Nanoflower Microreactor Based Versatile Enhancer for Recognition Cofactor-Dependent Enzyme Biocatalysis toward Saxitoxin Detection.

Investigating organic carriers' utilization efficiency and bioactivity within organic-inorganic hybrid nanoflowers is critical to constructing sensitive immunosensors. Nevertheless, the sensitivity of immunosensors is interactively regulated by different classes of biomolecules such as antibodies and enzymes. In this work, we introduced a new alkaline phosphatase-antibody-CaHPO4 hybrid nanoflowers (AAHNFs) microreactor based colorimetric immunoprobe. This system integrates a biometric unit (antibody) with a signal amplification element (enzyme) through the biomineralization process. Specifically, the critical factors affecting antibody recognition activity in the formation mechanism of AAHNFs are investigated. The designed AAHNFs retain antibody recognition ability with enhanced protection for encapsulated proteins against high temperature, organic solvents, and long-term storage, facilitating the selective construction of lock structures against antigens. Additionally, a colorimetric immunosensor based on AAHNFs was developed. After ascorbic acid 2-phosphate hydrolysis by alkaline phosphatase (ALP), the generated ascorbic acid decomposes I2 to I-, inducing the localized surface plasmon resonance in the silver nanoplate, which is effectively tuned through shape conversion to develop the sensor. Further, a 3D-printed portable device is fabricated, integrated with a smartphone sensing platform, and applied to the data of collection and analysis. Notably, the immunosensor exhibits improved analytical performance with a 0.1-6.25 ng·mL-1 detection range and a 0.06 ng·mL-1 detection limit for quantitative saxitoxin (STX) analysis. The average recoveries of STX in real samples ranged from 85.9% to 105.9%. This study presents a more in-depth investigation of the recognition element performance, providing insights for improved antibody performance in practical applications.

Epstein-Barr Viruses: Their Immune Evasion Strategies and Implications for Autoimmune Diseases.

Epstein-Barr virus (EBV), a member of the γ-herpesvirus family, is one of the most prevalent and persistent human viruses, infecting up to 90% of the adult population globally. EBV's life cycle includes primary infection, latency, and lytic reactivation, with the virus primarily infecting B cells and epithelial cells. This virus has evolved sophisticated strategies to evade both innate and adaptive immune responses, thereby maintaining a lifelong presence within the host. This persistence is facilitated by the expression of latent genes such as EBV nuclear antigens (EBNAs) and latent membrane proteins (LMPs), which play crucial roles in viral latency and oncogenesis. In addition to their well-known roles in several types of cancer, including nasopharyngeal carcinoma and B-cell lymphomas, recent studies have identified the pathogenic roles of EBV in autoimmune diseases such as multiple sclerosis, rheumatoid arthritis, and systemic lupus erythematosus. This review highlights the intricate interactions between EBV and the host immune system, underscoring the need for further research to develop effective therapeutic and preventive strategies against EBV-associated diseases.

Portable Near-Infrared to Near-Infrared Platform for Homogeneous Quantification of Biomarkers in Complex Biological Samples.

Förster resonance energy transfer (FRET)-based homogeneous immunoassay obviates tedious washing steps and thus is a promising approach for immunoassays. However, a conventional FRET-based homogeneous immunoassay operating in the visible region is not able to overcome the interference of complex biological samples, thus resulting in insufficient detection sensitivity and poor accuracy. Here, we develop a near-infrared (NIR)-to-NIR FRET platform (Ex = 808 nm, Em = 980 nm) that enables background-free high-throughput homogeneous quantification of various biomarkers in complex biological samples. This NIR-to-NIR FRET platform is portable and easy to operate and is mainly composed of a high-performance NIR-to-NIR FRET pair based on lanthanide-doped nanoparticles (LnNPs) and a custom-made microplate reader for readout of NIR luminescence signals. We demonstrate that this NIR-to-NIR FRET platform is versatile and robust, capable of realizing highly sensitive and accurate detection of various critical biomarkers, including small molecules (morphine and 1,25-dihydroxyvitamin D), proteins (human chorionic gonadotropin), and viral particles (adenovirus) in unprocessed complex biological samples (urine, whole blood, and feces) within 5-10 min. We expect this NIR-to-NIR FRET platform to provide low-cost healthcare for populations living in resource-limited areas and be widely used in many other fields, such as food safety and environmental monitoring.

Portable sensing methods based on carbon dots for food analysis.

Food analysis is significantly important in monitoring food quality and safety for human health. Traditional methods for food detection mainly rely on benchtop instruments and require a certain amount of analysis time, which promotes the development of portable sensors. Portable sensing methods own many advantages over traditional techniques such as flexibility and accessibility in diverse environments, real-time monitoring, cost-effectiveness, and rapid deployment. This review focuses on the portable approaches based on carbon dots (CDs) for food analysis. CDs are zero-dimensional carbon-based material with a size of less than 10 nm. In the manner of sensing, CDs exhibit rich functional groups, low biotoxicity, good biocompatibility, and excellent optical properties. Furthermore, there are many methods for the synthesis of CDs using various precursor materials. The incorporation of CDs into food science and engineering for enhancing food safety control and risk assessment shows promising prospects.

Rapid label-free impedimetric detection of progesterone enhanced by immunomagnetic bead-based competitive immunoreaction.

Detecting progesterone (P4) concentration in cow serum is essential for monitoring the pregnancy progress after fertilization and is significant for the dairy farming industry and veterinary medicine. This study reports enzyme-free immunomagnetic beads (IMBs)-based competitive immunoassay for detecting P4 by P4-bovine serum albumin (BSA)-modified biosensors. The anti-P4 antibody-conjugated IMBs serve as collectors to capture P4 in undiluted serum samples to prevent the biosensor surface from biosample contamination and as insulated labels to report the electron-transfer resistance signal of electrochemical impedance spectroscopy (EIS) measurement. The IMBs and P4-containing samples were mixed for 15-30 min, capable of obtaining stable P4@IMB complexes. The 0.2-kGauss pulsed magnetic field (PMF) of the 20-s pulse width and 20-s relaxation time applied for 5 min can shorten the immunoreaction time between the P4@IMBs and the P4-BSA-modified biosensor and reduce the IMB's nonspecific adsorption on the biosensor surface. This competitive immunoassay's cut-off value and detection limit were 7.71 ng/mL and 7.33 ng/mL, respectively, which is lower than the serum's P4 plateau concentration (over 8 ng/mL) of dairy cows on days 6-16 of estrus cycles and that in pregnancy. The IMB-based immunoassay combining the PMF attraction and the label-free EIS measurement exhibits promising potential for rapidly detecting P4 in undiluted serum.

N-doped carbon dots for the determination of Al3+ and Fe3+ using aggregation-induced emission.

New portable hydrogel sensors for Al3+ and Fe3+ detection were designed based on the aggregation-induced emission (AIE) and color change of N-doped carbon dots (N-CDs). N-CDs with yellow fluorescence were prepared by a one-pot hydrothermal method from 2,5-dihydroxyterephthalic acid and acrylamide. The fluorescence of N-CDs was enhanced by Al3+ about 20 times and quenched by Fe3+. It was interesting that although Fe3+ showed obvious quenching on the fluorescence of N-CDs it did not cause a noticeable change in the fluorescence of N-CDs + Al3+. The colorless solution of N-CDs appeared blue in the presence of Fe3+ without the influence of Al3+. Therefore, the turn-on fluorometry and colorimetry systems based on N-CDs were constructed for the simultaneous detection of Al3+ and Fe3+. Furthermore, the portable sensing of Al3+ and Fe3+ was realized with the assistance of hydrogel, filter paper, cellulose acetate, and cellulose nitrate film. The proposed approach was successfully applied to the detection of Al3+ and Fe3+ in food samples and cell imaging.

A multi-cycle signal amplification-mediated single quantum dot nanosensor for PIWI-interacting RNA detection.

We construct a single quantum dot-based nanosensor for piRNA detection based on ligation-mediated multi-cycle signal amplification. This nanosensor is homogenous, selective, and sensitive with a detection limit of 0.104 fM. Moreover, it can detect the endogenous piRNA level in different cell lines, and discriminate cancer tissues from normal tissues.

Methylation-Powered Assembly of a Single Quantum Dot-Based FRET Nanosensor for Antibody-Free and Enzyme-Free Monitoring of Locus-Specific N6-Methyladenosine in Clinical Tissues.

N6-Methyladenosine (m6A) is the most pervasive and evolutionarily conserved epitranscriptomic modification in long noncoding RNA (lncRNA), and its dysregulation may induce aberrant transcription and translation programs. Herein, we demonstrate the methylation-powered assembly of a single quantum dot (QD)-based fluorescence resonance energy transfer (FRET) nanosensor for antibody- and enzyme-free monitoring of locus-specific m6A in clinical tissues. The m6A-sensitive DNAzyme VMC10 is employed to identify a specific m6A site in lncRNA, and it catalyzes the hydrolytic cleavage of unmethylated lncRNA. The cleaved lncRNA fails to trigger the subsequent catalytic hairpin assembly (CHA) reaction due to the energy barrier. In contrast, when m6A-lncRNA is present, the methyl group in m6A protects lncRNA from VMC10-mediated cleavage. With the aid of an assistant probe, the retained intact m6A-lncRNA is released from the VMC10/lncRNA complex and subsequently triggers the CHA reaction, generating abundant AF647/biotin dual-labeled duplexes. The assembly of AF647/biotin dual-labeled duplexes onto 605QD results in efficient FRET between 605QD and AF647. The FRET signal can be simply quantified by single-molecule detection. Notably, this assay can be implemented in an antibody-free and enzyme-free manner. This nanosensor can sensitively quantify target m6A with a detection limit of 0.47 fM, and it can discriminate as low as a 0.001% m6A level from excess coexisting counterparts. Importantly, this nanosensor can monitor the cellular m6A level with single-cell sensitivity and profile target m6A expression in breast cancer and healthy para-cancerous tissues, providing a powerful tool for studying the physiological and pathological functions of m6A.

A Bioinspired Flexible Sensor for Electrochemical Probing of Dynamic Redox Disequilibrium in Cancer Cells.

Malignant tumors pose a serious risk to human health. Ascorbic acid (AA) has potential for tumor therapy; however, the mechanism underlying the ability of AA to selectively kill tumor cells remains unclear. AA can cause redox disequilibrium in tumor cells, resulting in the release of abundant reactive oxygen species, represented by hydrogen peroxide (H2 O2 ). Therefore, the detection of H2 O2 changes can provide insight into the selective killing mechanism of AA against tumor cells. In this work, inspired by the ion-exchange mechanism in coral formation, a flexible H2 O2 sensor (PtNFs/CoPi@CC) is constructed to monitor the dynamics of H2 O2 in the cell microenvironment, which exhibits excellent sensitivity and spatiotemporal resolution. Moreover, the findings suggest that dehydroascorbic acid (DHA), the oxidation product of AA, is highly possible the substance that actually acts on tumor cells in AA therapy. Additionally, the intracellular redox disequilibrium and H2 O2 release caused by DHA are positively correlated with the abundance and activity of glucose transporter 1 (GLUT1). In conclusion, this work has revealed the potential mechanism underlying the ability of AA to selectively kill tumor cells through the construction and use of PtNFs/CoPi@CC. The findings provide new insights into the clinical application of AA.

Electrical Characterization and Analysis of Single Cells and Related Applications.

Biological parameters extracted from electrical signals from various body parts have been used for many years to analyze the human body and its behavior. In addition, electrical signals from cancer cell lines, normal cells, and viruses, among others, have been widely used for the detection of various diseases. Single-cell parameters such as cell and cytoplasmic conductivity, relaxation frequency, and membrane capacitance are important. There are many techniques available to characterize biomaterials, such as nanotechnology, microstrip cavity resonance measurement, etc. This article reviews single-cell isolation and sorting techniques, such as the micropipette separation method, separation and sorting system (dual electrophoretic array system), DEPArray sorting system (dielectrophoretic array system), cell selector sorting system, and microfluidic and valve devices, and discusses their respective advantages and disadvantages. Furthermore, it summarizes common single-cell electrical manipulations, such as single-cell amperometry (SCA), electrical impedance sensing (EIS), impedance flow cytometry (IFC), cell-based electrical impedance (CEI), microelectromechanical systems (MEMS), and integrated microelectrode array (IMA). The article also enumerates the application and significance of single-cell electrochemical analysis from the perspectives of CTC liquid biopsy, recombinant adenovirus, tumor cells like lung cancer DTCs (LC-DTCs), and single-cell metabolomics analysis. The paper concludes with a discussion of the current limitations faced by single-cell analysis techniques along with future directions and potential application scenarios.

Patterning amyloid-ß aggregation under the effect of acetylcholinesterase using a biological nanopore - an in vitro study.

Aggregation of amyloid-ß peptide (Aß) is hypothesized to be the primary cause of Alzheimer's disease (AD) progression. Aß aggregation has been widely studied using conventional sensing tools like emission fluorescence, electron microscopy, mass spectroscopy, and circular dichroism. However, none of these techniques can provide cost-efficient, highly sensitive quantification of Aß aggregation kinetics at the molecular level. Among the influences on Aß aggregation of interest to disease progression is the acceleration of Aß aggregation by acetylcholinesterase (AChE), which is present in the brain and inflicts the fast progression of disease due to its direct interaction with Aß. In this work, we demonstrate the ability of a biological nanopore to map and quantify AChE accelerated aggregation of Aß monomers to mixed oligomers and small soluble aggregates with single-molecule precision. This method will allow future work on testing direct and indirect effects of therapeutic drugs on AChE accelerated Aß aggregation as well as disease prognosis.

A Randomized Clinical Trial of Intravenous Methylprednisolone With 2 Protocols in Patients With Graves Orbitopathy.

CONTEXT: Intravenous glucocorticoid (IVGC) is an accessible and affordable treatment for Graves orbitopathy (GO); the 4.5-g protocol is well studied, but many details of treatment protocols need to be clarified. OBJECTIVE: To compare the efficacy and safety of weekly and monthly protocol of IVGC in GO. METHODS: A prospective, randomized, observer-masked, single-center clinical trial, followed up to week 24, at the third affiliated hospital of Southern Medical University; 58 patients with active and moderate to severe GO, aged 18-60 years old, who had not received relevant treatment were included. The intervention was weekly protocol or monthly protocol of IVGC; both received a cumulative dose of methylprednisolone 4.5 g and had a duration of 12 weeks. The overall effective rate, improvement of quality of life (QOL) and signal intensity ratio (SIR) were measured. RESULTS: There was no significant difference in the effective rate between the 2 groups at week 12 and week 24 (86.21% vs 72.41%, P = .195; 86.21% vs 82.61%, P = .441), there was no significant difference in the improvement of clinical activity score, exophthalmos, soft tissue involvement, diplopia, and QOL. At week 24, the mean SIR and maximum SIR of the 2 groups were lower than those before treatment, and there were no statistically significant difference between the 2 groups. There was no significant difference in the incidence of adverse events between the 2 groups (31.03% vs 27.59%, P = .773). CONCLUSION: The efficacy and safety of the 2 protocols are comparable; the monthly protocol could be used as an alternative to the weekly protocol.

Automatically showing microbial growth kinetics with a high-performance microbial growth analyzer.

It is difficult to show microbial growth kinetics online when they grow in complex matrices. We presented a novel strategy to address this challenge by developing a high-performance microbial growth analyzer (HPMGA), which employed a unique 32-channel capacitively coupled contactless conductivity detector as a sensing element and fixed with a CellStatz software. It was capable of online showing accurate and repeatable growth curves of well-dispersed and bad-dispersed microbes, whether they grew in homogeneous simple culture broth or heterogeneous complex matrices. Moreover, it could automatically report key growth kinetics parameters. In comparison to optical density (OD), plate counting and broth microdilution (BMD) methods, we demonstrated its practicability in five scenarios: 1) the illustration of the growth, growth rate, and acceleration curves of Escherichia coli (E. coli); 2) the antimicrobial susceptibility testing (AST) of Oxacillin against Staphylococcus aureus (S. aureus); 3) the determination of Ag nanoparticle toxicity on Providencia rettgeri (P. rettgeri); 4) the characterization of milk fermentation; and 5) the enumeration of viable pathogenic Vibrio in shrimp body. Results highlighted that the HPMGA method had the advantages of universality and effectivity. This technology would significantly facilitate the routine analysis of microbial growth in many fields (biology, medicine, clinic, life, food, environment, and ecology), paving an avenue for microbiologists to achieve research goals that have been inhibited for years due to a lack of practical analytical methods.

EvIPqPCR, Target Circulating Tumorous Extracellular Vesicles for Detection of Pancreatic Cancer.

Pancreatic cancer patients predominantly present with advanced disease at diagnosis, contributing to its high mortality. A noninvasive, fast screening method to detect this disease is an unmet need. Tumor-derived extracellular vesicles (tdEVs) bearing information from parental cells have emerged as a promising cancer diagnostic biomarker. However, most tdEV-based assays have impractical sample volumes and time-consuming, complex, and costly techniques. To overcome these limitations, we developed a novel diagnostic method for pancreatic cancer screening. Our approach utilizes the mitochondrial DNA to nuclear DNA ratio of EVs as a collective cell-specific characteristic. We introduce EvIPqPCR, a fast method that combines immunoprecipitation (IP) and qPCR quantification to detect tumor-derived EVs directly from serum. Importantly, our method employs DNA isolation-free and duplexing probes for qPCR, saving at least 3 h. This technique has the potential to serve as a translational assay for cancer screening with a weak correlation to prognosis biomarkers and sufficient discriminatory power among healthy controls, pancreatitis, and pancreatic cancer cases.

Droplet microarray platforms for high-throughput drug screening.

High-throughput screening platforms are fundamental for the rapid and efficient processing of large amounts of experimental data. Parallelization and miniaturization of experiments are important for improving their cost-effectiveness. The development of miniaturized high-throughput screening platforms is essential in the fields of biotechnology, medicine, and pharmacology. Currently, most laboratories use 96- or 384-well microtiter plates for screening; however, they have disadvantages, such as high reagent and cell consumption, low throughput, and inability to avoid cross-contamination, which need to be further optimized. Droplet microarrays, as novel screening platforms, can effectively avoid these shortcomings. Here, the preparation method of the droplet microarray, method of adding compounds in parallel, and means to read the results are briefly described. Next, the latest research on droplet microarray platforms in biomedicine is presented, including their application in high-throughput culture, cell screening, high-throughput nucleic acid screening, drug development, and individualized medicine. Finally, the challenges and future trends in droplet microarray technology are summarized.

Biosensors integrated 3D organoid/organ-on-a-chip system: A real-time biomechanical, biophysical, and biochemical monitoring and characterization.

As a full-fidelity simulation of human cells, tissues, organs, and even systems at the microscopic scale, Organ-on-a-Chip (OOC) has significant ethical advantages and development potential compared to animal experiments. The need for the design of new drug high-throughput screening platforms and the mechanistic study of human tissues/organs under pathological conditions, the evolving advances in 3D cell biology and engineering, etc., have promoted the updating of technologies in this field, such as the iteration of chip materials and 3D printing, which in turn facilitate the connection of complex multi-organs-on-chips for simulation and the further development of technology-composite new drug high-throughput screening platforms. As the most critical part of organ-on-a-chip design and practical application, verifying the success of organ model modeling, i.e., evaluating various biochemical and physical parameters in OOC devices, is crucial. Therefore, this paper provides a logical and comprehensive review and discussion of the advances in organ-on-a-chip detection and evaluation technologies from a broad perspective, covering the directions of tissue engineering scaffolds, microenvironment, single/multi-organ function, and stimulus-based evaluation, and provides a more comprehensive review of the progress in the significant organ-on-a-chip research areas in the physiological state.

Circulating Plasma Exosomal Proteins of Either SHIV-Infected Rhesus Macaque or HIV-Infected Patient Indicates a Link to Neuropathogenesis.

Despite the suppression of human immunodeficiency virus (HIV) replication by combined antiretroviral therapy (cART), 50-60% of HIV-infected patients suffer from HIV-associated neurocognitive disorders (HAND). Studies are uncovering the role of extracellular vesicles (EVs), especially exosomes, in the central nervous system (CNS) due to HIV infection. We investigated links among circulating plasma exosomal (crExo) proteins and neuropathogenesis in simian/human immunodeficiency virus (SHIV)-infected rhesus macaques (RM) and HIV-infected and cART treated patients (Patient-Exo). Isolated EVs from SHIV-infected (SHIV-Exo) and uninfected (CTL-Exo) RM were predominantly exosomes (particle size < 150 nm). Proteomic analysis quantified 5654 proteins, of which 236 proteins (~4%) were significantly, differentially expressed (DE) between SHIV-/CTL-Exo. Interestingly, different CNS cell specific markers were abundantly expressed in crExo. Proteins involved in latent viral reactivation, neuroinflammation, neuropathology-associated interactive as well as signaling molecules were expressed at significantly higher levels in SHIV-Exo than CTL-Exo. However, proteins involved in mitochondrial biogenesis, ATP production, autophagy, endocytosis, exocytosis, and cytoskeleton organization were significantly less expressed in SHIV-Exo than CTL-Exo. Interestingly, proteins involved in oxidative stress, mitochondrial biogenesis, ATP production, and autophagy were significantly downregulated in primary human brain microvascular endothelial cells exposed with HIV+/cART+ Patient-Exo. We showed that Patient-Exo significantly increased blood-brain barrier permeability, possibly due to loss of platelet endothelial cell adhesion molecule-1 protein and actin cytoskeleton structure. Our novel findings suggest that circulating exosomal proteins expressed CNS cell markers-possibly associated with viral reactivation and neuropathogenesis-that may elucidate the etiology of HAND.

Advancements in microfluidics for skin cosmetic screening.

With the global penetration of skin care awareness and upgrading of personal care awareness, the use rate of cosmetics and personal skin care products has been increasing worldwide. It is particularly important to monitor the quality and safety of skin cosmetics. In accordance with the requirements of the 7th Amendment of the European Cosmetics Directive 1223/2009, in vitro test methods have been developed to replace animal experiments, such as the 2D test, 3D test, microfluidic skin chip test, etc. The microfluidic skin chip overcomes the shortcomings of the 2D test and the 3D test that lack the complexity of human skin through fine control of the human skin microenvironment and induction of relevant mechanical stimulation. High similarity to real human skin through simulation of the vascular system and immune response. Therefore, the microfluidic skin chip is considered as a valuable and effective tool for the in vitro screening of cosmetics. In this paper, we reviewed the detection methods and technologies of common chemical substances, toxic elements, active substances and adverse reactions in vitro in quality monitoring of cosmetics. The most advantageous microfluidic skin chip technology is also introduced. The material and technology progress of skin chips used in cosmetic screening is reviewed and discussed. Then the application of microfluidic design in cosmetic screening in vitro is summarized.

Reining in Cas13a activity with N-terminal removable tags expands Cas13a based molecular sensing and enables precise gene interference.

Activation of Cas13 is exclusively dependent on crRNA-target RNA hybridization according to the canonical mode of Cas13 action. Upon activation Cas13 can cleave both target RNA and any surrounding RNA. The latter has been well adopted by therapeutic gene interference and biosensor development. This work for the first time, rationale designs and validates a multi-component controlled activation system of Cas13 by N-terminus tagging. A composite SUMO tag comprised of His, Twinstrep, and Smt3 tags fully suppresses target dependent activation of Cas13a by interfering with crRNA docking. The suppression releases upon proteases mediated proteolytic cleavage. The modular composition of the composite tag can be altered to fulfill customized response to alternative proteases. The biosensor SUMO-Cas13a is able to resolve a broad concentration range of protease Ulp1 with a calculated LOD of 48.8pg/µL in aqueous buffer. Further, in accordance with this finding Cas13a was successfully programmed to exert target gene knock down preferentially in SUMO protease high cell types. In summary the discovered regulatory component not only fulfills Cas13a based protease detection for the first time, but also delivers a novel strategy for multi-component controlled activation of Cas13a toward temporal and spacial precision.

Recent Advances in Electrochemical Immunosensors with Nanomaterial Assistance for Signal Amplification.

Electrochemical immunosensors have attracted immense attention due to the ease of mass electrode production and the high compatibility of the miniature electric reader, which is beneficial for developing point-of-care diagnostic devices. Electrochemical immunosensors can be divided into label-free and label-based sensing strategies equipped with potentiometric, amperometric, voltammetric, or impedimetric detectors. Emerging nanomaterials are frequently used on electrochemical immunosensors as a highly rough and conductive interface of the electrodes or on nanocarriers of immobilizing capture antibodies, electroactive mediators, or catalyzers. Adopting nanomaterials can increase immunosensor characteristics with lower detection limits and better sensitivity. Recent research has shown innovative immobilization procedures of nanomaterials which meet the requirements of different electrochemical immunosensors. This review discusses the past five years of advances in nanomaterials (metal nanoparticles, metal nanostructures, carbon nanotubes, and graphene) integrated into the electrochemical immunosensor. Furthermore, the new tendency and endeavors of nanomaterial-based electrochemical immunosensors are discussed.