Recognition Events Research Articles

A split-type electrochemical biosensor using enzyme-linked DNA magnetic beads realizes the detection of BCR/ABLp210 fusion gene in clinical samples: Duplex ligation chain reaction coupled with OR logic gate design

Detection of BCR/ABLp210 transcript levels, allowing for the early diagnosis of chronic myelocytic leukemia (CML) and its minimal residual disease (MRD) monitoring, plays a decisive role in the cure of CML. Herein, to realize the sensitive detection of BCR/ABLp210 transcript by electrchemical DNA (E-DNA) sensors in clinical samples, a split-type strategy using enzyme-linked DNA magnetic beads (MBs) was intelligently provided. Specifically, the utilization of magnetic separation makes the target recognition event separate from electrochemical measurements, avoiding the interference of complex matrices on the sensing interface. Besides, the MBs adopted as dispersible electrodes could improve mass transfer of target compared to that of a planarelectrode. Consequently, the proposed E-DNA sensor combined with the ligation chain reaction (LCR) could achieve an exceptional performance with a low detection limit of 1 aM, a broad linear concentration range, and a remarkable specificity with a single-base resolution. Also, K562 cells in up to a 1000-fold excess of NB4 cells could be successfully detected by this E-DNA sensor. Finally, the integration of duplex LCR and OR logic gate into the E-DNA sensor has enabled the detection of e13a2/e14a2/both isoforms of BCR/ABLp210 transcript in a single assay in both newly diagnosed CML patients and those undergoing or having undergone treatment by tyrosine kinase inhibitors. Therefore, this method offers an alternative for the early diagnosis of CML and its MRD monitoring, and holds a huge potential of clinical translation due to its advantage of low cost, ease of miniaturization and generalizability.

Cyclic Peptides as Aggregation Inhibitors for Sickle Cell Disease.

Sickle cell disease is a missense genetic disorder characterized by the aggregation of deoxy-HbS into helical fibers that distort erythrocytes into a sickle-like shape. Herein, we investigate, through molecular dynamics, the effect of nine 5-mer cyclic peptides (CPs), tailor-designed to block key lateral contacts of the fibers. Our results show that the CPs bind orthogonally to the main HbS pocket involved in the latter contacts, with some revealing exceedingly long residence times. These CPs display moderate to high specificity, exhibiting molecular recognition events even at a HbS/CP (1:1) ratio. A much lower HbS-CP binding free energy, longer residence times, and higher specificity are also found relative to a previously reported CP with modest in vitro antisickling activity. These results indicate that some of these CPs have the potential to reduce the concentration of aggregation-competent deoxy-HbS, precluding or delaying the formation of lateral contact at the homogeneous nucleation stage.

Direct Z-scheme NiTiO3/polyaniline heterojunction based photoelectrochemical aptasensor for the efficient detection of ochratoxin A in corn and soil

Preparation of Z-scheme heterojunctions requires band-gap matched semiconductors with tedious procedures, which is challenging for the application of photoelectrochemical (PEC) sensing. Herein, a sensitive PEC aptasensor for ochratoxin A (OTA) detection was constructed based on direct Z-scheme NiTiO3/polyaniline (NiTiO3/PANI) heterojunction. NiTiO3/PANI was firstly prepared via electrostatic adsorption between monomers; PANI with high conductivity and hydrophilicity can engineer the surface of NiTiO3 to enhance the separation of charge carriers and enlarge the PEC signal (e.g., 71.8-fold higher of that at NiTiO3). By virtue of superior properties, a NiTiO3/PANI-based PEC aptasensor was developed by using the recognition event of OTA with aptamer to vary photocurrent. When OTA is present, the aptamer is pulled away from the electrode interface by OTA, leading to a decrease in steric hindrance and the recovery of photocurrent, thereby enabling the detection of OTA. The constructed PEC aptasensor offered a linear range of 1 pg mL−1–2000 pg mL−1 with a detection limit of 0.33 pg mL−1. Moreover, this PEC aptasenosr displayed satisfactory selectivity and stability. The strategies to fabricate high-performance direct Z-scheme heterojunction as well as PEC aptasensor offer new solutions for the efficient detection of mycotoxins in the field of food safety and environmental protection.

The Why and How of Ultrasmall Nanoparticles.

ConspectusIn this Account, we describe our research into ultrasmall nanoparticles, including their unique properties, and outline some of the new opportunities they offer. We will summarize our perspective on the current state of the field and highlight what we see as key questions that remain to be solved. First, there are several nanostructure size-scale regimes, with qualitatively distinct functional biological attributes. Broadly generalized, larger particles (e.g., larger than 300 nm) tend to be more efficiently swept away by the first line of the immune system (for example macrophages). In the "middle-sized" regime (20-300 nm), nanoparticle surfaces and shapes can be recognized by energy-dependent cellular reorganizations, then organized locally in a spatial and temporally coherent way. That energy is gated and made available by specific cellular recognition processes. The relationship between particle surface design, endogenously derived nonspecific biomolecular corona, and architectural features recognized by the cell is complex and only purposefully and very precisely designed nanoparticle architectures are able to navigate to specific targets. At sufficiently small sizes (<10 nm including the ligand shell, associated with a core diameter of a few nm at most) we enter the "quasi-molecular regime" in which the endogenous biomolecular environment exchanges so rapidly with the ultrasmall particle surface that larger scale cellular and immune recognition events are often greatly simplified. As an example, ultrasmall particles can penetrate cellular and biological barriers within tissue architectures via passive diffusion, in much the same way as small molecule drugs do. An intriguing question arises: what happens at the interface of cellular recognition and ultrasmall quasi-molecular size regimes? Succinctly put, ultrasmall conjugates can evade defense mechanisms driven by larger scale cellular nanoscale recognition, enabling them to flexibly exploit molecular interaction motifs to interact with specific targets. Numerous advances in control of architecture that take advantage of these phenomena have taken place or are underway. For instance, syntheses can now be sufficiently controlled that it is possible to make nanoparticles of a few hundreds of atoms or metalloid clusters of several tens of atoms that can be characterized by single crystal X-ray structure analysis. While the synthesis of atomically precise clusters in organic solvents presents challenges, water-based syntheses of ultrasmall nanoparticles can be upscaled and lead to well-defined particle populations. The surface of ultrasmall nanoparticles can be covalently modified with a wide variety of ligands to control the interactions of these particles with biosystems, as well as drugs and fluorophores. And, in contrast to larger particles, many advanced molecular analytical and separation tools can be applied to understand their structure. For example, NMR spectroscopy allows us to obtain a detailed image of the particle surface and the attached ligands. These are considerable advantages that allow further elaboration of the level of architectural control and characterization of the ultrasmall structures required to access novel functional regimes and outcomes. The ultrasmall nanoparticle regime has a unique status and provides a potentially very interesting direction for development.

Rozvytok instytutu rozpodilu maina podruzhzhia za zakonodavstvom, yake diialo na ukrainskykh zemliakh

The scientific article examines the issue of the development of the institution of the division of marital property according to the legislation that was in force on Ukrainian lands. On the basis of the conducted research, the author comes to a conclusion that during the period of operation of Roman private law on the territory of the Greek polis states of the Northern Black Sea region, the institution of the division of joint property of spouses existed only de facto through the mechanism of transfer of property transferred as a «premarital gift». Norms of canonical and princely law of the Kyiv state did not contain provisions on the division of property of spouses, however, in the customary law of that time there was a practice of separating property into the ownership of a family member, in particular, the division of joint property could also apply to one spouse who received the corresponding property rights, primarily in order of inheritance. In the Lithuanian Statute of 1588, next to separate property, for the first time the possibility of joint ownership of property acquired during marriage was recognized, but only for representatives of the peasant class, but the procedure for the division of such property was not defined. In the «Laws by which the Little Russian People are judged», for the first time at the level of a draft of a written legal act, the specifics of the division of the property of the spouses in the event of a divorce are defined; the provisions of the Code of Laws of the Russian empire provided for the possibility of only contractual regulation of the division of joint property of spouses and only with respect to that property that was located within the boundaries of the Kyiv, Volyn, and Podillia governorates. The judicial procedure for resolving disputes about the division of joint property of spouses was introduced in the Galician Civil Code, and the Austrian civil code allowed the possibility of establishing joint property by a separate contract of the spouses and also determined that the joint property of the spouses is terminated, and the property in question is subject to division in the event of recognition of one of the spouses incapacitated, «voluntary divorce», «judicial divorce», annulment of marriage or its dissolution. For the first time, the legal regime of joint ownership of property acquired by spouses during the marriage on Ukrainian lands was determined by the Law of the Ukrainian People’s Republic «On Marriage and Divorce and on the Registration of Acts of Marriage, Divorce, Birth and Death», and subsequently by Soviet acts of family legislation, starting with the Code of family, guardianship, marriage and acts of civil status UPCP of 1926. Despite the fact that the Decree of the Soviet People’s Committee of the Ukrainian SSR «On civil marriage and keeping records of acts of civil status» and the Code of Laws on acts of civil status, on family and guardianship of 1919, it was assumed that property acquired in marriage was not considered joint property of the spouses, however, in the judicial practice of that time regarding the resolution of disputes about divorce, the opposite was recognized and the judicial division of such property, acquired by their joint labor and funds, was carried out. The procedure and methods of division of common property were initially determined by acts of civil legislation and since 1969 — family legislation. At the same time, the principle of equal shares of spouses in common property was also introduced in 1969.

Deep learning model using ensemble based approach for walking activity recognition and gait event prediction with grey level co-occurrence matrix

Deep learning model using ensemble based approach for walking activity recognition and gait event prediction with grey level co-occurrence matrix

CRISPR/Cas-Assisted Nanoneedle Sensor for Adenosine Triphosphate Detection in Living Cells.

The clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein (Cas) (CRISPR/Cas) systems have recently emerged as powerful molecular biosensing tools based on their collateral cleavage activity due to their simplicity, sensitivity, specificity, and broad applicability. However, the direct application of the collateral cleavage activity for in situ intracellular detection is still challenging. Here, we debut a CRISPR/Cas-assisted nanoneedle sensor (nanoCRISPR) for intracellular adenosine triphosphate (ATP), which avoids the challenges associated with intracellular collateral cleavage by introducing a two-step process of intracellular target recognition, followed by extracellular transduction and detection. ATP recognition occurs by first presenting in the cell cytosol an aptamer-locked Cas12a activator conjugated to nanoneedles; the recognition event unlocks the activator immobilized on the nanoneedles. The nanoneedles are then removed from the cells and exposed to the Cas12a/crRNA complex, where the activator triggers the cleavage of an ssDNA fluorophore-quencher pair, generating a detectable fluorescence signal. NanoCRISPR has an ATP detection limit of 246 nM and a dynamic range from 1.56 to 50 μM. Importantly, nanoCRISPR can detect intracellular ATP in 30 min in live cells without impacting cell viability. We anticipate that the nanoCRISPR approach will contribute to broadening the biomedical applications of CRISPR/Cas sensors for the detection of diverse intracellular molecules in living systems.

Fluoride ion detection in aqueous medium: Colorimetric and turn-off fluorescent Schiff base chemosensor

The work described is focused on a newly developed colorimetric Schiff base fluorescent sensor that exhibits a “turn-off” response when detecting fluoride ions (F−) in an aqueous environment. The confirmation of the recognition event is accomplished through fluorescence titration, absorbance titration, and Density Functional Theory (DFT) calculation study. From the results, it is determined that the detection limit of F− is 2.35 × 10−8 M which is much lower than the World Health Organization (WHO) permissible limit for drinking water while the binding constant was obtained to be 0.1 × 106 M−1 indicating a moderate affinity for the fluoride ions. Furthermore, the binding stoichiometry between the Probe L and F− was found to be 1:1 which is evidenced by the Job’s plot and DFT calculation study. Overall, the novel sensor's impressive sensitivity and selectivity make it a promising candidate for the detection of fluoride ions in aqueous media, particularly for monitoring drinking water quality to ensure compliance with WHO guidelines.

Engineering a Two-in-one Ni(II)-azophenine switch: Intelligent Lab-on a-box device for decentralized recognition event

A new Ni2+ complex of an azophenine-based sensory receptor, CN-1•Ni2+ is judiciously designed for selective aqueous phase fluoride recognition based on specific hydrogen bonding interaction with sharp chromogenic variation from brownish orange to purple. The mechanistic pathway of interaction has been thoroughly investigated via ESI-MS, FT-IR, differential pulse voltammetry (DPV) studies with theoretical DFT support and Loewdin spin population analysis. The detection limit was acquired to be 148 ppb, which is lower than the WHO permissible limit. The real-time sensitivity of the probe has been displayed by TLC strip-based solid-state sensing assay. The probe is also responsive towards recognition of fluoride from various real field water specimens with promising recoveries (95–119%). One step ahead, the chemically obtained outcomes have been extensively synchronized with electronic circuitry to fabricate lab-on a-box based Layman’s prototype for on-spot, decentralized detection of fluoride, which would be beneficial for the end users, particularly in the fluoride endemic areas. Moreover, based on the reversible spectroscopic outcome, herein AND-NOT-OR-NOR gate-based complex logic circuitry and sequence-dependent molecular keypad lock has been fabricated. In addition, silica gel supported composite material, CN-1•Ni2+@SG has been prepared for effectual remediation of aqueous fluoride below safe limit. Thus, the present work relies on recognition as well as proficient remediation of aqueous fluoride based on selective tuning of acidity, validating the ‘Two-in-one’ approach.

Enantioselective Target Transport-Mediated Nanozyme Decomposition for the Identification of Reducing Enantiomers in Asymmetric Nanochannel Arrays.

Enantioselective identification of chiral molecules is regarded as one of the key issues in biological and medical sciences because of their configuration-dependent effects on biological systems. In this study, we developed an electrochemical platform based on a tandem recognition-reaction zone design in TiO2 nanochannels for the specific recognition of reducing enantiomers. In this system, MIL-125(Ti) Ti-metal-organic frameworks, in situ grown in TiO2 nanochannels, provided a homochiral recognition environment via postmodification with l-tartaric acid (l-TA); MnO2 nanosheets possessing both glucose oxidase (GOD)- and peroxidase (POD)-mimicking activities served as the target-reactive zone at the end of the nanochannels. The use of penicillamine (Pen) enantiomers as model-reducing targets facilitated the passage of d-Pen through the homochiral recognition zone, owing to its lower affinity with l-TA. The passed Pen molecules reached the responsive zone and induced a target concentration-dependent MnO2 disassembly. Such target recognition event impaired the cascade GOD- and POD-like activities of MnO2. Combining the enantioselectivity of the recognition nanochannels with the cascade enzyme-like activity of MnO2 toward glucose and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonate), the quantitative identification of l- and d-Pen was achieved through the changes in transmembrane ionic current induced by the generated charged products. This recognition-reaction zone design paves an effective way for developing a promising electrochemical platform for the identification of reducing enantiomers with improved selectivity and sensitivity.

NLR receptors in plant immunity: making sense of the alphabet soup.

Plants coordinately use cell-surface and intracellular immune receptors to perceive pathogens and mount an immune response. Intracellular events of pathogen recognition are largely mediated by immune receptors of the nucleotide binding and leucine rich-repeat (NLR) classes. Upon pathogen perception, NLRs trigger a potent broad-spectrum immune reaction, usually accompanied by a form of programmed cell death termed the hypersensitive response. Some plant NLRs act as multifunctional singleton receptors which combine pathogen detection and immune signaling. However, NLRs can also function in higher order pairs and networks of functionally specialized interconnected receptors. In this article, we cover the basic aspects of plant NLR biology with an emphasis on NLR networks. We highlight some of the recent advances in NLR structure, function, and activation and discuss emerging topics such as modulator NLRs, pathogen suppression of NLRs, and NLR bioengineering. Multi-disciplinary approaches are required to disentangle how these NLR immune receptor pairs and networks function and evolve. Answering these questions holds the potential to deepen our understanding of the plant immune system and unlock a new era of disease resistance breeding.

Amplifying sensing performance through gold micropatterns-induced modulation of graphene's vertical electron transfer

High-quality graphene monolayers exhibit limited vertical electron transfer properties due to their inert basal plan structure, hampering their use in electrochemical sensing devices. Overcoming this limitation while maintaining the graphene's basal plane electrical conductivity is crucial for developing sensitive and efficient electrochemical sensors. In this study, we demonstrate that incorporating a gold support beneath graphene enhances its electrochemical activity by facilitating effective hybridizations between carbon atoms in graphene and gold atoms. To achieve this, micrometer-sized gold micropatterns were fabricated on Si/SiO2 substrates to support the graphene monolayer. The presence of gold significantly improved the electrochemical activity of graphene towards redox probes in solution (ferri-/ferrocyanide) as well as for adsorbed ferrocene, indicating a synergistic effect between graphene and gold. We further investigated the adsorption effects of double-stranded DNA on pristine graphene, bare gold, and graphene with a gold underlayer, revealing that the graphene-gold combination achieved a significantly lower limit of detection for DNA hybridization events, surpassing the other configurations. These findings highlight the importance of appropriate substrate selection in the design of graphene-based electrochemical sensors, particularly in the context of DNA recognition events. The proposed approach of incorporating a gold support beneath graphene holds promise for enhancing the performance of electrochemical sensing devices without compromising the intrinsic properties of graphene.

“OR” logic gate multiplexed photoelectrochemical sensor for high-risk human papillomaviruses: “One pot” recombinase polymerase amplification and logic discrimination

Multiple targets analysis in complex samples is of great importance in medical and health sciences. Limited by independent laborious operational procedures, multiple targets determination remains a challenge. Herein, we report an “OR” logic gate multiplexed photoelectrochemical (PEC) sensor based on “one pot” recombinase polymerase amplification (RPA) strategy. “One pot” RPA triggers exponential growth of multiple DNA in complex samples. Subsequently, the amplification products interact separately with lambda exonuclease (λ exo) or Cas12a-crRNA. Following the multiple targets recognition event, the dual enzyme-mediated cleavage separates the signal labels from the photocathode. The resulting photocurrent change is utilized for logical discrimination and detection. The feasibility of the sensor is verified by analyzing the two typical duplex DNA (high-risk human papillomaviruses (HPV)). Ultralow detection limit (0.088 fg/μL, 0.081 fg/μL) with broad detection range (0.1 fg/μL to 10 ng/μL, 0.1 fg/μL to 1 ng/μL) for HPV16 and HPV18 are obtained. Eliminating instrumentation constraints (light source/potential modulation) and simplifying operation procedures, this work opens an avenue for developing multiplexed sensing devices for clinical diagnosis and disease treatment.

Nonmonotonic Modulation of the Protein-Ligand Recognition Event by Inert Crowders.

The ubiquitous event of a protein recognizing small molecules or ligands at its native binding site is crucial for initiating major biological processes. However, how a crowded environment, as is typically represented by a cellular interior, would modulate the protein-ligand search process is largely debated. Excluded volume-based theory suggests that the presence of an inert crowder would reinforce a steady stabilization and enhancement of the protein-ligand recognition process. Here, we counter this long-held perspective via the molecular dynamics simulation and Markov state model of the protein-ligand recognition event in the presence of inert crowders. Specifically, we demonstrate that, depending on concentration, even purely inert crowders can exert a nonmonotonic effect via either stabilizing or destabilizing the protein-ligand binding event. Analysis of the kinetic network of binding pathways reveals that the crowders would either modulate precedent non-native on-pathway intermediates or would devise additional ones in a multistate recognition event across a wide range of concentrations. As an important insight, crowders gradually shift the relative transitional preference of these intermediates toward a native-bound state, with ligand residence time at the binding pocket dictating the trend of nonmonotonic concentration dependence by simple inert crowders.

From Gene-for-Gene to Resistosomes: Flor's Enduring Legacy.

The gene-for-gene model proposed by H. H. Flor has been one of the fundamental precepts of plant-pathogen interactions that has underpinned decades of research towards our current concepts of plant immunity. The broad validity of this model as an elegant and accurate genetic description of specific recognition events between the products of plant resistance (R) and pathogen avirulence (Avr) genes has been demonstrated many times over in a wide variety of plant disease systems. In recent years detailed molecular and structural analyses have provided a deep understanding of the principles by which plant immune receptors recognize pathogen effectors, including providing molecular descriptions of many of the genetic loci in flax and flax rust characterized by Flor. Recent advances in molecular and structural understanding of immune receptor recognition and activation mechanisms have brought the field to a new level, where rational design of novel receptors through engineering approaches is becoming a realizable goal. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Atomic visualization of flipped-back conformations of high mannose glycans interacting with cargo lectins: An MD simulation perspective.

Protein-carbohydrate interactions play a crucial role in mediating several biomolecular recognition events. We attempt to unravel its intricacies by understanding how carbohydrate-binding proteins interpret the glycan code. We aim to decipher lectin-mediated recognition in the endoplasmic reticulum (ER), which plays a crucial role in ER-mediated quality control (ER-QC). The ER-QC functions in three phases-protein folding, transport, and degradation. Altered protein QC leads to ER-related storage disorders. Cargo transport proteins-Ergic53 and Vip36-necessary for maintaining cellular homeostasis-are our primary focus. They recognize monoglucosylated/high mannose N-glycans on the folded glycoproteins. This article reports on the first dynamic investigation of the ER cargo lectins in complex with the high mannose glycans using an advanced sampling technique-replica exchange molecular dynamics to decipher the inherent conformational heterogeneity and the binding mechanism. The study involves simulations for the proteins complexed with three high mannose glycans-Man8B, Man9, and mono-glucosylated glycan. The recognition process is captured using MD simulations to achieve mechanistic insights and characterize the dynamics of glycans in their native and bound states via dihedral angle analysis. Results indicate that the flipped conformation of the glycans was crucial in differentiating their interaction with the proteins. Similar conformers of the glycans are preferred for Ergic53 and Vip36 in their glycan recognition events. Ergic53 preferred Man8B while it was Man9 for Vip36, in coherence with the previous experimental reports. These simulations provide a computational microscopic purview of the mechanism at both spatial and temporal scales. The results correlate with the published experimental data on the specificities of these lectins.

Synthetic Receptor Based on a Peptide Antibiotic-Functionalized Chimera for Hybridization-Based Polynucleotide Detection.

Nanopores offer highly sensitive, low-cost, and single-molecule sensing capabilities, and the societal impact of this approach is best captured by the advent of nanopore-based DNA detection and sequencing technologies, which extract genomic information without amplification. To address a critical difficulty plaguing such undertakings involving especially protein-based nanopores isolated in lipid bilayers, namely, the formation of a stable, long-lasting single nanopore, we pioneer herein an approach for generating functional nanostructures enabling small single-stranded DNA (ssDNA) detection. We designed a dynamic hybrid construct by appending extramembrane peptide nucleic acid (PNA) segments to the C-terminus of modified ion channel-forming alamethicin monomers. We found that the resulting chimeric molecules successfully coassemble in a voltage-dependent manner in planar lipid membranes generating diameter-variable oligomers. The subsequent interaction at the flexible extramembrane segment of such formed dynamic nanopores with aqueously added complementary ssDNA fragments leads to overall conformational alterations affecting the peptide assembly state kinetics and mediated ionic current. Such recognition events were found specific to the primary structure of target ssDNA and uninhibited the presence of serum. Our platform demonstrates the feasibility of designing an entirely new class of versatile chimeric biosensors, for which, dependent upon the nature of the attached receptor moiety and underlying recognition chemistry, the applicability area may extend to other analytes.

A self-supply crRNA-mediated CRISPR/Cas12a-driven controlled-release homogeneous biosensor for ultrasensitive detection of microRNA

The abnormal expression of microRNAs (miRNAs) will lead to the occurrence of cancer and serious genetic related diseases. In this work, a novel self-supply crRNA-mediated CRISPR/Cas12a-driven controlled-release homogeneous biosensor was developed for ultra-sensitivity detection of miRNA-155, using single stranded DNA1 (ssDNA1, S1) as a bio-gate and UiO-66-NH2 as a nanocontainer. Initially, the exponential amplification reaction (EXPAR) was designed to transform and amplify the target recognition event into the T7 promoter, which could function as a trigger for transcription amplification to generate a large number of crRNA. The abundant crRNA formed a ternary complex with Cas12a and activator, and the trans-cleavage activity of the CRISPR/Cas12a was activated. Then the ssDNA2 completely complementary to ssDNA1 was cleaved, resulting in inability to form double-stranded DNA (dsDNA), which made ssDNA1 inseparable from UiO-66-NH2. Eventually, the stored methylene blue (MB) signal cannot be released from UiO-66-NH2. On the contrary, the ssDNA1 bio-gate was opened, and MB signal molecules escaped from the container, producing an obvious current signal. Through triple signal amplification and precise control and release system, the prepared biosensor could realize an ultralow detection limit of miRNA-155 as low as 31.6 aM. More importantly, the proposed strategy showed excellent sensitivity and specificity in complex serum environments and tumor cell lysates, providing a new approach to develop a robust tool for disease diagnosis.

Nucleic Acid Enzyme-Activated CRISPR-Cas12a With Circular CRISPR RNA for Biosensing.

clustered regularly interspaced short palindromic repeats (CRISPR)-Cas systems are increasingly used in biosensor development. However, directly translating recognition events for non-nucleic acid targets by CRISPR into effective measurable signals represents an important ongoing challenge. Herein, it is hypothesized and confirmed that CRISPR RNAs (crRNAs) in a circular topology efficiently render Cas12a incapable of both site-specific double-stranded DNA cutting and nonspecific single-stranded DNA trans cleavage. Importantly, it is shown that nucleic acid enzymes (NAzymes) with RNA-cleaving activity can linearize the circular crRNAs, activating CRISPR-Cas12a functions. Using ligand-responsive ribozymes and DNAzymes as molecular recognition elements, it is demonstrated that target-triggered linearization of circular crRNAs offers great versatility for biosensing. This strategy is termed as "NAzyme-Activated CRISPR-Cas12a with Circular CRISPR RNA (NA3C)." Use of NA3C for clinical evaluation of urinary tract infections using an Escherichia coli-responsive RNA-cleaving DNAzyme to test 40 patient urine samples, providing a diagnostic sensitivity of 100% and specificity of 90%, is further demonstrated.

Politics, geography and recognition in the emerging multipolar world order

ABSTRACT This article examines international politics of recognition in the increasingly multipolar global environment, with a specific focus on recognition events in Kosovo, Abkhazia and South Ossetia, Crimea, the Occupied Golan Heights, Jerusalem, Western Sahara, Luhansk and Donetsk. It reveals that the recent surge in recognition is propelled by geopolitical and geographical motivations, which undermine the normative significance of territorial integrity, non-recognition and self-determination in recognition politics. The article proposes a thesis that situates recognition within the context of geopolitics, spatial management and power dynamics that expand our understanding of the broader impacts of recognition beyond the traditional emphasis on statehood emergence and prevention. Recognition has become entangled in geopolitical contestations between influential global players seeking to normatively legitimise and normalise conquest, occupation and colonisation. In this context, geopolitical considerations have overridden the normative and legal weight of the well-institutionalised norms of territorial recognition. This underscores the gap between recognition norms and practices and demonstrates the explanatory power of geopolitics.