Proactive functional classification of all possible missense single-nucleotide variants in KCNQ4.

Clinical exome sequencing has yielded extensive disease-related missense single-nucleotide variants (SNVs) of uncertain significance, leading to diagnostic uncertainty. KCNQ4 is one of the most commonly responsible genes for autosomal dominant nonsyndromic hearing loss. According to the gnomAD cohort, approximately one in 100 people harbors missense variants in KCNQ4 (missense variants with minor allele frequency > 0.1% were excluded), but most are of unknown consequence. To prospectively characterize the function of all 4085 possible missense SNVs of human KCNQ4, we recorded the whole-cell currents using the patch-clamp technique and categorized 1068 missense SNVs as loss of function, as well as 728 loss-of-function SNVs located in the transmembrane domains. Further, to mimic the heterozygous condition in Deafness nonsyndromic autosomal dominant 2 (DFNA2) patients caused by KCNQ4 variants, we coexpressed loss-of-function variants with wild-type KCNQ4 and found 516 variants showed impaired or only partially rescued heterogeneous channel function. Overall, our functional classification is highly concordant with the auditory phenotypes in Kcnq4 mutant mice and the assessments of pathogenicity in clinical variant interpretations. Taken together, our results provide strong functional evidence to support the pathogenicity classification of newly discovered KCNQ4 missense variants in clinical genetic testing.

Chiral CuxCoyS-CuzS Nanoflowers with Bioinspired Enantioselective Catalytic Performances.

Nanomaterials with biomimetic catalytic abilities have attracted significant attention. However, the stereoselectivity of natural enzymes determined by their unique configurations is difficult to imitate. In this work, a kind of chiral CuxCoyS-CuzS nanoflowers (L/D-Pen-NFs) is developed, using porous CuxCoyS nanoparticles (NPs) as stamens, CuzS sheets as petals, and chiral penicillamine as surface stabilizers. Compared to the natural laccase enzyme, L/D-Pen-NFs exhibit significant advantages in catalytic efficiency, stability against harsh environments, recyclability, and convenience in construction. Most importantly, they display high enantioselectivity toward chiral neurotransmitters, which is proved by L- and D-Pen-NFs' different catalytic efficiencies toward chiral enantiomers. L-Pen-NFs are more efficient in catalyzing the oxidation of L-epinephrine and L-dopamine compared with D-Pen-NFs. However, their catalytic efficiency in oxidizing L-norepinephrine and L-DOPA is lower than that of D-Pen-NFs. The reason for the difference in catalytic efficiency is the distinct binding affinities between CuxCoyS-CuzS nano-enantiomers and chiral molecules. This work can spur the development of chiral nanostructures with biomimetic functions.

Carbon Dots with Integrated Photothermal Antibacterial and Heat-Enhanced Antioxidant Properties for Diabetic Wound Healing.

Diabetic wounds pose a persistent challenge due to their slow healing nature, primarily caused by bacterial infection and excessive reactive oxygen species (ROS)-induced inflammation. In this study, carbon dots with synergistic antibacterial and antioxidant properties, referred to as AA-CDs, are developed specifically for diabetic wound healing using a straightforward solvothermal method. By utilizing cost-effective precursors like citric acid and ascorbic acid, AA-CDs are engineered to possess tailored functions of photothermal sterilization and ROS scavenging. The resulting AA-CDs demonstrats broad-spectrum antibacterial activity, particularly against multidrug-resistant strains, along with efficient ROS scavenging both in solution and within cells. Additionally, AA-CDs exhibits a protective effect against oxidative stress-induced damage. Notably, with a high photothermal conversion efficiency (41.18%), AA-CDs displays heat-enhanced antioxidant performance, providing not only augmented ROS scavenging but also additional protection against oxidative stress, yielding a true "1 + 1 > 2" effect. To facilitate their use in vivo, AA-CDs are incorporated into a thermally responsive hydrogel, which exhibits evident anti-inflammatory properties by modulating inflammatory factors and significantly promots the healing of diabetic wounds. This study underscores the value of integrated platforms for diabetic wound healing and highlights the potential of versatile CDs as promising therapeutic agents in biomedical applications.

Modulation by Co (II) Ion of Optical Activities of L/D-glutathione (GSH)-modified Chiral Copper Nanoclusters for Sensitive Adenosine Triphosphate Detection.

Chiral nanoscale enantiomers exhibit different biological effects in living systems. However, their chirality effect on the detection sensitivity for chiral biological targets still needs to be explored. Here, we discovered that Co2+ can modulate the luminescence performance of L/D-glutathione (GSH)-modified copper nanoclusters (L/D-Cu NCs) and induce strong chiroptical activities as the asymmetric factor was enhanced 223-fold with their distribution regulating from the ultraviolet to visible region. One Co2+ coordinated with two GSH molecules that modified on the surface of Cu NCs in the way of CoN2O2. On this basis, dual-modal chiral and luminescent signals of Co2+ coordinated L/D-Cu NCs (L/D-Co-Cu NCs) were used to detect the chiral adenosine triphosphate (ATP) based on the competitive interaction between surficial GSH and ATP molecules with Co2+. The limits of detection of ATP obtained with fluorescence and circular dichroism intensity were 9.15 µM and 15.75 nM for L-Co-Cu NCs, and 5.35 µM and 4.69 nM for D-Co-Cu NCs. This demonstrated that selecting suitable chiral configurations of nanoprobes effectively enhances detection sensitivity. This study presents not only a novel method to modulate and enhance the chiroptical activity of nanomaterials but also a unique perspective of chirality effects on the detection performances for bio-targets.

Specific Chemiluminescence Imaging and Enhanced Photodynamic Therapy of Bacterial Infections by Hemin-Modified Carbon Dots.

Antibacterial photodynamic therapy (aPDT) is a promising antibiotics-alternative strategy for bacterial infectious diseases, which features broad-spectrum antibacterial activity with a low risk of inducing bacterial resistance. However, clinical applications of aPDT are still hindered by the hydrophobicity-caused inadequate photodynamic activity of conventional photosensitizers and the hypoxic microenvironment of bacterial infections. To address these problems, herein, a promising strategy is developed to achieve specific chemiluminescence (CL) imaging and enhanced PDT of bacterial infections using hemin-modified carbon dots (H-CDs). The H-CDs can be facilely prepared and exhibit favorable water solubility, augmented photodynamic activity, and unique peroxidase-mimicking capacity. Compared with the free CDs, the photodynamic efficacy of H-CDs is significantly augmented due to the increased electron-hole separation efficiency. Moreover, the peroxidase catalytic performance of H-CDs enables not only infection identification via bacterial infection microenvironment-responsive CL imaging but also oxygen self-supplied aPDT with hypoxia-relief-enhanced bacteria inactivation effects. Finally, the enhanced aPDT efficiencies of H-CDs are validated in both in vivo abscess and infected wound models. This work may provide an effective antibacterial platform for the selective imaging-guided treatment of bacterial infections.

Enantiomeric NIR-II Emitting Rare-Earth-Doped Ag2 Se Nanoparticles with Differentiated In Vivo Imaging Efficiencies.

Here, chiral second near-infrared (NIR-II) emitting rare-earth doped silver selenide nanoparticles (R- or S-Ag2 Se:Nd/Yd/Er NPs) were fabricated, exhibiting circular dichroism peak at 850 nm and fluorescence peak at 1550 nm, with 145.7-fold enhanced intensity compared to the reported Ag2 Se NPs. Compared with S-Ag2 Se:Nd/Yd/Er NPs, imaging efficiency of R-Ag2 Se:Nd/Yd/Er NPs in living cells was significantly improved due to a higher cellular uptake rate and 927.7-fold higher affinity. Furthermore, R-Ag2 Se:Nd/Yd/Er NPs reached at the tumor 2-fold faster than S type of NPs in vivo. We discover that chirality leads to differences in the affinity between chiral Ag2 Se:Nd/Yd/Er NPs and cluster of differentiation 44 (CD44) onto the surface of murine mammary carcinoma cell to cause different in vivo imaging efficiency. These results reveal that chiral Ag2 Se:Nd/Yd/Er NPs have high photoluminescence intensity and high in vivo imaging efficiency reflecting wide applications in biomedical diagnosis.

Photo-Stimulated Polychromatic Room Temperature Phosphorescence of Carbon Dots.

Single-component multicolor luminescence, particularly phosphorescence materials are highly attractive both in numerous applications and in-depth understanding the light-emission processes, but formidable challenges still exist for preparing such materials. Herein, a very facile approach is reported to synthesize carbon dots (CDs) (named MP-CDs) that exhibit multicolor fluorescence (FL), and more remarkably, multicolor long-lived room temperature phosphorescence (RTP) under ambient conditions. The FL and RTP colors of the CDs powder are observed to change from blue to green and cyan to yellow, respectively, with the excitation wavelength shifting from 254 to 420 nm. Further studies demonstrate that the multicolor emissions can be attributed to the existence of multiple emitting centers in the CDs and the relatively higher reaction temperature plays a critical role for achieving RTP. Given the unique optical properties, a preliminary application of MP-CDs in advanced anti-counterfeiting is presented. This study not only proposes a strategy to prepare photo-stimulated multicolor RTP materials, but also reveals great potentials of CDs in exploiting novel optical materials with unique properties.

A colorimetric sensor array for the discrimination of Chinese liquors.

Although some colorimetric sensor arrays have been developed for the identification of Chinese liquors, they usually require the confirmation of volatile markers in the liquors by chromatography and mass spectrometry firstly. Herein, we present a simple colorimetric sensor array to identify various Chinese liquors in the liquid phase without the aid of other analytical techniques. The colorimetric sensor array consists of six commercially available and inexpensive solvatochromic dyes, and the sensing mechanism of this array is based on the response of solvatochromic dyes to their local polarity. On the basis of the colour changes of the sensor array, different Chinese liquors are discerned readily using pattern recognition methods, and the statistical analysis results (i.e., hierarchical clustering analysis and principal component analysis) reveal that the as-fabricated sensor array can distinguish the subtle differences between different liquors from the same winery and the same flavor type. Moreover, the developed sensor array can even distinguish diverse diluted liquors from the pristine ones.

Disulfide bond-based self-crosslinked carbon-dots for turn-on fluorescence imaging of GSH in living cells.

Herein, we report a turn-on fluorimetric nanoprobe for intracellular glutathione (GSH) imaging. The principle of this probe is designed on the basis of the selective reduction between GSH and disulfide bond-based self-crosslinked red emissive carbon dots (abbreviated as SCCDs). The nanoprobe (i.e., SCCDs) was facilely fabricated from thiol-modified carbon dots (CDs) through oxidation in the presence of H2O2, and its fluorescence was greatly reduced due to the effect of aggregation induced quenching (AIQ). However, in the presence of GSH, the SCCDs were separated into many single CDs. As a result, the fluorescence of the nanoprobe was recovered in a GSH concentration-dependent manner, which is the basis for the quantitative analysis of GSH. The nanoprobe shows excellent specificity and a linear range from 0 to 0.15 mM towards GSH with a limit of detection (LOD) of 5.7 µM. Finally, the nanoprobe was demonstrated to have extremely low cytotoxicity, and was successfully applied for monitoring the GSH level in living cells. This work would provide a promising probe for the research of GSH in cytobiology.

Carbon Dots with Dual-Emissive, Robust, and Aggregation-Induced Room-Temperature Phosphorescence Characteristics.

Carbon dots (CDs) with dual-emissive, robust, and aggregation-induced RTP characteristics are reported for the first time. The TA-CDs are prepared via hydrothermal treatment of trimellitic acid and exhibit unique white prompt and yellow RTP emissions in solid state under UV excitation (365 nm) on and off, respectively. The yellow RTP emission of TA-CDs powder should be resulted from the formation of a new excited triplet state due to their aggregation, and the white prompt emission is due to their blue fluorescence and yellow RTP dual-emissive nature. The RTP emission of TA-CDs powder was highly stable under grinding, which is very rare amongst traditional pure organic RTP materials. To employ the unique characteristics of TA-CDs, advanced anti-counterfeiting and information encryption methodologies (water-stimuli-response producing RTP) were preliminarily investigated.

Tumor Microenvironment Stimuli-Responsive Fluorescence Imaging and Synergistic Cancer Therapy by Carbon-Dot-Cu2+ Nanoassemblies.

A method is developed to fabricate tumor microenvironment (TME) stimuli-responsive nanoplatform for fluorescence (FL) imaging and synergistic cancer therapy via assembling photosensitizer (chlorine e6, Ce6) modified carbon dots (CDs-Ce6) and Cu2+ . The as-obtained nanoassemblies (named Cu/CC nanoparticles, NPs) exhibit quenched FL and photosensitization due to the aggregation of CDs-Ce6. Their FL imaging and photodynamic therapy (PDT) functions are recovered efficiently once they entering tumor sites by the stimulation of TME. Introducing of Cu2+ not only provides extra chemodynamic therapy (CDT) function through reaction with hydrogen peroxide (H2 O2 ), but also depletes GSH in tumors by a redox reaction, thus amplifying the intracellular oxidative stress and enhancing the efficacy of reactive oxygen species (ROS) based therapy. Cu/CC NPs can act as a FL imaging guided trimodal synergistic cancer treatment agent by photothermal therapy (PTT), PDT, and thermally amplified CDT.

A conjugated carbon-dot-tyrosinase bioprobe for highly selective and sensitive detection of dopamine.

In this work, a bioprobe for the detection of dopamine was designed and fabricated through covalently linking fluorescent carbon dots (CDs) and tyrosinase (TYR). The bioprobe (named CDs-TYR) can catalyze oxidation of dopamine and produce dopaquinone, and consequently the fluorescence of the CDs was quenched due to an efficient electron transfer mechanism from excited CDs to dopaquinone. The fluorescence intensity of CDs decreased in a dopamine-concentration-dependent manner, which built the foundation of dopamine quantification. The bioprobe provided a wide linear range from 0.1 to 6.0 µM for dopamine sensing. Additionally, excellent selectivity of the bioprobe to dopamine was achieved because of the specific catalytic character of the conjugated TYR. Furthermore, the bioprobe was successfully employed for the detection of dopamine in spiked human serum. To the best of our knowledge, this is the first example of the construction of a bioprobe through conjugating CDs and an enzyme. This work would open new opportunities to develop CD-based photoinduced electron transfer bioprobes for other analytes via linking typical enzymes onto CDs.

Carbon-Dots-Based Lab-On-a-Nanoparticle Approach for the Detection and Differentiation of Antibiotics.

Fluorescent carbon dots (CDs) have received considerable attention in recent years due to their superior optical properties. To take further advantages of these unique features, herein, a CDs-based "lab-on-a-nanoparticle" approach for the detection and discrimination of antibiotics is developed. The sensing platform was designed based on the different channel's fluorescence recoveries or further quenching of the full-color emissive CDs (F-CDs) and metal ion ensembles upon the addition of antibiotics. The F-CDs exhibited unusually comparable emission intensity nearly across the entire visible spectrum even as the excitation wavelength is shifted, making it very suitable for the construction of multi-channel sensing systems. The sensing platform was fabricated on the basis of the competing interaction of metal ions with the F-CDs and antibiotics. Three metal ions (i.e., Cu2+ , Ce3+ and Eu3+ ) can efficiently quench the fluorescence of the F-CDs. Upon the addition of antibiotics, the fluorescent intensities either recovered at different emission wavelengths or were further quenched to various degrees. The fluorescence response patterns at different emission wavelength were characteristic for each antibiotic and can be quantitatively differentiated by standard statistical methods (e.g., hierarchical clustering analysis and principal component analysis). Moreover, as an example, the proposed method was applied for quantitative detection of oxytetracycline with a limit of detection to be 0.06 µm. Finally, the sensing system was successfully employed for residual antibiotics detection and identification in real food samples.

Facile, Quick, and Gram-Scale Synthesis of Ultralong-Lifetime Room-Temperature-Phosphorescent Carbon Dots by Microwave Irradiation.

Long-lifetime room-temperature phosphorescence (RTP) materials are important for many applications, but they are highly challenging materials owing to the spin-forbidden nature of triplet exciton transitions. Herein, a facile, quick and gram-scale method for the preparation of ultralong RTP (URTP) carbon dots (CDs) was developed via microwave-assisted heating of ethanolamine and phosphoric acid aqueous solution. The CDs exhibit the longest RTP lifetime, 1.46 s (more than 10 s to naked eye) for CDs-based materials to date. The doping of N and P elements is critical for the URTP which is considered to be favored by a n→π* transition facilitating intersystem crossing (ISC) for effectively populating triplet excitons. In addition, possibilities of formation of hydrogen bonds in the interior of the CDs may also play a significant role in producing RTP. Potential applications of the URTP CDs in the fields of anti-counterfeiting and information protection are proposed and demonstrated.

Applying Carbon Dots-Metal Ions Ensembles as a Multichannel Fluorescent Sensor Array: Detection and Discrimination of Phosphate Anions.

Fluorescent carbon dots (CDs) are attracting much attention in sensing recently thanks to their superior optical properties and abundant surface functional groups. To take further advantages of these unique features, CDs are considered to be possible for facilely fabricating multichannel sensor arrays. As a proof-of-concept research, CDs-metal ions ensembles are screened and designed as a triple-channel fluorescent sensor array in this study for the identification of various phosphate anions (e.g., ATP, ADP, AMP, PPi, and Pi) for the first time. Further studies reveal that the selected three metal ions (i.e., Ce3+, Fe3+, and Cu2+) could induce aggregation of the CDs, resulting in quenching of their fluorescence. However, disaggregation or further aggregation of the CDs-metal ions ensembles occurs with the addition of phosphate anions. Consequently, fluorescence of the CDs is recovering or further quenching. On account of various numbers of phosphate group and steric hindrance effects of phosphate anions, their affinities to the sensor array can be distinguished through fluorescence changes of the CDs-metal ions ensembles. By means of statistical analysis methods, the as-developed array is shown excellent capabilities in the detection and discrimination of phosphate anions. Furthermore, practicability of the sensor array is validated by the successful identification of phosphates in serum and blind samples. Compared to previous reports, the as-developed multichannel sensor array manifests numerous advantages, such as simple fabrication process, flexible adjusting detection ranges, and possible extension to other analytes having similar chemical structures or properties.

Triple-Mode Emission of Carbon Dots: Applications for Advanced Anti-Counterfeiting.

Photoluminescence (PL), up-conversion PL (UCPL), and phosphorescence are three kinds of phenomena common to light-emitting materials, but it is very difficult to observe all of them simultaneously when they are derived from a single material at room temperature. For the first time, triple-mode emission (that is, PL, UCPL, and room temperature phosphorescence (RTP)) is reported, which relies on a composite of the luminescent carbon dots (CDs) prepared from m-phenylenediamine and poly(vinyl alcohol) (PVA). Moreover, the CDs-PVA aqueous dispersion is nearly colorless and demonstrates promise as a triple-mode emission ink in the field of advanced anti-counterfeiting.

Colorimetric sensor array for detection and identification of organophosphorus and carbamate pesticides.

Due to relatively low persistence and high effectiveness for insect and pest eradication, organophosphates (OPs) and carbamates are the two major classes of pesticides that broadly used in agriculture. Hence, the sensitive and selective detection of OPs and carbamates is highly significant. In this current study, a colorimetric sensor array comprising five inexpensive and commercially available thiocholine and H2O2 sensitive indicators for the simultaneous detection and identification of OPs and carbamates is developed. The sensing mechanism of this array is based on the irreversible inhibition capability of OPs and carbamates to the activity of acetylcholinesterase (AChE), preventing production of thiocholine and H2O2 from S-acetylthiocholine and acetylcholine and thus resulting in decreased or no color reactions to thiocholine and H2O2 sensitive indicators. Through recognition patterns and standard statistical methods (i.e., hierarchical clustering analysis and principal component analysis), the as-developed array demonstrates not only discrimination of OPs and carbamates from other kinds of pesticides but, more interestingly, identification of them exactly from each other. Moreover, this array is experimentally confirmed to have high selectivity and sensitivity, good anti-interference capability, and potential applications in real samples for OPs and carbamates.

Red, green, and blue luminescence by carbon dots: full-color emission tuning and multicolor cellular imaging.

A facile approach for preparation of photoluminescent (PL) carbon dots (CDs) is reported. The three resulting CDs emit bright and stable red, green and blue (RGB) colors of luminescence, under a single ultraviolet-light excitation. Alterations of PL emission of these CDs are tentatively proposed to result from the difference in their particle size and nitrogen content. Interestingly, up-conversion (UC)PL of these CDs is also observed. Moreover, flexible full-color emissive PVA films can be achieved through mixing two or three CDs in the appropriate ratios. These CDs also show low cytotoxicity and excellent cellular imaging capability. The facile preparation and unique optical features make these CDs potentially useful in numerous applications such as light-emitting diodes, full-color displays, and multiplexed (UC)PL bioimaging.

Paper-based colorimetric array test strip for selective and semiquantitative multi-ion analysis: simultaneous detection of Hg²âº, Ag⁺, and Cu²âº.

A novel approach is presented in this article to qualitatively and semiquantitatively analyze multiple heavy metal ions simultaneously by a colorimetric array test strip. As a proof-of-concept application, a multi-ion analysis array test strip (for Hg(2+), Ag(+), and Cu(2+)) was fabricated through immobilizing five specifically responsive indicators in typical matrixes with tunable sensitivities. The as-obtained test strip shows not only high selectivity for Hg(2+), Ag(+), and Cu(2+), respectively, but also can be applied for their mixtures. The detection limit of the test strip is well below the Chinese wastewater discharge standard concentrations. Moreover, the array test strip demonstrates excellent anti-interference capability, detection, and production reproducibility, long-term storage stability, and real water sample applicability.

The use of a microreactor for rapid screening of the reaction conditions and investigation of the photoluminescence mechanism of carbon dots.

A microreactor is applied and reported, for the first time, in the field of research of carbon dots (CDs), including rapid screening of the reaction conditions and investigation of the photoluminescence (PL) mechanism. Various carbonaceous precursors and solvents were selected and hundreds of reaction conditions were screened (ca. 15 min on average per condition). Through analyzing the screened conditions, tunable PL emission maxima, from about 330 to 550 nm with respectable PL quantum yields, were achieved. Moreover, the relationship between different developmental stages of the CDs and the PL properties was explored by using the microreactor. The PL emission was observed to be independent of the composition, carbonization extent, and morphology/size of the CDs. This study unambiguously presents that a microreactor could serve as a promising tool for the research of CDs.