitations of traditional self-assembled monolayer (SAM) modifications. This dual-layer methodology establishes a robust foundation for constructing capacitive sensors optimized specifically for liquid medium-based biosensing applications. The NC coating in this study represents a breakthrough in effectively immobilizing BSA, unraveling the capacitive response intricately linked to the quantity of adsorbed recognizing elements. The results underscore the prowess of the proposed immunosensor, showcasing a meticulously defined linear calibration curve for anti-BSA (ranging from 0 to 25 µg/ml). Additionally, specific interactions with anti-HAS and anti-TNF-α further validate the versatility and efficacy of the developed immunosensor. This work presents a streamlined and highly efficient protocol for developing label-free immunosensors for antibody determination and introduces a paradigm shift by utilizing readily available electrodes and sensing systems. The findings are poised to catalyze a significant acceleration in the advancement of biosensor technology, opening new avenues for innovative applications in point-of-care diagnostics.

Preparation of immunomagnetic composite nanostructures with bifunctional four-arm PEG derivatives as linkers for the ultrafast enrichment of zearalenone and its metabolites.

It is challenging to prepare sample pretreatment materials with simple use, strong selectivity and satisfactory enrichment performance. In this study, the antibody (3D4) that can specifically recognize zearalenone (ZEN) and its metabolites was immobilized on the surface of gold-coated magnetic Fe3O4 nanoparticles (GMN) by streptavidin (SA)-biotin interaction using GMN as the substrate and our designed four-arm PEG derivative (HS-4ARMPEG10K-(CM)3) as the linker. The immunomagnetic nanoparticles (GMN-4ARMPEG10K-SA-3D4) prepared by this strategy can achieve rapid enrichment (only 5 min) of analytes directly in the matrix, and higher enrichment capacity compared with the previous immunomagnetic particles. The sensitive and accurate analysis of ZEN and its metabolites can be achieved coupled with HPLC-MS/MS. The LODs and LOQs were 0.02-0.05 µg/kg and 0.05-0.10 µg/kg, respectively. The recoveries were 84.13%-112.67%, and the RSDs were 1.09%-9.39%. The method can provide a powerful tool for highly sensitive and rapid monitoring of mycotoxins in complex matrices due to its' strong selectivity and resistance to matrix interference.

Bilateral efforts to improve SERS detection efficiency of exosomes by Au/Na7PMo11O39 Combined with Phospholipid Epitope Imprinting.

Detection of cancer-related exosomes in body fluids has become a revolutionary strategy for early cancer diagnosis and prognosis prediction. We have developed a two-step targeting detection method, termed PS-MIPs-NELISA SERS, for rapid and highly sensitive exosomes detection. In the first step, a phospholipid polar site imprinting strategy was employed using magnetic PS-MIPs (phospholipids-molecularly imprinted polymers) to selectively isolate and enrich all exosomes from urine samples. In the second step, a nanozyme-linked immunosorbent assay (NELISA) technique was utilized. We constructed Au/Na7PMo11O39 nanoparticles (NPs) with both surface-enhanced Raman scattering (SERS) property and peroxidase catalytic activity, followed by the immobilization of CD9 antibodies on the surface of Au/Na7PMo11O39 NPs. The Au/Na7PMo11O39-CD9 antibody complexes were then used to recognize CD9 proteins on the surface of exosomes enriched by magnetic PS-MIPs. Lastly, the high sensitivity detection of exosomes was achieved indirectly via the SERS activity and peroxidase-like activity of Au/Na7PMo11O39 NPs. The quantity of exosomes in urine samples from pancreatic cancer patients obtained by the PS-MIPs-NELISA SERS technique showed a linear relationship with the SERS intensity in the range of 6.21 × 107-2.81 × 108 particles/mL, with a limit of detection (LOD) of 5.82 × 107 particles/mL. The SERS signal intensity of exosomes in urine samples from pancreatic cancer patients was higher than that of healthy volunteers. This bidirectional MIPs-NELISA-SERS approach enables noninvasive, highly sensitive, and rapid detection of cancer, facilitating the monitoring of disease progression during treatment and opening up a new avenue for rapid early cancer screening.

Exploring various carbon nanomaterials-based electrodes modified with polymelamine for the reagentless electrochemical immunosensing of Claudin18.2.

Claudin18.2 (CLDN18.2) is a tight junction protein often overexpressed in various solid tumors, including gastrointestinal and esophageal cancers, serving as a promising target and potential biomarker for tumor diagnosis, treatment assessment, and prognosis. Despite its significance, no biosensor has been reported to date for the detection of CLDN18.2. Here, we present the inaugural immunosensor for CLDN18.2. In this study, an amine-rich conducting polymer of polymelamine (PM) was electrografted onto different carbon nanomaterial-based screen-printed electrodes (SPEs), including carbon (C), graphene (Gr), graphene oxide (GO), carbon nanotube (CNT), and carbon nanofiber (CNF) via cyclic voltammetry. A comparative study was performed to explore the best material for the preparation of the PM-modified electrodes to be used as in-situ redox substrate for the immunosensor fabrication. The surface chemistry and structural features of pristine and PM-deposited electrodes were analyzed using Raman and scanning electron microscopy (SEM) techniques. Our results showed that the PM deposited on Gr and CNT/SPEs exhibited the most significant and stable redox behavior in PBS buffer. The terminal amine moieties on the PM-modified electrode surfaces were utilized for immobilizing anti-CLDN18.2 monoclonal antibodies via N-ethyl-N'-(3-(dimethylamino)propyl)carbodiimide/N-hydroxysuccinimide chemistry to construct the electrochemical immunosensor platform. Differential pulse voltammetry-based immunosensing of CLDN18.2 protein on BSA/anti-CLDN18.2/PM-Gr/SPE and BSA/anti-CLDN18.2/PM-CNT/SPE exhibited excellent selectivity against other proteins such as CD1, PDCD1, and ErBb2. The limits of detection of these two immunosensor platforms were calculated to be 7.9 pg/mL and 0.104 ng/mL for the CNT and Gr immunosensors, respectively. This study demonstrated that the PM-modified Gr and CNT electrodes offer promising platforms not only for the reagentless signaling but also for covalent immobilization of biomolecules. Moreover, these platforms offer excellent sensitivity and selectivity for the detection of CLDN18.2 due to its enhanced stable redox activity. The immunosensor demonstrated promising results for the sensitive detection of CLDN18.2 in biological samples, addressing the critical need for early gastric cancer diagnosis.

The Evaluation of a Lateral Flow Strip Based on the Covalently Fixed "End-On" Orientation of an Antibody for Listeria monocytogenes Detection.

In this study, the covalently fixed "end-on" orientation of a monoclonal Listeria monocytogenes antibody (mAb-Lis) to amino terminated oligo (ethylene glycol)-capped gold nanoparticles (NH2-TEG-AuNPs) was used to fabricate an in-house lateral flow strip (LFS), namely, the fixed "end-on" Lis-mAb-NH-TEG-AuNPs LFS. The aim was to evaluate the performance of the fixed "end-on" Lis-mAb-NH-TEG-AuNPs LFS in detecting L. monocytogenes. The proposed LFS enabled the sensitive detection of L. monocytogenes in 15 min with a visual limit of detection of 102 CFU/mL. Quantitative analysis indicated an LOD at 10 CFU/mL. The fixed "end-on" Lis-mAb-NH-TEG-AuNPs LFS showed no cross-reactivity with other pathogenic bacteria and practical performance across different food matrices, including human blood, milk, and mushroom samples. Furthermore, the clinical performance of the fixed "end-on" Lis-mAb-NH-TEG-AuNPs LFS for detecting L. monocytogenes was evaluated by using 12 clinical samples validated by the hemoculture method. It demonstrated excellent concordance with the reference methods, with no false-positive or false-negative results observed. Therefore, the fixed "end-on" Lis-mAb-NH-TEG-AuNPs LFS serves as a promising candidate for a point-of-care test (POCT), enabling the rapid, precise, and highly sensitive detection of L. monocytogenes in clinical samples and contaminated food.

Nanobody-based immunosensor for the detection of H. pylori in saliva.

Helicobacter pylori (H. pylori) infection is highly prevalent worldwide, affecting more than 43% of world population. The infection can be transmitted through different routes, like oral-oral, fecal-oral, and gastric-oral. Electrochemical sensors play a crucial role in the early detection of various substances, including biomolecules. In this study, the development of nanobody (Nb)-based immunosensor for the detection of H. pylori antigens in saliva samples was investigated. The D2_Nb was isolated and characterized using Western blot and ELISA and employed in the fabrication of the immunosensor. The sensor was prepared using gold screen-printed electrodes, with the immobilization of Nb achieved through chemical linkage using cysteamine-glutaraldehyde. The surface of the electrode was characterized using EIS, FTIR and SEM. Initially, the Nb-based immunosensor's performance was evaluated through cyclic voltammetry (CV), differential pulse voltammetry (DPV), and square wave voltammetry (SWV). The sensor exhibited excellent linearity with an R2 value of 0.96. However, further assessment with the DPV technique revealed both a low limit of detection (5.9 ng/mL, <1 cfu/mL) and high selectivity when exposed to a mixture of similar antigens. Moreover, the immunosensor demonstrated robust recovery rates (96.2%-103.4%) when spiked into artificial saliva and maintained its functionality when stored at room temperature for 24 days.

Point-of-Care Electrochemical Immunosensor Applied against Nosocomial Infection: Staphylococcus aureus Detection in Human Hand Skin.

Staphylococcus aureus is an important pathogen that causes nosocomial infections, resulting in unacceptable morbidity and mortality rates. In this work, we proposed the construction of a nanostructured ZnO-based electrochemical immunosensor for qualitative and semiquantitative detection of S. aureus using simple methods for growing zinc oxide nanorods (ZnO NRs) on a sensor board and immobilizing the anti-S. aureus antibody on ZnO NRs through cystamine and glutaraldehyde. The immunosensor detected S. aureus in the 103-107 colony-forming unit (CFU) mL-1 range and showed a limit of detection (LoD) around 0.792 × 103 CFU mL-1. Beyond a satisfactory LoD, the developed immunosensor presented other advantages, such as high versatility for point-of-care assays and a suitable selective factor that admits the detection of the S. aureus concentration range in human hand skin after washing. Moreover, the immunosensor showed the potential to be an excellent device to control nosocomial infection by detecting the presence of S. aureus in human hand skin.

A disposable immunosensor array using cellulose paper assembled chemiresistive biosensor for simultaneous monitoring of mycotoxins AFB1 and FB1.

Due to the common contamination of multiple mycotoxins in food, which results in stronger toxicity, it is particularly important to simultaneously test for various mycotoxins for the protection of human health. In this study, a disposable immunosensor array with low-cost was designed and fabricated using cellulose paper, polydimethylsiloxane (PDMS), and semiconducting single-walled carbon nanotubes (s-SWCNTs), which was modified with specific antibodies for mycotoxins AFB1 and FB1 detection. The strategy for fabricating the immunosensor array with two individual channels involved a two-step protocol starting with the form of two kinds of carbon films by depositing single-wall carbon nanotubes (SWCNTs) and s-SWCNTs on the cellulose paper as the conductive wire and sensing element, followed by the assembly of chemiresistive biosensor with SWCNTs strip as the wire and s-SWCNTs as the sensing element. After immobilizing AFB1-bovine serum albumin (AFB1-BSA) and FB1-bovine serum albumin (FB1-BSA) separately on the different sensing regions, the formation of mycotoxin-BSA-antibody immunocomplexes transfers to electrochemical signal, which would change with the different concentrations of free mycotoxins. Under optimal conditions, the immunosensor array achieved a limit of detection (LOD) of 0.46 pg/mL for AFB1 and 0.34 pg/mL for FB1 within a wide dynamic range from 1 pg/mL to 20 ng/mL. Furthermore, the AFB1 and FB1 spiked in the ground corn and wheat extracts were detected with satisfactory recoveries, demonstrating the excellent practicality of this established method for simultaneous detection of mycotoxins.