To date, various fluorescent probes for G-quadruplex have been reported. However, the development of fluorescent dyes for split G-quadruplex formed on double-stranded DNA structure still remained largely unexplored. Herein, we have demonstrated that thioflavin T (ThT) was able to intercalate into split G-quadruplex sequence and the formed split G-quadruplex-ThT structure exhibited remarkable fluorescence enhancement. By integrating cobalt oxyhydroxide (CoOOH) nanoflakes and hybridization chain reaction (HCR), a sensitive and label-free biosensing strategy for miRNA detection was proposed. In our design, miRNA-21 was chosen as the model target and it was capable of triggering HCR with two split G-quadruplex sequence-contained hairpin probes, resulting in the generation of long chain-like HCR product. Unlike hairpin probes, the HCR product cannot be adsorbed onto CoOOH nanoflakes because of lower affinity. Consequently, in the presence of ThT, numerous split G-quadruplex-ThT structures on HCR product were formed and the target miRNA was converted into amplified and activatable fluorescence signal. Experimental results demonstrated that this novel sensing strategy can be applied for sensitive and selective detection of miRNA. Moreover, the analysis of miRNA in complex biological media was successfully realized, offering a robust, label-free and cost-effective platform for miRNA-related disease diagnosis and biomedical research.

Abstract The utilization of light as an external stimulus to promote the generation of hot electrons in plasmonic heterostructures and thus augment their catalytic efficacy presents significant potential for bacterial biofilm eradication. However, the inefficient harnessing of photoinduced hot electrons in conventional developed strategies greatly impede their therapeutic application in bone tissues. To overcome these challenges, we herein engineered a near-infrared II (NIR-II)-triggered plasmonic catalysis, which was fabricated through the integration of gold nanobipyramids (Au NBPs) with tip-deposited platinum nanoparticles (Pt NPs), for effective elimination of hypoxic bacterial biofilms on bone implants. The strategic deposition of Pt NPs at the tip of Au NBPs (ePt-Au NBPs) not only brought the redshift of the NIR absorption peak, but also accelerated charge separation and electromagnetic field localization, which endowed the ePt-Au NBPs plasmonic heterostructures with enhanced catalytic activity. Under NIR-II laser irradiation, the plasmonic catalysis with tip-localized enhancement enabled robust generation of hydroxyl radicals (•OH), thereby facilitating the cleavage of extracellular DNA (eDNA) within biofilms, disrupting biofilm integrity, and ultimately sensitizing bacteria to thermal ablation. These attributes collectively contribute to the effective elimination of hypoxic bacterial biofilms. Furthermore, surface functionalization with RGDC peptides conferred the implant with superior biocompatibility and osteogenic integration capabilities. This rationally designed plasmonic catalysis, combining the NIR-II-triggered simultaneous production of enhanced catalytic activity and localized hyperthermia, demonstrates significant potential for translational applications in light-responsive therapeutic strategies for implant-associated infections.

Growth factor holds great promise for bone regeneration, and spatiotemporal control of their expressing through site-specific reactions is crucial but challenging for on-demand therapy. In this study, we present the development of a novel unnatural amino acids (UAAs)-triggered therapeutic switch (UATS) system, composed of an orthogonal aminoacyl-tRNA-synthase (aaRS)-tRNA pair and a bone morphogenetic protein 2 (BMP2) gene harboring premature stop codon, which enable in situ and on-demand initiation of the expression of BMP2. The resulting UATS system allowed specifically control of base expressing on the BMP2 mRNA that switched to the BMP2 protein with complete structure and function to facilitate bone regeneration. Our investigations showed that the UATS system exhibits remarkable attributes of rapid, sensitive, reversible, and sustained BMP2 expression both in vitro and in vivo settings. Moreover, the implantation of microencapsulated cells with UATS system is applied to a mouse femur defect model, demonstrating high effciency in controlled expressing of BMP2 protein and substantial repair of bone defect following oral administration of UAAs. Therefore, our findings underscore the great potential of UATS system for on-demand awakening of functional growth factor, thus offering promising prospects in the realm of regenerative medicine.

DNA walkers have attracted considerable attention in biosensing and bioimaging. Compared with the conventional single leg-based DNA walker, the bipedal DNA walker has remarkable advantages, with improved sensitivity and fast kinetics, and can work efficiently in a crowded cellular environment. However, most reported bipedal DNA walkers are powered by exogenous supplementation, and elaborate DNA sequence designs, auxiliary additives or extra carriers are often needed. A highly integrated bipedal DNA walker that can address robustness, sensitivity and consistency issues in a single system is highly desirable but remains a great challenge. We herein report a novel bipedal DNA nanowalker system through simple assembly of a DNA substrate, hairpin functionalized-AuNPs (AuNPs-H2), and a blocked Mn2+-dependent DNAzyme hairpin (H1) on degradable MnO2 nanosheets, which holds great potential for living cell operation. Highly integrated features enable the simultaneous delivery of core components of the bipedal DNA walker, including a walking track (AuNPs-H2), a walking strand (H1 cleaved by APE1), and a driving force (Mn2+-dependent DNAzyme cleavage) as a whole, thereby enhancing the control of the spatiotemporal distribution of these components at the intracellular target sites. The redox reaction between the MnO2 nanosheets and GSH inside the cells not only consumed the intracellular GSH to improve the biostability of the walking track but also generated abundant Mn2+ as a cofactor of the DNAzyme. As a proof of concept, the developed nanowalker was demonstrated to work efficiently for monitoring base excision repair (BER)-related human apurinic/apyrimidinic endonuclease 1 (APE1) in living cells, highlighting the great potential of the bipedal DNA nanowalker in biological systems.

Nanomaterials possess unusual physicochemical properties including unique optical, magnetic, electronic properties, and large surface-to-volume ratio. However, nanomaterials face some challenges when they were applied in the field of biomedicine. For example, some nanomaterials suffer from the limitations such as poor selectivity and biocompatibility, low stability, and solubility. To address the above-mentioned obstacles, functional nucleic acid has been widely served as a powerful and versatile ligand for modifying nanomaterials because of their unique characteristics, such as ease of modification, excellent biocompatibility, high stability, predictable intermolecular interaction and recognition ability. The functionally integrating functional nucleic acid with nanomaterials has produced various kinds of nanocomposites and recent advances in applications of functional nucleic acid decorated nanomaterials for cancer imaging and therapy were summarized in this review. Further, we offer an insight into the future challenges and perspectives of functional nucleic acid decorated nanomaterials.

Accurate, rapid, and quantitative detection of D-penicillamine (D-PA) in pharmaceutical formulations is imperative for drug quality and clinical medication guidance. Various colorimetric and fluorescent assays have been constructed for D-PA based on nanomaterials. However, the reliable and rapid detection of D-PA in pharmaceutical formulations remains a challenge because these methods suffer from shortcomings such as time-consuming procedures, poor uniformity, and complicated nanomaterial preparation. In this work, an effective and facile homogeneous dual-mode optical sensing strategy was established based on the strong complexation between D-PA and Cu2+ and catalyzed the oxidation of O-phenylenediamine. This platform displayed a sensitive response to D-PA from 5 to 100 mu mol/L, with a detection limits of 0.54 mu mol/L (fluorescence) and 0.76 mu mol/L (colorimetry). As expected, the method was successfully applied to the determination of D-PA in pharmaceutical formulations with satisfactory results. Overall, this developed strategy provides a facile and rapid dual-mode optical platform for D-PA and has great potential in D-PA-related pharmaceutical quality control.

In this work, a novel fluorescence nanoprobe based on MnO2 nanosheets and DNAzyme cyclic amplification was developed for label-free detection of GSH. In our design, the DNA@MnO2 nanoprobe was facilely prepared through the adsorption of DNA probes on the surface of MnO2 nanosheets. Importantly, MnO2 nanosheets in the DNA@MnO2 nanoprobe can act as the recognizer of analytical target GSH. In the presence of GSH, the DNA@MnO2 nanoprobe are decomposed owing to redox-responsive reaction between GSH and MnO2 nanosheets, resulting in the generation of abundant Mn2+ and the desorption of DNA probes. Subsequently, the Mn2+dependent DNAzyme recycling amplification was rapidly initiated and a large number of the blocked G-rich sequences in DNA probes was released. The G-rich sequences were efficiently folded into G-quadruplex structures, which can remarkably light up the fluorescence intensity of thioflavin T (ThT). Experimental data demonstrated that the DNA@MnO2 nanoprobes for GSH detection have a wide linear range from 0 to 2000 mu M. Moreover, this fluorescence sensing strategy was successfully applied for the detection of GSH in complex biological media with satisfying results, providing a simple, facile and cost-efficient nanoplatform for GSH-related clinical disease diagnosis.

A novel fluorescent strategy has been developed by using an enzymatic reaction modulated DNA assembly on graphitic carbon nitride nanosheets (CNNS) for the detection of acetylcholinesterase (AChE) activity and its inhibitors. The two-dimensional and ultrathin-layer CNNS-material was successfully synthesized through a chemical oxidation and ultrasound exfoliation method. Because of its excellent adsorption selectivity to ssDNA over dsDNA and superior quenching ability toward the fluorophore labels, CNNS were employed to construct a sensitive fluorescence sensing platform for the detection of AChE activity and inhibition. The detection was based on enzymatic reaction modulated DNA assembly on CNNS, which involved the specific AChE-catalyzed reaction-mediated DNA/Hg2+ conformational change and subsequent signal transduction and amplification via hybridization chain reaction (HCR). Under the excitation at 485 nm, the fluorescence signal from 500 to 650 nm (λmax = 518 nm) of the developed sensing system was gradually increased with increasing concentration of AChE. The quantitative determination range of AChE is from 0.02 to 1 mU/mL and the detection limit was 0.006 mU/mL. The developed strategy was successfully applied to the assay of AChE in human serum samples, and can also be used to effectively screen AChE inhibitors, showing great promise providing a robust and effective platform for AChE-related diagnosis, drug screening, and therapy.

DNA molecular probes have been increasingly used for biological imaging and tumor theranostics. In particular, live-cell imaging of low abundance microRNA (miRNA) with high accuracy and efficiency was extremely important for understanding functions and dynamics of miRNA. Herein, an easy-to-obtain DNA-nucleated spherical nucleic acid (DSNA) was constructed by one pair of hairpin probes "sticked" on self-assembled threedimensional (3D) urchin-like DNA nanostructures (UDNs) to realize sensitive imaging of tumor-related miRNA in living cells. UDNs were prepared through four short single-stranded DNA by a simple "one-pot" annealing process that took only a few minutes. Moreover, sticking the reactants of hairpin probes on the UDNs not only sped up the reaction kinetic rate, but also improved the sensitivity and efficiency of miRNA-triggered hairpin cascade circuits. Compared with traditional AuNP-based spherical nucleic acid, the DSNA possessed superb biocompatibility and stability, excellent cell internalization capability and accessibility, which endowed them tremendous advantages for the monitoring of miRNA in living cells. Therefore, our developed strategy exhibited great potential in sensitive imaging of intracellular miRNA, offering a new avenue for early tumor diagnosis and related biomedical research.

Two-photon carbon-based nanoprobes hold great potential for biomedical applications as a result of their advantages of low fluorescence background, deep tissue imaging penetration and enhanced spatial resolution. However, the development of an activatable two-photon fluorescence carbon-based nanoprobe that simultaneously has the ability to target desired organs or cells is highly desired but remained a largely unsolved challenge. Herein, we developed boronate affinity BCNP@MnO2 nanocomposites, constructed by one step in situ growth of MnO2 nanosheets on the surface of aminophenylboronic acid-functionalized CNPs (BCNPs) via a redox reaction, which can feature efficient fluorescence energy transfer quenching to the BCNPs, allowing for tumor-specific affinity recognition and two-photon fluorescence activation imaging. By utilizing the inherent two-photon optical properties and sialic acid (SA) specific targeting ability of the BCNPs, good biocompatibility of the nanocomposites as well as highly sensitive and selective responses of MnO2 nanosheets towards GSH, the developed nanocomposites have demonstrated specific two-photon fluorescence activation imaging in target cancer cells and nude mouse tissues. Therefore, our proposed novel strategy could be used for monitoring GSH-triggered two-photon fluorescence activation events in SA-overexpressed cancer cells and has promising applications in both biological exploration and clinical diagnosis.

光声成像是一种新兴的非侵入式的生物成像方式,具有极高的空间分辨率和良好的成像对比度,已逐步应用于肿瘤成像诊断基础研究.光声成像主要依赖于光声信号转换,而光声信号转换能力主要取决于造影剂的选择.近年来,随着无机纳米材料在生物医学成像领域的研究逐渐深入,越来越多的二维无机纳米材料也应用于光声成像造影剂,尤其是新型类石墨烯二维纳米材料,其优异的近红外吸收率类似于石墨烯,光热转换效率高,生物相容性良好,而且部分材料还具备带隙可调特性以及良好的生物降解性,因此有望成为肿瘤光声成像理想的造影剂.其中,二维过渡金属硫化物、二维过渡金属碳/氮化物以及二维单元素材料已被多次报道应用于肿瘤光声成像造影剂的研发.该文将综述上述几类二维纳米材料在肿瘤光声成像诊断中的应用进展.

DNA molecular probes have emerged as powerful tools for fluorescence imaging of microRNAs (miRNAs) in living cells and thus elucidating functions and dynamics of miRNAs. In particular, the highly integrated DNA probes that can be able to address the robustness, sensitivity and consistency issues in a single assay system were highly desired but remained largely unsolved challenge. Herein, we reported for the first time that the development of the novel DNA nanomachines that split-DNAzyme motif was highly integrated in a single DNA triangular prism (DTP) reactor and can undergo target-activated DNAzyme catalytic cascade circuits, allowing amplified sensing and imaging of tumor-related microRNA-21 (miR-21) in living cells. The DNA nanomachines have shown dynamic responses for target miR-21 with excellent sensitivity and selectivity and demonstrated the potential for living cell imaging of miR-21. With the advantages of facile modular design and assembly, high biostability, low cytotoxicity and excellent cellular internalization, the highly integrated DNA nanomachines enabled accurate and effective monitoring of miR-21 expression levels in living cells. Therefore, our developed strategy may afford a reliable and robust nanoplatform for tumor diagnosis and for related biological research.

DNA molecular probes have emerged as a powerful tool for RNA imaging. Hurdles in cell-specific delivery and other issues such as insufficient stability, limited sensitivity, or slow reaction kinetics, however, hinder the further application of DNA molecular probes in vivo. Herein, we report an aptamer-tethered DNA polymer for cell-specific transportation and amplified imaging of RNA in vivo via a DNA cascade reaction. DNA polymers are constructed through an initiator-triggered hybridization chain reaction using two functional DNA monomers. The prepared DNA polymers show low cytotoxicity and good stability against nuclease degradation and enable cell-specific transportation of DNA circuits via aptamer-receptor binding. Moreover, assembling the reactants of hairpins C1 and C2 on the DNA polymers accelerates the response kinetics and improves the sensitivity of the cascade reaction. We also show that the DNA polymers enable efficient imaging of microRNA-21 in live cells and in vivo via intravenous injection. The DNA polymers provide a valuable platform for targeted and amplified RNA imaging in vivo, which holds great implications for early clinical diagnosis and therapy.

DNA nanowalkers moving progressively along a prescribed DNA track are useful tools in biosensing, molecular theranostics and biosynthesis. However, stochastic DNA nanowalkers that can perform in living cells have been largely unexplored. We report the development of a novel stochastic bipedal DNA walker that, for the first time, realizes direct intracellular base excision repair (BER) fluorescence activation imaging. In our design, the bipedal walker DNA was generated by BER-related human apurinic/apyrimidinic endonuclease 1 (APE1)-mediated cleavage of DNA sequences at an abasic site in the intracellular environment, and it autonomously travelled on spherical nucleic acid (SNA) surfaces via catalyzed hairpin assembly (CHA). Our nanomachine outperforms the conventional single leg-based DNA walker with an improved sensitivity, kinetics and walking steps. Moreover, in contrast to the single leg-based DNA walker, the bipedal DNA walker is capable of monitoring the fluorescence signal of reduced APE1 activity, thus indicating amplified intracellular imaging. This bipedal DNA-propelled DNA walker presents a simple and modular amplification mechanism for intracellular biomarkers of interest, providing an invaluable platform for low-abundance biomarker discovery leading to the accurate identification and effective treatment of cancers.

Detecting the interactions between small molecules and proteins was critical for disease theranostics and drug development. Here we propose a novel universal assay strategy for monitoring small molecule-protein interactions in solution using strand displacement amplification (SDA) mediated by protein binding to small molecule with DNAzyme-based chemiluminescence detection. The DNA polymerase and nicking enzyme assisted SDA could yield a great amount of peroxidase-mimicking DNAzyme sequences which cause significantly chemiluminescence signals, while protein binding to the small molecule label would prevent DNA polymerase from extending nick site and DNAzyme sequence, and thus the chemiluminescence signals would obviously decrease. This strategy was demonstrated using folate and its binding protein (folate receptor), and the results revealed that the developed strategy enable offer a label-free, homogeneous, and highly sensitive chemiluminescence detection of folate receptor with a detection limit of 1pM. At the same time, it has been successfully used for folate receptor detection in human serum. The proposed chemiluminescence sensing method might provide a generic, robust, and high-throughput platform for detecting various small molecule-protein interactions for biological applications.

A multifunctional theranostic nanoplatform, which integrates diagnostic and therapeutic functions in a single nanosystem, holds great promise for guiding disease treatment and improving the corresponding therapy efficacy. We report the development of a novel g-C(3)N(4)nanosheet-based theranostic nanoassembly for both enhanced imaging of cancer-relevant mRNA in living cells and imaging-guided on-demand photodynamic therapy (PDT) for tumors. The nanoassembly was constructed by using highly fluorescent and water-dispersible g-C(3)N(4)nanosheets which act as nanocarriers, enabling efficient and self-tracking transfection of the DNA hairpin probes. The presence of intracellular mRNA will initiate the DNA hairpin probes, ultimately resulting in an amplified fluorescence signalviahybridization and displacement with mRNA. Moreover, enhanced fluorescence imaging-guided precise PDT for tumors in living cells was also demonstrated, allowing the selective ablation of tumors without any obvious side effects. Therefore, the developed theranostic approach can provide a promising platform for low-abundance biomarker discovery and early treatment of related diseases.

DNA hydrogels are biocompatible and are suitable for many biomedical applications. However, to be useful imaging probes or drug carriers, the ordinary bulk size of DNA hydrogels must be overcome. Here we put forward a new strategy for fabricating a novel and simple protein-scaffolded DNA nanohydrogel, constructed through a direct DNA self-assembly using three types of streptavidin (SA)-based DNA tetrad for the activation of imaging and targeting therapy of cancer cells. The DNA nanohydrogels are easily prepared, and we show that by varying the initial concentration of DNA tetrad, it is possible to finely control their size within nanoscale range, which are favorable as carriers for intracellular imaging and transport. By further incorporating therapeutic agents and tumor-targeting MUC1 aptamer, these multifunctionalized SA-scaffolded DNA nanohydrogels (SDH) can specifically target cancer cells and selectively release the preloaded therapeutic agents via a structure switching when in an ATP-rich intracellular environment, leading to the activation of the fluorescence and efficient treatment of cancer cells. With the advantages of facile modular design and assembly, effective cellular uptake, and excellent biocompatibility, the method reported here has the potential for the development of new tunable DNA nanohydrogels with multiple synergistic functionalities for biological and biomedical applications.

It is of great value to develop a simple, rapid, label-free and sensitive strategy for tyrosinase activity detection and corresponding inhibitor screening in both biomedical diagnosis and cosmetic industry. However, many recently reported fluorescent assays may suffer from instability to light irradiation, poor water solubility, and complex modification. Herein, a novel fluorescent biosensor strategy has been developed capable of sensitive, label-free and rapid screening of tyrosinase activity as well as corresponding inhibitors based on fluorescent block copolymer nanoparticles (BCNs). The reported BCNs were prepared by coprecipitation assay using the mixture of amphiphilic block copolymer and fluorescent conjugated polymer, which show many advantages including high brightness, excellent photostability and biocompatibility, enabling the biosensor with high sensitivity and good reproducibility in a single-step operation. The reported assay exhibited high sensitivity and selectivity with a detection limit of 1.5 ng/mL for tyrosinase (TRY) activity detection and hold potential in its inhibitor screening. The satisfying recoveries offered great potential for complicated sample analysis. Hence, this reported assay may provide a novel platform for biosensor development.

A novel and versatile platform for single-step amplified fluorescence detection of antibodies via specific proximity-induced hybridization chain assembly is developed.