Molecular Origins of Long-Term Changes in a Competitive Continuous Biosensor with Single-Molecule Resolution.

Biosensing by particle motion is a biosensing technology that relies on single-molecule interactions and enables the continuous monitoring of analytes from picomolar to micromolar concentration levels. However, during sensor operation, the signals are observed to change gradually. Here, we present a comprehensive methodology to elucidate the molecular origins of long-term changes in a particle motion sensor, focusing on a competitive sensor design under conditions without flow. Experiments were performed wherein only the particles or only the surfaces were aged in order to clarify how each individual component changes over time. Furthermore, distributions of particle motion patterns and switching activity were studied to reveal how particle populations change over timespans of several days. For a cortisol sensor with anticortisol antibodies on the particles and cortisol analogues on the sensing surface, the leading hypotheses for the long-term changes are (i) that the particles lose antibodies and develop nonspecific interactions and (ii) that analogue molecules dissociate from the sensing surface. The developed methodologies and the acquired insights pave a way for realizing sensors that can operate over long timespans.

A temperature compensated fiber probe for highly sensitive detection in virus gene biosensing.

This research presents an innovative reflective fiber optic probe structure, mutinously designed to detect H7N9 avian influenza virus gene precisely. This innovative structure skillfully combines multimode fiber (MMF) with a thin-diameter seven-core photonic crystal fiber (SCF-PCF), forming a semi-open Fabry-Pérot (FPI) cavity. This structure has demonstrated exceptional sensitivity in light intensity-refractive index (RI) response through rigorous theoretical and experimental validation. The development of a quasi-distributed parallel sensor array, which provides temperature compensation during measurements, has achieved a remarkable RI response sensitivity of up to 532.7 dB/RIU. The probe-type fiber optic sensitive unit, expertly functionalized with streptavidin, offers high specificity in detecting H7N9 avian influenza virus gene, with an impressively low detection limit of 10-2 pM. The development of this biosensor marks a significant development in biological detection, offering a practical engineering solution for achieving high sensitivity and specificity in light-intensity-modulated biosensing. Its potential for wide-ranging applications in various fields is now well-established.

Ureido-Ionic Liquid Mediated Conductive Hydrogel: Superior Integrated Properties for Advanced Biosensing Applications.

Ionic conductive hydrogels (ICHs) have recently gained prominence in biosensing, indicating their potential to redefine future biomedical applications. However, the integration of these hydrogels into sensor technologies and their long-term efficacy in practical applications pose substantial challenges, including a synergy of features, such as mechanical adaptability, conductive sensitivity, self-adhesion, self-regeneration, and microbial resistance. To address these challenges, this study introduces a novel hydrogel system using an imidazolium salt with a ureido backbone (UL) as the primary monomer. Fabricated via a straightforward one-pot copolymerization process that includes betaine sulfonate methacrylate (SBMA) and acrylamide (AM), the hydrogel demonstrates multifunctional properties. The innovation of this hydrogel is attributed to its robust mechanical attributes, outstanding strain responsiveness, effective water retention, and advanced self-regenerative and healing capabilities, which collectively lead to its superior performance in various applications. Moreover, this hydrogel  exhibited broad-spectrum antibacterial activity. Its potential for biomechanical monitoring, especially in tandem with contact and noncontact electrocardiogram (ECG) devices, represents a noteworthy advancement in precise real-time cardiac monitoring in clinical environments. In addition, the conductive properties of the hydrogel make it an ideal substrate for electrophoretic patches aimed at treating infected wounds and consequently enhancing the healing process.

Inspiring a convergent engineering approach to measure and model the tissue microenvironment.

Understanding the molecular and physical complexity of the tissue microenvironment (TiME) in the context of its spatiotemporal organization has remained an enduring challenge. Recent advances in engineering and data science are now promising the ability to study the structure, functions, and dynamics of the TiME in unprecedented detail; however, many advances still occur in silos that rarely integrate information to study the TiME in its full detail. This review provides an integrative overview of the engineering principles underlying chemical, optical, electrical, mechanical, and computational science to probe, sense, model, and fabricate the TiME. In individual sections, we first summarize the underlying principles, capabilities, and scope of emerging technologies, the breakthrough discoveries enabled by each technology and recent, promising innovations. We provide perspectives on the potential of these advances in answering critical questions about the TiME and its role in various disease and developmental processes. Finally, we present an integrative view that appreciates the major scientific and educational aspects in the study of the TiME.

Spatially confined dual-emission nanoprobes assembled from silicon nanoparticles and gold nanoclusters for ratiometric biosensing.

Gold nanoclusters (AuNCs) possess weak intrinsic fluorescence, limiting their sensitivity in biosensing applications. This study addresses these limitations by developing a spatially confined dual-emission nanoprobe composed of silicon nanoparticles (SiNPs) and AuNCs. This amplified and stabilized fluorescence mechanism overcomes the limitations associated with using AuNCs alone, achieving superior sensitivity in the sensing platform. The nanoprobe was successfully employed for ratiometric detection of bleomycin (BLM) in serum samples, operating at an excitation wavelength of 365 nm, with emission wavelengths at 480 nm and 580 nm. The analytical performance of the system is distinguished by a linear detection range of 0-3.5 µM, an impressive limit of detection (LOD) of 35.27 nM, and exceptional recoveries ranging from 96.80 to 105.9%. This innovative approach significantly enhances the applicability and reliability of AuNC-based biosensing in complex biological media, highlighting its superior analytical capabilities.

Impact of Particle Size on the Nonlinear Magnetic Response of Iron Oxide Nanoparticles during Frequency Mixing Magnetic Detection.

Magnetic nanoparticles (MNPs), particularly iron oxide nanoparticles (IONPs), play a pivotal role in biomedical applications ranging from magnetic resonance imaging (MRI) enhancement and cancer hyperthermia treatments to biosensing. This study focuses on the synthesis, characterization, and application of IONPs with two different size distributions for frequency mixing magnetic detection (FMMD), a technique that leverages the nonlinear magnetization properties of MNPs for sensitive biosensing. IONPs are synthesized through thermal decomposition and subsequent growth steps. Our findings highlight the critical influence of IONP size on the FMMD signal, demonstrating that larger particles contribute dominantly to the FMMD signal. This research advances our understanding of IONP behavior, underscoring the importance of size in their application in advanced diagnostic tools.

Nanoscale Metal-Organic Frameworks as a Photoluminescent Platform for Bioimaging and Biosensing Applications.

The tracking of nanomedicines in their concentration and location inside living systems has a pivotal effect on the understanding of the biological processes, early-stage diagnosis, and therapeutic monitoring of diseases. Nanoscale metal-organic frameworks (nano MOFs) possess high surface areas, definite structure, regulated optical properties, rich functionalized sites, and good biocompatibility that allow them to excel in a wide range of biomedical applications. Controllable syntheses and functionalization endow nano MOFs with better properties as imaging agents and sensing units for the diagnosis and treatment of diseases. This minireview summarizes the tunable synthesis strategies of nano MOFs with controllable size, shape, and regulated luminescent performance, and pinpoints their recent advanced applications as optical elements in bioimaging and biosensing. The current limitations and future development directions of nano MOF-contained materials in bioimaging and biosensing applications are also discussed, aiming to expand the biological applications of nano MOF-based nanomedicine and facilitate their production or clinical translation.

A smart tablet-phone-based high-performance pancreatic cancer cell biosensing system for drug screening.

Exploring more efficient pancreatic cancer drug screening platforms is of significant importance for accelerating the drug development process. In this study, we developed a high-sensitivity bioluminescence system based on smartphones and smart tablets, and constructed a pancreatic cancer drug screening platform (PCDSP) by combining the pancreatic cancer cell sensing model (PCCSM) on the multiwell plates (MTP). A smart tablet was used as the light source and a smartphone as the colorimetric sensing device. The smartphone dynamically controls the color and brightness displayed on the smart tablet to achieve lower LOD and wider detection ranges. We constructed PCCSM for 24 h, 48 h, and 72 h , and performed colorimetric experiments using both PCDSP and a commercial plate reader (CPR). The results showed that the PCDSP had a lower LOD than that of CPR. Moreover, PCDSP even exhibited a lower LOD for 24 h PCCSM testing compared to CPR for 48 h PCCSM testing, effectively shortening the drug evaluation process. Additionally, the PCDSP offers higher portability and efficiency compared with CPR, making it a promising platform for efficient pancreatic cancer drug screening.

A Mesoporous Gold Sensor Unveils Phospho PD-L1 in Extracellular Vesicles as a Proxy for PD-L1 Expression in Lung Cancer Tissue.

Immune checkpoint inhibitors (ICIs) targeting programmed cell death ligand 1 (PD-L1), or its receptor, PD-1 have improved survival in patients with non-small-cell lung cancer (NSCLC). Assessment of PD-L1 expression requires tissue biopsy or fine needle aspiration that are currently used to identify patients most likely to respond to single agent anti-PD-1/PD-L1 therapy. However, obtaining sufficient tissue to generate a PD-L1 tissue proportion score (TPS) ≥ 50% using immunohistochemistry remains a challenge that potentially may be overcome by liquid biopsies. This study utilized a mesoporous gold sensor (MGS) assay to examine the phosphorylation status of PD-L1 in plasma extracellular vesicles (EV pPD-L1) and PD-L1 levels in plasma from NSCLC patient samples and their association with tumor PD-L1 TPS. The 3-dimensional mesoporous network of the electrodes provides a large surface area, high signal-to-noise ratio, and a superior electro-conductive framework, thereby significantly improving the detection sensitivity of PD-L1 nanosensing. Test (n = 20) (Pearson's r = 0.99) and validation (n = 45) (Pearson's r = 0.99) cohorts show that EV pPD-L1 status correlates linearly with the tumor PD-L1 TPS assessed by immunohistochemistry irrespective of the tumor stage, with 64% of patients overall showing detectable EV pPD-L1 levels in plasma. In contrast to the EV pPD-L1 results, plasma PD-L1 levels did not correlate with the tumor PD-L1 TPS score or EV pPD-L1 levels. These data demonstrate that EV pPD-L1 levels may be used to select patients for appropriate PD-1 and PD-L1 ICI therapy regimens in early, locally advanced, and advanced NSCLC and should be tested further in randomized controlled trials. Most importantly, the assay used has a less than 24h turnaround time, facilitating adoption of the test into the routine diagnostic evaluation of patients prior to therapy.

Trends in surface plasmon resonance biosensing: materials, methods, and machine learning.

Surface plasmon resonance (SPR) proves to be one of the most effective methods of label-free detection and has been integral for the study of biomolecular interactions and the development of biosensors. This trend delves into the latest SPR research and progress built upon the Kretschmann configuration, a pivotal platform, and highlights three key developments that have enhanced the capabilities of the technique. We will first cover a range of explorations of novel plasmonic materials that have shaped SPR performance. Innovative signal transduction and collection, which leverages traditional materials and emerging alternatives, will then be discussed. Finally, the evolving landscape of data analysis, including the integration of machine learning algorithms to navigate complex SPR datasets, will be reviewed. We will also discuss the implementation of these improvements that have enabled new biosensing functions. These advancements not only pave the way for enhanced biosensing in general but also open new avenues for the technique to play a more significant role in research concerning human health.

From Structure to Application: The Evolutionary Trajectory of Spherical Nucleic Acids.

Since the proposal of the concept of spherical nucleic acids (SNAs) in 1996, numerous studies have focused on this topic and have achieved great advances. As a new delivery system for nucleic acids, SNAs have advantages over conventional deoxyribonucleic acid (DNA) nanostructures, including independence from transfection reagents, tolerance to nucleases, and lower immune reactions. The flexible structure of SNAs proves that various inorganic or organic materials can be used as the core, and different types of nucleic acids can be conjugated to realize diverse functions and achieve surprising and exciting outcomes. The special DNA nanostructures have been employed for immunomodulation, gene regulation, drug delivery, biosensing, and bioimaging. Despite the lack of rational design strategies, potential cytotoxicity, and structural defects of this technology, various successful examples demonstrate the bright and convincing future of SNAs in fields such as new materials, clinical practice, and pharmacy.

Biologically Active Micropatterns of Biomolecules and Living Matter Using Microbubble Lithography.

In situ patterning of biomolecules and living organisms while retaining their biological activity is extremely challenging, primarily because such patterning typically involves thermal stresses that could be substantially higher than the physiological thermal or stress tolerance level. Top-down patterning approaches are especially prone to these issues, while bottom-up approaches suffer from a lack of control in developing defined structures and the time required for patterning. A microbubble generated and manipulated by optical tweezers (microbubble lithography) is used to self-assemble and pattern living organisms in continuous microscopic structures in real-time, where the material thus patterned remains biologically active due to their ability to withstand elevated temperatures for short exposures. Successful patterns of microorganisms (Escherichia coli, Lactococcus. lactis and the Type A influenza virus) are demonstrated, as well as reporter proteins such as green fluorescent protein (GFP) on functionalized substrates with high signal-to-noise ratio and selectivity. Together, the data presented herein may open up fascinating possibilities in rapid in situ parallelized diagnostics of multiple pathogens and bioelectronics.

Dynamically predicting comprehension difficulties through physiological data and intelligent wearables.

Comprehending digital content written in natural language online is vital for many aspects of life, including learning, professional tasks, and decision-making. However, facing comprehension difficulties can have negative consequences for learning outcomes, critical thinking skills, decision-making, error rate, and productivity. This paper introduces an innovative approach to predict comprehension difficulties at the local content level (e.g., paragraphs). Using affordable wearable devices, we acquire physiological responses non-intrusively from the autonomous nervous system, specifically pulse rate variability, and electrodermal activity. Additionally, we integrate data from a cost-effective eye-tracker. Our machine learning algorithms identify 'hotspots' within the content and regions corresponding to a high cognitive load. These hotspots represent real-time predictors of comprehension difficulties. By integrating physiological data with contextual information (such as the levels of experience of individuals), our approach achieves an accuracy of 72.11% ± 2.21, a precision of 0.77, a recall of 0.70, and an f1 score of 0.73. This study opens possibilities for developing intelligent, cognitive-aware interfaces. Such interfaces can provide immediate contextual support, mitigating comprehension challenges within content. Whether through translation, content generation, or content summarization using available Large Language Models, this approach has the potential to enhance language comprehension.

An Exo III-powered closed-loop DNA circuit architecture for biosensing/imaging.

With their regulated Boolean logic operations in vitro and in vivo, DNA logic circuits have shown great promise for target recognition and disease diagnosis. However, significant obstacles must be overcome to improve their operational efficiency and broaden their range of applications. In this study, we propose an Exo III-powered closed-loop DNA circuit (ECDC) architecture that integrates four highly efficient AND logic gates. The ECDC utilizes Exo III as the sole enzyme-activated actuator, simplifying the circuit design and ensuring optimal performance. Moreover, the use of Exo III enables a self-feedback (autocatalytic) mechanism in the dynamic switching between AND logic gates within this circulating logic circuit. After validating the signal flow and examining the impact of each AND logic gate on the regulation of the circuit, we demonstrate the intelligent determination of miR-21 using the carefully designed ECDC architecture in vitro. The proposed ECDC exhibits a linear detection range for miR-21 from 0 to 300 nM, with a limit of detection (LOD) of approximately 0.01 nM, surpassing most reported methods. It also shows excellent selectivity for miR-21 detection and holds potential for identifying and imaging live cancer cells. This study presents a practical and efficient strategy for monitoring various nucleic acid-based biomarkers in vitro and in vivo through specific sequence modifications, offering significant potential for early cancer diagnosis, bioanalysis, and prognostic clinical applications.

From lab to wearables: Innovations in multifunctional hydrogel chemistry for next-generation bioelectronic devices.

Functional hydrogels have emerged as foundational materials in diagnostics, therapy, and wearable devices, owing to their high stretchability, flexibility, sensing, and outstanding biocompatibility. Their significance stems from their resemblance to biological tissue and their exceptional versatility in electrical, mechanical, and biofunctional engineering, positioning themselves as a bridge between living organisms and electronic systems, paving the way for the development of highly compatible, efficient, and stable interfaces. These multifaceted capability revolutionizes the essence of hydrogel-based wearable devices, distinguishing them from conventional biomedical devices in real-world practical applications. In this comprehensive review, we first discuss the fundamental chemistry of hydrogels, elucidating their distinct properties and functionalities. Subsequently, we examine the applications of these bioelectronics within the human body, unveiling their transformative potential in diagnostics, therapy, and human-machine interfaces (HMI) in real wearable bioelectronics. This exploration serves as a scientific compass for researchers navigating the interdisciplinary landscape of chemistry, materials science, and bioelectronics.

Plasmonics nanorod biosensor for in situ intracellular detection of gene expression biomarkers in intact plant systems.

The intracellular developmental processes in plants, particularly concerning lignin polymer formation and biomass production are regulated by microRNAs (miRNAs). MiRNAs including miR397b are important for developing efficient and cost-effective biofuels. However, traditional methods of monitoring miRNA expression, like PCR, are time-consuming, require sample extraction, and lack spatial and temporal resolution, especially in real-world conditions. We present a novel approach using plasmonics nanosensing to monitor miRNA activity within living plant cells without sample extraction. Plasmonic biosensors using surface-enhanced Raman scattering (SERS) detection offer high sensitivity and precise molecular information. We used the Inverse Molecular Sentinel (iMS) biosensor on unique silver-coated gold nanorods (AuNR@Ag) with a high-aspect ratio to penetrate plant cell walls for detecting miR397b within intact living plant cells. MiR397b overexpression has shown promise in reducing lignin content. Thus, monitoring miR397b is essential for cost-effective biofuel generation. This study demonstrates the infiltration of nanorod iMS biosensors and detection of non-native miRNA 397b within plant cells for the first time. The investigation successfully demonstrates the localization of nanorod iMS biosensors through TEM and XRF-based elemental mapping for miRNA detection within plant cells of Nicotiana benthamiana. The study integrates shifted-excitation Raman difference spectroscopy (SERDS) to decrease background interference and enhance target signal extraction. In vivo SERDS testing confirms the dynamic detection of miR397b in Arabidopsis thaliana leaves after infiltration with iMS nanorods and miR397b target. This proof-of-concept study is an important stepping stone towards spatially resolved, intracellular miRNA mapping to monitor biomarkers and biological pathways for developing efficient renewable biofuel sources.

A label-free and rapid fluorometric strategy for microRNA detection using CRISPR-Cas12a coupled with copper nanoparticles.

CRISPR-Cas12a with robust trans-cleavage activity were employed to mitigate background fluorescence signal, achieving sensitive detection of miRNA-21. The activation of trans-cleavage activity of Cas12a was achieved by utilizing cDNA as a trigger. Upon the presence of target miRNA-21, cDNA hybridizes with it forming a DNA/RNA double-stranded structure. Exonuclease III (ExoIII) facilitates the degradation of cDNA, releasing the target for subsequent cycles. Due to cDNA degradation, the trans-cleavage activity of Cas12a remains unactivated and does not disrupt the synthesis template of copper nanoparticles. Addition of Cu2+ and AA leads to the formation of highly fluorescent copper nanoparticles. Conversely, in absence of miRNA-21, intact cDNA activates trans-cleavage activity of Cas12a, resulting in degradation of the synthesis template and failure in synthesizing fluorescent copper nanoparticles. This method exhibits excellent selectivity with a low limit of detection (LOD) at 5 pM. Furthermore, we successfully applied this approach to determine miRNA-21 in cell lysates and human serum samples, providing a new approach for sensitive determination of biomarkers in biochemical research and disease diagnosis.

Objective-Free Ultrasensitive Biosensing on Large-Area Metamaterial Surfaces in the Near-IR.

Plasmonic metamaterials have opened new avenues in medical diagnostics. However, the transfer of the technology to the markets has been delayed due to multiple challenges. The need of bulky optics for signal reading from nanostructures patterned on submillimeter area limits the miniaturization of the devices. The use of objective-free optics can solve this problem, which necessitates large area patterning of the nanostructures. In this work, we utilize laser interference lithography (LIL) to pattern nanodisc-shaped metamaterial absorber nanoantennas over a large area (4 cm2) within minutes. The introduction of a sacrificial layer during the fabrication process enables an inverted hole profile and a well-controlled liftoff, which ensures perfectly defined uniform nanopatterning almost with no defects. Furthermore, we use a macroscopic reflection probe for optical characterization in the near-IR, including the detection of the binding kinematics of immunologically relevant proteins. We show that the photonic quality of the plasmonic nanoantennas commensurates with electron-beam-lithography-fabricated ones over the whole area. The refractive index sensitivity of the LIL-fabricated metasurface is determined as 685 nm per refractive index unit, which demonstrates ultrasensitive detection. Moreover, the fabricated surfaces can be used multiple times for biosensing without losing their optical quality. The combination of rapid and large area nanofabrication with a simple optical reading not only simplifies the detection process but also makes the biosensors more environmentally friendly and cost-effective. Therefore, the improvements provided in this work will empower researchers and industries for accurate and real-time analysis of biological systems.

Bait and Cleave: Exosite-Binding Peptides on Quantum Dots Selectively Accelerate Protease Activity for Sensing with Enhanced Sensitivity.

The advantageous optical properties of quantum dots (QDs) motivate their use in a wide variety of applications related to imaging and bioanalysis, including the detection of proteases and their activity. Recent studies have shown that surface chemistry on QDs is able to modulate protease activity, but only nonspecifically. Here, we present a strategy to selectively accelerate the activity of a particular target protease by as much as two orders of magnitude. Exosite-binding "bait" peptides were derived from proteins that span a range of biological roles─substrate, receptor, and inhibitor─and were used to increase the affinity of the QD-peptide conjugates for either thrombin or factor Xa, resulting in increased rates of proteolysis for coconjugated substrates. Unlike effects from QD surface chemistry, the acceleration was specific to the target protease with negligible acceleration of other proteases. Benefits of this "bait and cleave" sensing approach included detection limits that improved by more than an order of magnitude, reenabled detection of target protease against an overwhelming background of nontarget proteolysis, and mitigation of the action of inhibitors. The cumulative results point to a generalizable strategy, where the mechanism of acceleration, considerations for the design of bait peptides and conjugates, and routes to expanding the scope of this approach are discussed. Overall, this research represents a major step forward in the rational design of nanoparticle-based enzyme sensors that enhance sensitivity and selectivity.

Real-time label-free three-dimensional invasion assay for anti-metastatic drug screening using impedance sensing.

Tumor metastasis presents a formidable challenge in cancer treatment, necessitating effective tools for anti-cancer drug development. Conventional 2D cell culture methods, while considered the "gold standard" for invasive studies, exhibit limitations in representing cancer hallmarks and phenotypes. This study proposes an innovative approach that combines the advantages of 3D tumor spheroid culture with impedance-based biosensing technologies to establish a high-throughput 3D cell invasion assay for anti-metastasis drug screening through multicellular tumor spheroids. In addition, the xCELLigence device is employed to monitor the time-dependent kinetics of cell behavior, including attachment and invasion out of the 3D matrix. Moreover, an iron chelator (deferoxamine) is employed to monitor the inhibition of epithelial-mesenchymal transition in 3D spheroids across different tumor cell types. The above results indicate that our integrated 3D cell invasion assay with impedance-based sensing could be a promising tool for enhancing the quality of the drug development pipeline by providing a robust platform for predicting the efficacy and safety of anti-metastatic drugs before advancing into preclinical or clinical trials.