Background Signal Research Articles

High-efficiency electrochemiluminescence of host-guest ligand-assembled gold nanoclusters preoxidated for ultrasensitive biosensing of protein

Herein, host–guest ligand-assembled gold nanoclusters (AuNCs) with highly efficient electrochemiluminescence (ECL) as an emitter and a programable tripedal deoxyribonucleic acid (DNA) walker with a well-regulated DNA track as the signal amplifier were used to develop an ultrasensitive biosensing platform for determining soluble urokinase-type plasminogen activator receptor (suPAR) related to chronic kidney disease. Notably, the host–guest ligand-assembled AuNCs with 6-aza-2-thiothymine (ATT) as the ligand and guanidine butyric acid (Gba) as the model (Gba/ATT-AuNCs) exhibited a strong and stable ECL signal under preoxidation, 25.6 times higher than that of traditional ATT-protected AuNCs (ATT-AuNCs). This is because of the dual-enhancement ECL mechanism of the suppression of nonradiative transitions by host–guest recognition and the improvement of the electrochemical redox rate by preoxidation. Furthermore, the trace target protein suPAR could be efficiently converted into a tripedal DNA walker to walk along a well-regulated square DNA orbit with high movement efficiency and low background signal, thereby considerably improving detection sensitivity. Thus, the established ECL sensing platform achieved the ultrasensitive detection of suPAR with a linear range from 10 fg/mL to 10 ng/mL and a limit of detection of 3.15 fg/mL, whose detection sensitivity was five orders of magnitude higher than that of the gold-standard enzyme-linked immunosorbent assay with the lowest detection concentration of 1 ng/mL. This strategy provides a novel enhanced ECL mode of AuNCs for constructing a sensitive biosensing platform and is expected to be extended to the trace detection of other protein markers in human body fluids for disease diagnosis.

Label-free photoacoustic computed tomography of visually evoked responses in the primary visual cortex and four subcortical retinorecipient nuclei of anesthetized mice.

Many techniques exist for screening retinal phenotypes in mouse models in vision research, but significant challenges remain for efficiently probing higher visual centers of the brain. Photoacoustic computed tomography (PACT), with optical sensitivity to hemodynamic response (HR) in brain and ultrasound resolution, provides unique advantages in comprehensively assessing higher visual function in the mouse brain. We aim to examine the reliability of PACT in the functional phenotyping of mouse models for vision research. A PACT-ultrasound (US) parallel imaging system was established with a one-dimensional (1D) US transducer array and a tunable laser. Imaging was performed at three coronal planes of the brain, covering the primary visual cortex and the four subcortical nuclei, including the superior colliculus, the dorsal lateral geniculate nucleus, the suprachiasmatic nucleus, and the olivary pretectal nucleus. The visual-evoked HR was isolated from background signals using an impulse-based data processing protocol. rd1 mice with rod/cone degeneration, melanopsin-knockout (mel-KO) mice with photoreceptive ganglion cells that lack intrinsic photosensitivity, and wild-type mice as controls were imaged. The quantitative characteristics of the visual-evoked HR were compared. Quantitative analysis of the HRs shows significant differences among the three mouse strains: (1)rd1 mice showed both smaller and slower responses compared with wild type ( , ) and (2)mel-KO mice had lower amplitude but not significantly delayed photoresponses than wild-type mice ( , ). These results agree with the known visual deficits of the mouse strains. PACT demonstrated sufficient sensitivity to detecting post-retinal functional deficits.

Ultrasensitive detection of mycotoxins using a novel single-Atom, CRISPR/Cas12a-Based nanozymatic colorimetric biosensor

Aflatoxin B1 (AFB1) is a typical mycotoxin in cereals, posing a significant threat to the ecology and public health. Overall, the development of nanozymatic sensors with good specificity, high sensitivity, and a clear catalytic mechanism seems to be a challenge for CRISPR/Cas detection of non-nucleic acid targets. In this study, a series of Fe-N-C single-atom enzymes (SAzymes) and Fe-Co magnetic nanoparticles (MNPs) were synthesized via pyrolysis. Fe-N-C SAzymes with high peroxidase-mimetic activity were successfully utilized as TMB chromogenic catalysts for detecting AFB1 via the CRISPR/Cas12a colorimetric aptamer sensor. In the absence of the AFB1 target, its aptamer bound to crRNA and activated the trans-cleavage activity of Cas12a, indiscriminately cutting single-stranded DNA and releasing significant amounts of Fe-N-C SAzymes into the supernatant, which catalyzed the change of TMB to blue. The synthesized Fe-Co MNPs not only effectively reduced the background signal but also significantly reduced the assay cost. This study combined SAzymes with CRISPR/Cas12a technology for the first time for non-nucleic acid target detection. The combination of Fe-N-C SAzymes’ high catalytic activity and stability, and the CRISPR/Cas12a system’s high specificity and signal amplification, significantly improved the detection system’s sensitivity and specificity, enabling the detection of other targets by substituting the aptamers. Additionally, the potential active sites and catalytic mechanism of Fe-N-C SAzymes were investigated using extended X-ray absorption fine structure (EXAFS) spectroscopy and density functional theory (DFT) simulations. This scheme provides a general method for constructing high-density Fe-N-C SAzymes and offers both experimental and theoretical guidance for understanding their catalytic mechanism.

Ultrasensitive carbon nanotube-bridged MXene conductive network arrays for one-step homogeneous electrochemical immunosensing of tumor markers

Developing non-passivating and fully integrated electrode arrays for point-of-care testing of carcinoembryonic antigen (CEA) is crucial, as the serum level of CEA is closely associated with colorectal cancer. Herein, we propose a simple, low-cost, and eco-friendly template-assisted filtration method for the scalable preparation of carbon nanotube-bridged Ti3C2Tx MXene (MX@CNT) electrode arrays with a conductive network. Furthermore, we fabricate a homogeneous electrochemical (HEC) sensor for CEA detection by integrating a magnetic-bead-based alkaline phosphatase-linked immunoassay (MB-aElisa), which enables the in-situ generation of the electroactive substance 1-naphthol (1-NP). Benefiting from the unique electrochemical characteristics of a MX@CNT electrode array, such as ultra-low background signal and superior electrocatalytic activity towards the hydrolyzed 1-NP, the MB-aElisa-based HEC sensor specifically measures CEA within a detection range spanning from 0.005 to 1.0 ng mL−1, achieving a detection limit of 1.6 pg mL−1. Subsequently, this biosensing prototype is successfully utilized for the detection of CEA in serum specimens obtained from colorectal cancer patients. More importantly, the integration of MB-aElisa with a MX@CNT electrode array not only marks a significant advancement but also enables the creation of a one-step homogeneous electrochemical immunosensing platform, serving as a paradigm for the highly sensitive and selective measurement of trace tumor markers in complex biological samples.

Optogenetic Strategies for Optimizing the Performance of Phospholipids Biosensors.

High-performance biosensors play a crucial role in elucidating the intricate spatiotemporal regulatory roles and dynamics of membrane phospholipids. However, enhancing the sensitivity and imaging performance remains a significant challenge. Here, optogenetic-based strategies are presented to optimize phospholipid biosensors. These strategies involves presequestering unbound biosensors in the cell nucleus and regulating their cytosolic levels with blue light to minimize background signal interference in phospholipid detection, particularly under conditions of high expression levels of biosensor. Furthermore, optically controlled phase separation and the SunTag system are employed to generate punctate probes for substrate detection, thereby amplifying biosensor signals and enhancing visualization of the detection process. These improved phospholipid biosensors hold great potential for enhancing the understanding of the spatiotemporal dynamics and regulatory roles of membrane lipids in live cells and the methodological insights in this study might be valuable for developing other high-performance biosensors.

A Deep Learning Lidar Denoising Approach for Improving Atmospheric Feature Detection

Space-based atmospheric backscatter lidars provide critical information about the vertical distribution of clouds and aerosols, thereby improving our understanding of the climate system. They are additionally useful for detecting hazards to aviation and human health, such as volcanic plumes and man-made pollution events. The Cloud-Aerosol Lidar with Orthogonal Polarization (CALIOP, 2006–2023), Cloud-Aerosol Transport System (CATS, 2015–2017), and Advanced Topographic Laser Altimeter System (ATLAS 2018–present) are three such lidars that operated within the past 20 years. The signal-to-noise ratio (SNR) for these lidars is significantly lower in daytime data compared with nighttime data due to the solar background signal increasing the detector response noise. Averaging horizontally across profiles has been the standard way to increase SNR, but this comes at the expense of resolution. Modern, deep learning-based denoising algorithms can be applied to improve the SNR without coarsening resolution. This paper describes how one such model architecture, Dense Dense U-Net (DDUNet), was trained to denoise CATS 1064 nm raw signal data (photon counts) using artificially noised nighttime data. Simulated CATS daytime 1064 nm data were then created to assess the model’s performance. The denoised simulated data increased the daytime SNR by a factor of 2.5 (on average) and decreased minimum detectable backscatter (MDB) to ~7.3×10−4 km−1sr−1, which is lower than the CALIOP 1064 nm night MDB value of 8.6×10−4 km−1sr−1. Layer detection was performed on simulated 2 km horizontal resolution denoised and 60 km averaged data. Despite the finer resolution input, the denoised layers had more true positives, fewer false positives, and an overall Jaccard Index of 0.54 versus 0.44 when compared to the layers detected on averaged data. Layer detection was also performed on a full month of denoised daytime CATS data (Aug. 2015) to detect layers for comparison with CATS standard Level 2 (L2) product layers. The detection on the denoised data yielded 2.33 times more, higher-quality bins within detected layers at 2.7–33 times finer resolution than the CATS L2 products.

PAIT effect: Padlock activator inhibits the trans-cleavage activity of CRISPR/Cas12a

The CRISPR/Cas12a system is increasingly used in biosensor development. However, high background signal and low sensitivity for the non-nucleic acid targets detection is challenging. Here, a padlock activator which could inhibit the trans-cleavage activity of CRISPR/Cas12a system in the intact form by steric hindrance effect (PAIT effect) was designed for non-nucleic acid targets detection. The PAIT effect disappeared when padlock activator was separated into two split activators. To verify the feasibility of padlock activator, a Ca2+ sensor was developed based on PAIT effect with the assistance of DNAzyme, activity of which was Ca2+ dependent. In the presence of Ca2+, DNAzyme was activated to cleave its substrate, a padlock activator modified with adenine ribonucleotide, into split padlock activators which would trigger the trans-cleavage activity of Cas12a to generate fluorescence. There was a mathematical relationship between the fluorescence intensity and the logarithm of Ca2+ concentration ranging from 10 pM to 1 nM, with a limit of detection of 3.98 pM. The little interference of Mg2+, Mn2+, Cd2+, Cu2+, Na+, Al3+, K+, Fe2+, and Fe3+ indicated high selectivity. Recovery ranged from 93.32% to 103.28% with RSDs from 1.87% to 12.74% showed a good accuracy and reliability. Furthermore, the proposed sensor could be applied to detect Ca2+ in mineral water, milk powder and urine. The results were consistent with that of flame atomic absorption spectroscopy. Thus, PAIT effect is valuable for expanding the application boundary of CRISPR/Cas12a system.

Single-Shot MRI in parahydrogen hyperpolarized samples

The site-specific signal enhancement provided by parahydrogen induced polarization (PHIP) may be combined with magnetic resonance imaging (MRI) to study chemical and biomolecular processes. However, imaging of hydrogen nuclei (1H) is hampered by background signals arising from the presence of thermally polarized nuclei. Additionally, fast imaging sequences are commonly based on multiple radio-frequency pulses, where the signals resulting from PHIP oscillate due to the evolution with a J-coupling Hamiltonian. In this article, an innovative imaging scheme for single-scan MRI is presented that effectively detects hyperpolarized components while simultaneously canceling out thermal contributions. This method is based on the quenching of inherent oscillations of PHIP-originated signals due to J-couplings during the multipulse sequence and the suppression of thermal signals by spin dynamics and a tailored restructuring of the k-space. A series of numerical simulations on specific two- and three-spin systems serve to support the feasibility of the approach. Furthermore, this theoretical study demonstrates the potential of combining hyperpolarization and long-lived states (PHIP and LLS) in the selected molecules, which could be seen as a preliminary step towards the development of fast imaging techniques, for example in the field of biomolecular research.

Probing flavor violation and baryogenesis via primordial gravitational waves

We show that observations of primordial gravitational waves of inflationary origin can shed light into the scale of flavor violation in a flavon model which also explains the mass hierarchy of fermions. The energy density stored in oscillations of the flavon field around the minimum of its potential redshifts as matter and is expected to dominate over radiation in the early universe. At the same time, the evolution of primordial gravitational waves acts as bookkeeping to understand the expansion history of the universe. Importantly, the gravitational wave spectrum is different if there is an early flavon dominated era compared to radiation domination expected from a standard cosmological model and this spectrum gets damped by the entropy released in flavon decays, determined by the mass of the flavon field mS and new scale of flavor violation ΛFV. We derive analytical expressions of the frequency above which the spectrum is damped, as-well-as the amount of damping, in terms of mS and ΛFV. We show that the damping of the gravitational wave spectrum would be detectable at BBO, DECIGO, U-DECIGO, μ−ARES, LISA, CE and ET detectors for ΛFV = 105−10 GeV and mS = OTeV\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$ \\mathcal{O}\\left(\ extrm{TeV}\\right) $$\\end{document}. Furthermore, the flavon decays can source the baryon asymmetry of the universe. We identify the mS – ΛFV parameter space where the observed baryon asymmetry η ~ 10−10 is produced and can be tested by gravitational wave detectors like LISA and ET. We also discuss our results in the context of the recently measured stochastic gravitational background signals by NANOGrav.

Hexagonal Plasmonic Arrays for High-Throughput Multicolor Single-Molecule Studies.

Nanophotonic biosensors offer exceptional sensitivity in the presence of strong background signals by enhancing and confining light in subwavelength volumes. In the field of nanophotonic biosensors, antenna-in-box (AiB) designs consisting of a nanoantenna within a nanoaperture have demonstrated remarkable single-molecule fluorescence detection sensitivities under physiologically relevant conditions. However, their full potential has not yet been exploited as current designs prohibit insightful correlative multicolor single-molecule studies and are limited in terms of throughput. Here, we overcome these constraints by introducing aluminum-based hexagonal close-packed AiB (HCP-AiB) arrays. Our approach enables the parallel readout of over 1000 HCP-AiBs with multicolor single-molecule sensitivity up to micromolar concentrations using an alternating three-color excitation scheme and epi-fluorescence detection. Notably, the high-density HCP-AiB arrays not only enable high-throughput studies at micromolar concentrations but also offer high single-molecule detection probabilities in the nanomolar range. We demonstrate that robust and alignment-free correlative multicolor studies are possible using optical fiducial markers even when imaging in the low millisecond range. These advancements pave the way for the use of HCP-AiB arrays as biosensor architectures for high-throughput multicolor studies on single-molecule dynamics.

Enzymatic cascade reactors on carbon nanotube transistor detecting trace prostate cancer biomarker

Biosensors based on carbon nanotube field-effect transistors (CNT-FETs) have shown great potential in biomarker detection due to their high sensitivity because of appreciable semiconducting electrical properties. However, background signal interferences in complex mediums may results in low signal-to-noise ratio, which may impose challenges for precise biomarker detection in physiological fluids. In this work, we develop an enzymatic CNT-FET, with scalable production at wafer scale, for detection of trace sarcosine that is a biopsy-correlated biomarker of prostate cancer. Enzymatic cascade rectors are constructed on the CNT to improve the reaction efficiency, thereby, enhancing the signal transduction. As such, a limit of detection as low as 105 zM is achieved in buffer solution. Owing to the enhanced reaction efficiency, the testing of clinical serum samples yields significant signal difference to discriminate the prostate cancer (PCa) samples from the benign prostatic hyperplasia (BPH) samples (P = 1.07 × 10−5), demonstrating immense potential in practical applications.

Light-Driven Electrochemical Biosensing with DNA Origami-Assisted Hybrid Nanoantenna for Fumonisin B1 Monitoring.

The electrochemical detection of biosensors is largely governed by the changes in physical properties of redox probes, which are susceptible to electrode substrate effects, inhibiting sensor sensitivity. In this work, a light-driven electrochemical biosensor based on a hybrid nanoantenna was developed for the sensitive detection of fumonisin B1 (FB1). The hybrid nanoantenna sensing interface was constructed by coupling CdSe quantum dots (QDs)-DNA nanowire and graphdiyne oxide composites loaded with methylene blue and gold nanorods (GDYO-MB-Au NRs) using a tetrahedral DNA nanostructure, which acted as a light-driven unit and an amplification unit, respectively. The hybrid nanoantenna with light-driven properties facilitated the alteration in the chemical properties of MB at the sensing interface; that is, MB was degraded under light illumination. The stripping of the CdSe QDs-DNA nanowire triggered by the binding of FB1 could degrade the light-driven capability, thereby improving the electrochemical signal through depressing MB degradation. Taking advantage of the photodegradation of MB by the hybrid nanoantenna, the developed biosensor reduced the background signal and increased the detection sensitivity. The developed biosensor exhibited a linear detection range from 0.5 fg mL-1 to 10 pg mL-1 and a detection limit down to 0.45 fg mL-1. This strategy shows great promise for the fabrication of highly sensitive electrochemical biosensors.

A "Dual-Key-and-Lock" MRI Contrast Agent with T1-T2 Switchable Function for Accurate Diagnosis of Tumors.

Extremely small iron oxide nanoparticle (ESIONP)-based stimuli-responsive switchable MRI contrast agents (CAs) show great promise for accurate detection of tumors due to their outstanding advantages of high specificity and low background signal. However, currently developed ESIONP-based switchable CAs often suffer single-biomarker-induced responses, which lack absolute specificity to pathological tissues, potentially diminishing diagnostic accuracy. In this study, weak acidity and hypoxia, two of the most remarkable characteristics of tumors, are introduced as dual biomarker stimuli to construct an ESIONP-based switchable MRI CA (DKL-CA), with its signal switch controlled by a "dual-key-and-lock" strategy. Only when DKL-CA is exposed to a coexisting weakly acidic and hypoxic environment can monodispersed ESIONPs form nanoclusters, thereby realizing a switch from the T1 to T2 contrast. Moreover, DKL-CA exhibits favorable biosafety and the capacity for precise tumor diagnosis in tumor-bearing mice. Overall, DKL-CA paves the way for designing highly accurate ESIONP-based MRI CAs for tumor diagnosis.

Development of a Colorimetric and SERS Dual-Signal Platform via dCas9-Mediated Chain Assembly of Bifunctional Au@Pt Nanozymes for Ultrasensitive and Robust Salmonella Assay.

Timely screening for harmful pathogens is a great challenge in emergencies where traditional culture methods suffer from long assay time and alternative methods are limited by poor accuracy and low robustness. Herein, we present a dCas9-mediated colorimetric and surface-enhanced Raman scattering (SERS) dual-signal platform (dCas9-CSD) to address this challenge. Strategically, the platform used dCas9 to accurately recognize the repetitive sequences in amplicons produced by loop-mediated isothermal amplification (LAMP), forming nucleic acid frameworks that assemble numerous bifunctional gold-platinum (Au@Pt) nanozymes into chains on the surface of streptavidin-magnetic beads (SA-MB). The collected Au@Pt converted colorless 3,3',5,5'-tetramethylbenzidine (TMB) to blue oxidized TMB (oxTMB) via its Pt shell and then enhanced the Raman signal of oxTMB by its Au core. Therefore, the presence of Salmonella could be dexterously converted into cross-validated colorimetric and SERS signals, providing more reliable conclusions. Notably, dCas9-mediated secondary recognition of amplicons reduced background signal caused by nontarget amplification, and two-round signal amplification consisting of LAMP reaction and Au@Pt catalysis greatly improved the sensitivity. With this design, Salmonella as low as 1 CFU/mL could be detected within 50 min by colorimetric and SERS modes. The robustness of dCas9-CSD was further confirmed by various real samples such as lake water, cabbage, milk, orange juice, beer, and eggs. This work provides a promising point-of-need tool for pathogen detection.

Ternary electrochemiluminescence biosensor based on Ce2Sn2O7@MSN luminophore and Au@NiO-CeO2 as a co-reaction accelerator for the detection of prostate specific antigen

A ternary ECL system was constructed with Ce2Sn2O7@MSN composite as luminophore, potassium persulfate as co-reactant and Au@NiO-CeO2 as co-reaction accelerator to realize ultra-sensitive biosensing detection of prostate specific antigen (PSA). The abundant oxygen vacancies in Ce2Sn2O7 endow it with powerful electrochemical redox ability, accelerate energy transfer, and ensure Ce2Sn2O7 excellent electrochemical luminescence performance as a luminophore. Furthermore, to optimize the luminescence energy, enhance stability, and reduce the background signal to improve its signal-to-noise ratio, a highly selective Ce2Sn2O7@MSN biological probe was obtained by coating cerium stannate (Ce2Sn2O7) with mesoporous silica nanospheres (MSN). Au@NiO-CeO2 with a large specific surface area and high oxygen vacancy activity was synthesized as the co-reaction accelerator, which promoted the generation of SO4•− through the rapid reversible oxidation and reduction of Ce4+/Ce3+, thereby the ECL signal amplified effectively. Under optimal conditions, the linear detection range of PSA spans from 100fg mL−1 to 100 ng mL−1, with a remarkable lowest detection limit of 0.037pg mL−1, showcasing significant application potential. This study provides a valuable reference for pyrochlore to be used as luminophore in the detection of various biomarkers by ECL biosensing.

High‐Efficiency Multilevel Phase Lenses with Nanostructures on Polyimide Membranes

AbstractThe emergence of planar meta‐lenses on flexible materials has profoundly impacted the long‐standing perception of diffractive optics. Despite their advantages, these lenses still face challenges in design and fabrication to obtain high focusing efficiency and resolving power. A nanofabrication technique is demonstrated based on photolithography and polyimide casting for realizing membrane‐based multilevel phase‐type Fresnel zone plates (FZPs) with high focusing efficiency. By employing advantageous techniques, these lenses with nanostructures are directly patterned into thin polyimide membranes. The computational and experimental results have indicated that the focusing efficiency of these nanostructures at the primary focus increases significantly with increasing the number of phase levels. Specifically, 16‐level phase lenses on a polyimide membrane can achieve a focusing efficiency of more than 91.6% of the input signal (9.5 times better than that of a conventional amplitude‐type FZP) and focus light into a diffraction‐limited spot together with very weak side‐lobes. Furthermore, these lenses exhibit considerably reduced unwanted diffraction orders and produce extremely low background signals. The potential impact of these lenses extends across various applications and techniques including microscopy, imaging, micro‐diffraction, remote sensing, and space flight instruments which require lightweight and flexible configurations.

Inversion of Underground Target by Satellite Gravity Gradient Signal Considering Terrain Interference

This paper explores using satellite gravity gradiometry to detect underground targets. It discusses measured signals including target body, terrain interference, and global background signals. Formulas and principles for calculating these signals are provided. Simulation results show that a 5 million cubic meter sphere generates a gravity gradient signal of to at 200 km altitude. Terrain interference signals are around for a 100 km × 100 km area. The paper presents a parametric inversion method based on full tensor gravity gradient signals for target detection. Inversion experiments consider noise and terrain deduction, with 100 iterations for error analysis.

Acid Treatment of FVIII-Containing Plasma Samples Unmasks a Broad Spectrum of FVIII-Specific Antibodies in ELISA.

During routine treatment, plasma samples of patients with hemophilia A or acquired hemophilia A are frequently analyzed for the presence of FVIII-specific antibodies. While only inhibitory antibodies can be detected by the Bethesda assay, inhibitory and non-inhibitory antibodies can be detected by ELISA. However, plasma samples of patients frequently contain endogenous or substituted FVIII, hence interfering with both types of analyses. One option for the inactivation of FVIII is heat denaturation, which unfortunately has been shown to lead to high background signals complicating the discrimination of negative and positive plasma samples. In the current study, we developed a method of acid denaturation for FVIII-containing plasma samples that can help identify samples containing FVIII-specific antibodies and compared the effects of heat and acid denaturation on the detection of FVIII-antibody interactions in a monoclonal setting. The aim of our study was to establish an analysis that allows safer treatment decisions in the context of tolerance to FVIII.

Near-infrared II fluorescence-guided glioblastoma surgery targeting monocarboxylate transporter 4 combined with photothermal therapy

Surgery is crucial for glioma treatment, but achieving complete tumour removal remains challenging. We evaluated the effectiveness of a probe targeting monocarboxylate transporter 4 (MCT4) in recognising gliomas, and of near-infrared window II (NIR-II) fluorescent molecular imaging and photothermal therapy as treatment strategies. We combined an MCT4-specific monoclonal antibody with indocyanine green to create the probe. An orthotopic mouse model and a transwell model were used to evaluate its ability to guide tumour resection using NIR-II fluorescence and to penetrate the blood-brain barrier (BBB), respectively. A subcutaneous tumour model was established to confirm photothermal therapy efficacy. Probe specificity was assessed in brain tissue from mice and humans. Finally, probe effectiveness in photothermal therapy was investigated. MCT4 was differentially expressed in tumour and normal brain tissue. The designed probe exhibited precise tumour targeting. Tumour imaging was precise, with a signal-to-background (SBR) ratio of 2.8. Residual tumour cells were absent from brain tissue postoperatively (SBR: 6.3). The probe exhibited robust penetration of the BBB. Moreover, the probe increased the tumour temperature to 50°C within 5min of laser excitation. Photothermal therapy significantly reduced tumour volume and extended survival time in mice without damage to vital organs. These findings highlight the potential efficacy of our probe for fluorescence-guided surgery and therapeutic interventions. Jilin Province Department of Science and Technology (20200403079SF), Department of Finance (2021SCZ06) and Development and Reform Commission (20200601002JC); National Natural Science Foundation of China (92059207, 92359301, 62027901, 81930053, 81227901, U21A20386); and CAS Youth Interdisciplinary Team (JCTD-2021-08).

Tuning phenolic hemicyanines for ratiometric DNA sensing and live cell nuclear bioimaging applications

Activatable fluorescent probes with turn-on emission in response to a biological target can reduce the background signal and improve the detection limit needed for biosensor and bioimaging resolution. Turn-on probes with dual emission (ratiometric probes) have further advantages, as they have self-calibration ability for more reliable detection. Herein, we demonstrate the tunability of the phenolic hemicyanine (HC) scaffold for both DNA biosensing and bioimaging applications using the quinine-binding DNA aptamer (MN4) as a host-guest biosensor platform and live ovarian cancer cells for nuclear imaging. The DNA aptamer MN4 contains a three-way junction (3WJ) consisting of a hydrophobic branch point connecting three double-stranded stems. Phenolic HCs bearing electron-withdrawing halogen substituents (FPhOBtz and Cl2PhOBTz) bind to the 3WJ of MN4 as the phenolate with strong turn-on emission. Subsequent displacement of the phenolate by the quinine target causes a turn-off emission response. In contrast, the parent phenolic HC (PhOBtz) and those bearing electron-donating groups (Me2PhOBtz and (MeO)2PhOBtz) exhibit dual (phenol and phenolate) fluorescence upon MN4 binding. Phenol excitation generates phenolate emission by an excited-state proton transfer (ESPT) process with DNA components serving as the proton acceptor. Quinine displacement of PhOBtz from the 3WJ of MN4 affords a turn-on ratiometric response with preferential light-up of phenol emission. The ability of the phenolic HCs to serve as cellular nuclear stains further highlights the tunability of the phenolic HC probes with the Cl2PhOBtz analog displaying superior staining ability. The simplicity and tunability of phenolic HCs make them attractive fluorescent probes for DNA sensing and bioimaging applications.