A Simple, pH-Activatable Fluorescent Aptamer Probe with Ultralow Background for Bispecific Tumor Imaging.

Activatable aptamer probes (AAPs) are promising in molecular imaging of tumors, but the reported shape-switching-dependent AAPs are still challenged by unsatisfied noise suppression, poor stability, and sophisticated sequence design. To address the problem, we constructed a pH-activatable aptamer probe (pH-AAP) by utilizing an acid-labile acetal linker as the responsive element to be fused with a tumor-targeted aptamer. Specifically, a Cy5-labeled aptamer was connected with the quencher BHQ2 through the acetal group, thus generating pH-AAP with quenched fluorescence. Due to the stable proximity of Cy5 to BHQ2, pH-AAP was found to have ultralow background with a quenching efficiency as high as 98%. In comparison with shape-switching-dependent AAPs, the noise suppression of pH-AAP was well maintained for a much longer time in both serum and mouse body, thus showing a robust fluorescence stability. By a combination of the fluorescence recovery induced by acid hydrolysis of acetal linkers and the tumor-targeted recognition of aptamers, pH-AAP could either specifically anchor the extracellular pH-activated signals on the target cell surface in an acidic tumor microenvironment or be activated by acidic lysosomes after it was internalized into target cells. As proof of concept, in vitro evaluation and in vivo imaging of A549 lung cancer cells were performed by using S6 aptamer as a demonstration. It was indicated that pH-AAP realized washing-free, bispecific, and contrast-enhanced tumor imaging. The strategy is simple and free of sequence modification, which promises to provide a universal platform for sensitive and precise tumor diagnosis.

Halobacteriovorax vibrionivorans sp. nov., a novel prokaryotic predator isolated from coastal seawater of China.

Three prokaryotic predator strains, BL9T, BL10 and BL28, were isolated with Vibrio alginolyticus from coastal seawater of PR China. Cells of the strains were Gram-negative, vibrioid-shaped and motile with a single sheathed flagellum (25-28 nm wide). Cells were around 0.3×0.5-1.0 µm in size. The three strains were obligate predators that exhibited a biphasic life cycle: a free-swimming attack phase and an intraperiplasmic growth phase within the prey. Bdelloplasts were formed. NaCl was required for growth. Optimum growth occurred at ~37 °C, with 2-4 % (w/v) NaCl and at pH 7.0-8.0. The results of phylogenetic analyses based on 16S rRNA gene sequences indicated that the three strains shared 99.9 % similarity to each other, were affiliated with the genus Halobacteriovorax in the class Oligoflexia, and represented the same new species. Strain BL9T (=MCCC 1K03527T=JCM 32962T) was designated as the type strain. Genome sequencing of strain BL9T revealed a genome size of 3.14 Mb and a G+C content of 35.8 mol%. The estimated digital DNA-DNA hybridization (dDDH) values and the whole genome average nucleotide identity (gANI) values between the genome of strain BL9T and those of Bdellovibrionales and Bacteriovoracales were 12.5-19 and 63.49-76.15 %, respectively. On the basis of life cycle features, results of physiological analyses, gANI data and dDDH data, strain BL9T represents a new species within the genus Halobacteriovorax, for which the name Halobacteriovoraxvibrionivorans sp. nov. is proposed.

Nature-Inspired Smart DNA Nanodoctor for Activatable In Vivo Cancer Imaging and In Situ Drug Release Based on Recognition-Triggered Assembly of Split Aptamer.

DNA-based activatable theranostic nanoprobes are still unmet for in vivo applications. Here, by utilizing the "induced-fit effect", a smart split aptamer-based activatable theranostic probe (SATP) was first designed as "nanodoctor" for cancer-activated in vivo imaging and in situ drug release. The SATP assembled with quenched fluorescence and stable drug loading in its free state. Once binding to target proteins on cell surface, the SATP disassembled due to recognition-triggered reassembly of split aptamers with activated signals and freed drugs. As proof of concept, split Sgc8c against CEM cancer was used for theranostic studies. Benefiting from the design without blocking aptamer sequence, the SATP maintained an excellent recognition ability similar to intact Sgc8c. An "incubate-and-detect" assay showed that the SATP could significantly lower background and improve signal-to-background ratio (∼4.8 times of "always on" probes), thus affording high sensitivity for CEM cell analysis with 46 cells detected. Also, its high selectivity to target cells was demonstrated in analyzing mixed cell samples and serum samples. Then, using doxorubicin as a model, highly specific drug delivery and cell killing was realized with minimized toxicity to nontarget cells. Moreover, in vivo and ex vivo investigations also revealed that the SATP was specifically activated by CEM tumors inside mice. Especially, contrast-enhanced imaging was achieved in as short as 5 min, thus, laying a foundation for rapid diagnosis and timely therapy. As a biocompatible and target-activatable strategy, the SATP may be widely applied in cancer theranostics.

Iodide-Responsive Cu-Au Nanoparticle-Based Colorimetric Platform for Ultrasensitive Detection of Target Cancer Cells.

Colorimetric analysis is promising in developing facile, fast, and point-of-care cancer diagnosis techniques, but the existing colorimetric cancer cell assays remain problematic because of dissatisfactory sensitivity as well as complex probe design or synthesis. To solve the problem, we here present a novel colorimetric analytical strategy based on iodide-responsive Cu-Au nanoparticles (Cu-Au NPs) combined with the iodide-catalyzed H2O2-TMB (3,3,5,5-tetramethylbenzidine) reaction system. In this strategy, bimetallic Cu-Au NPs prepared with an irregular shape and a diameter of ∼15 nm could chemically absorb iodide, thus indirectly inducing colorimetric signal variation of the H2O2-TMB system. By further utilizing its property of easy biomolecule modification, a versatile colorimetric platform was constructed for detection of any target that could cause the change of Cu-Au NPs concentration via molecular recognition. As proof of concept, an analysis of human leukemia CCRF-CEM cells was performed using aptamer Sgc8c-modified Cu-Au NPs as the colorimetric probe. Results showed that Sgc8c-modified Cu-Au NPs successfully achieved a simple, label-free, cost-effective, visualized, selective, and ultrasensitive detection of cancer cells with a linear range from 50 to 500 cells/mL and a detection limit of 5 cells in 100 µL of binding buffer. Moreover, feasibility was demonstrated for cancer cell analysis in diluted serum samples. The iodide-responsive Cu-Au NP-based colorimetric strategy might not only afford a new design pattern for developing cancer cell assays but also greatly extend the application of the iodide-catalyzed colorimetric system.

Masking agent-free and channel-switch-mode simultaneous sensing of Fe(3+) and Hg(2+) using dual-excitation graphene quantum dots.

A novel channel-switch-mode strategy for simultaneous sensing of Fe(3+) and Hg(2+) is developed with dual-excitation single-emission graphene quantum dots (GQDs). By utilizing the dual-channel fluorescence response performance of GQDs, this strategy achieved a facile, low-cost, masking agent-free, quantitative and selective dual-ion assay even in mixed ion samples and practical water samples.

A versatile activatable fluorescence probing platform for cancer cells in vitro and in vivo based on self-assembled aptamer/carbon nanotube ensembles.

Activatable aptamer probes (AAPs) have emerged as a promising strategy in cancer diagnostics, but existing AAPs remain problematic due to complex design and synthesis, instability in biofluids, or lack of versatility for both in vitro and in vivo applications. Herein, we proposed a novel AAP strategy for cancer cell probing based on fluorophore-labeled aptamer/single-walled carbon nanotube (F-apt/SWNT) ensembles. Through π-stacking interactions and proximity-induced energy transfer, F-apt/SWNT with quenched fluorescence spontaneously formed in its free state and realized signal activation upon targeting surface receptors of living cells. As a demonstration, Sgc8c aptamer was used for in vitro analysis and in vivo imaging of CCRF-CEM cancer cells. It was found that self-assembled Cy5-Sgc8c/SWNT held robust stability for biological applications, including good dispersity in different media and ultralow fluorescence background persistent for 2 h in serum. Flow cytometry assays revealed that Cy5-Sgc8c/SWNT was specifically activated by target cells with dramatic fluorescence elevation and showed improved sensitivity with as low as 12 CCRF-CEM cells detected in mixed samples containing ~100,000 nontarget cells. In vivo studies confirmed that specifically activated fluorescence was imaged in CCRF-CEM tumors, and compared to "always on" probes, Cy5-Sgc8c/SWNT greatly reduced background signals, thus resulting in contrast-enhanced imaging. The general applicability of the strategy was also testified by detecting Ramos cells with aptamer TD05. It was implied that F-apt/SWNT ensembles hold great potential as a simple, stable, sensitive, specific, and versatile activatable platform for both in vitro cancer cell detection and in vivo cancer imaging.

Concatemeric dsDNA-templated copper nanoparticles strategy with improved sensitivity and stability based on rolling circle replication and its application in microRNA detection.

DNA-templated copper nanoparticles (CuNPs) have emerged as promising fluorescent probes for biochemical assays, but the reported monomeric CuNPs remain problematic because of weak fluorescence and poor stability. To solve this problem, a novel concatemeric dsDNA-templated CuNPs (dsDNA-CuNPs) strategy was proposed by introducing the rolling circle replication (RCR) technique into CuNPs synthesis. In this strategy, a short oligonucleotide primer could trigger RCR and be further converted to a long concatemeric dsDNA scaffold through hybridization. After the addition of copper ions and ascorbate, concatemeric dsDNA-CuNPs could effectively form and emit intense fluorescence in the range of 500-650 nm under a 340 nm excitation. In comparison with monomeric dsDNA-CuNPs, the sensitivity of concatemeric dsDNA-CuNPs was greatly improved with ~10,000 folds amplification. And their fluorescence signal was detected to reserve ~60% at 2.5 h after formation, revealing ~2 times enhanced stability. On the basis of these advantages, microRNA let-7d was selected as the model target to testify this strategy as a versatile assay platform. By directly using let-7d as the primer in RCR, the simple, low-cost, and selective microRNA detection was successfully achieved with a good linearity between 10 and 400 pM and a detection limit of 10 pM. The concatemeric dsDNA-CuNPs strategy might be widely adapted to various analytes that can directly or indirectly induce RCR.

A label-free activatable aptamer probe for colorimetric detection of cancer cells based on binding-triggered in situ catalysis of split DNAzyme.

A novel label-free tailed hairpin-shaped activatable aptamer probe (THAAP) was developed by rationally integrating an aptamer and a split G-quadruplex into one sequence. Based on target recognition-triggered in situ catalysis of split DNAzyme, the THAAP strategy achieved a simple, fast, washing-free, specific and quantitative colorimetric assay of human leukemic CCRF-CEM cells.

Activatable aptamer probe for contrast-enhanced in vivo cancer imaging based on cell membrane protein-triggered conformation alteration.

Aptamers have emerged as promising molecular probes for in vivo cancer imaging, but the reported "always-on" aptamer probes remain problematic because of high background and limited contrast. To address this problem, we designed an activatable aptamer probe (AAP) targeting membrane proteins of living cancer cells and achieved contrast-enhanced cancer visualization inside mice. The AAP displayed a quenched fluorescence in its free state and underwent a conformational alteration upon binding to target cancer cells with an activated fluorescence. As proof of concept, in vitro analysis and in vivo imaging of CCRF-CEM cancer cells were performed by using the specific aptamer, sgc8, as a demonstration. It was confirmed that the AAP could be specifically activated by target cancer cells with a dramatic fluorescence enhancement and exhibit improved sensitivity for CCRF-CEM cell analysis with the cell number of 118 detected in 200 µl binding buffer. In vivo studies demonstrated that activated fluorescence signals were obviously achieved in the CCRF-CEM tumor sites in mice. Compared to always-on aptamer probes, the AAP could substantially minimize the background signal originating from nontarget tissues, thus resulting in significantly enhanced image contrast and shortened diagnosis time to 15 min. Furthermore, because of the specific affinity of sgc8 to target cancer cells, the AAP also showed desirable specificity in differentiating CCRF-CEM tumors from Ramos tumors and nontumor areas. The design concept can be widely adapted to other cancer cell-specific aptamer probes for in vivo molecular imaging of cancer.

Poly(thymine)-templated fluorescent copper nanoparticles for ultrasensitive label-free nuclease assay and its inhibitors screening.

Noble-metal fluorescent nanoparticles have attracted considerable interest on account of their excellent properties and potential applicable importance in many fields. Particularly, we recently found that poly(thymine) (poly T) could template the formation of fluorescent copper nanoparticles (CuNPs), offering admirable potential as novel functional biochemical probes. However, exploration of poly T-templated CuNPs for application is still at a very early stage. We report herein for the first example to develop a novel ultrasensitive label-free method for the nuclease (S1 nuclease as a model system) assay, and its inhibitors screening using the poly T-templated fluorescent CuNPs. In this assay, the signal reporter of poly T of 30 mer (T30) kept the original long state in the absence of nuclease, which could effectively template the formation of fluorescent CuNPs. In the presence of nuclease, poly T was digested to mono- or oligonucleotide fragments with decrease of fluorescence. The proposed method was low-cost and simple in its operation without requirement for complex labeling of probe DNA or sophisticated synthesis of the fluorescent compound. The assay process was very rapid with only 5 min for the formation of fluorescent CuNPs. The capabilities for target detection from complex fluids and screening of nuclease inhibitors were verified. A high sensitivity exhibited with a detectable minimum concentration of 5 × 10(-7) units µL(-1) S1 nuclease, which was about 1-4 orders of magnitude more sensitive than the developed approaches.

A facile graphene oxide-based DNA polymerase assay.

The DNA polymerase assay is fundamental for related molecular biology investigations and drug screenings, however, the commonly used radioactive method is laborious and restricted. Herein, we report a novel, simple and cost-effective fluorometric DNA polymerase detection method by utilizing graphene oxide (GO) as a signal switch. In this strategy, in the absence of DNA polymerase, the fluorophore-labeled template ssDNA could be strongly adsorbed and almost entirely quenched by GO. However, as DNA polymerase exists, the polymerized dsDNA product might lead to a much lower quenching efficiency after addition of GO due to the much weaker interaction of dsDNA with GO than ssDNA, thus resulting in a much higher fluorescence signal detected. As proof of concept, the quantitative DNA polymerase activity assay was performed using the Klenow fragment exo(-) (KF(-)) as a model. It was confirmed that, after optimization of detection conditions, KF(-) activity could be sensitively detected through facile fluorescence measurements, with a detection limit of 0.05 U mL(-1) and a good linear correlation between 0.05-2.5 U mL(-1) (R(2) = 0.9928). In addition, this GO-based method was further inspected to evaluate the inhibitive behaviors of several drugs toward KF(-) activity, the result of which firmly demonstrated its potential application in polymerization-targeted drug screening.

A persistent luminescent nanobeacon for practical detection of lead ions via avoiding background interference.

Heavy metal ions are considered to be the most serious sources for water pollution. Accurate detection of metal ions is important for pollution control and ecological protection. Background interference is an inevitable obstacle in the fluorescent analysis of complex samples. Herein, a persistent luminescent nanobeacon is communicated to detect metal ions in real samples via avoiding background interference. The nanobeacon is constituted by persistent luminescence nanomaterials and metal-specific DNAzymes. As a proof of concept, Zn2GeO4: Mn persistent luminescence nanorods (PLNRs) was synthesized and functionalized with the 17E DNAzyme for lead ion (Pb2+) detection. As a result, in the luminescent manner, the nanobeacon could recognize Pb2+ selectively and detect it with high signal-to-background ratios (SBR) both in buffer and real samples, but the fluorescent SBR declined significantly when used in real samples. Thus, this persistent luminescent nanobeacon can achieve practical detection of metal ions via avoiding background interference. Compared to previous methods of improving signal-to-background ratio, this persistent luminescent nanobeacon is more accessible, and all DNAzyme-specific ions can be directly adapted.

Copper ion and G-quadruplex-mediated fluorescent sensor for highly selective detection of bleomycin in actual samples.

Improper dosage of Bleomycin (BLM) can easily lead to a series of side effects such as pulmonary fibrosis and pulmonary toxicity. Therefore, detecting the content of BLM in actual sample is very helpful to make full use of its therapeutic efficacy and reduce its toxicity. Herein, we constructed a copper ion and G-quadruplex mediated label-free sensor to detect BLM. The strategy mainly relies on the chelation of BLM to copper ions, which makes the copper ions lose the quenching ability to the fluorescent dye N-methylmesoporphyrin (NMM) after chelation. With the assistance of the G-quadruplex, the BLM content in the sample can be detected by observing the change in fluorescence. A good linear relationship can be clearly observed within the BLM concentration range of 0.1 nM-75 nM, and the limit of detection was derived as 0.1 nM. This sensor did not involve any labeling or addition of Fe2+, even in the presence of 10 different antibiotics and 11 different metal ions, it still has a good monitoring effect, and can be successfully applied to the detection of BLM in serum and wastewater. Thus, we believe that this work hold great potential in antibiotic monitoring and environmental protection.

A novel CRISPR/Cas14a system integrated with 2D porphyrin metal-organic framework for microcystin-LR determination through a homogeneous competitive reaction.

Selective and sensitive detection of microcystin-LR (MC-LR) is of vital importance because of its high toxicity and broad distribution. Herein, a novel and versatile fluorescence sensor (Cas14-pMOFs fluorescence sensor) was developed by combining the CRISPR/Cas14a system with a 2D porphyrin metal-organic framework nanosheets (2D-pMOFs) for MC-LR determination. The designed CRISPR/Cas14a system was activated by the unbound complementary DNA (cDNA), which was positively correlated with MC-LR concentration. Furthermore, the activated Cas14a protein was utilized to indiscriminately cleave the FAM-labeled single-stranded DNA (ssDNA-FAM), which was pre-absorbed on Cu-TCPP(Fe) nanosheets. Because of the desorption of the cleaved ssDNA-FAM, the pre-quenched fluorescence signal was recovered. Owing to the excellent performance in quantifying cDNA using this Cas14-pMOFs fluorescence sensor with a limit of detection (LOD) of 0.12 nM, this Cas14-pMOFs fluorescence sensor was able to detect MC-LR in a range from 50 pg/mL to 1 µg/mL with the LOD of 19 pg/mL. This work not only provided a new insight for the exploration of fluorescence sensors based on 2D-pMOFs coupled with CRISPR/Cas14a, but also, demonstrated its universality in both nucleic acid and non-nucleic acid targets determination.

Recent advances in performance improvement of Metal-organic Frameworks to remove antibiotics: Mechanism and evaluation.

Antibiotic pollution is a serious global problem, which may threaten the health of human and ecosystem. Thereinto, water pollution is the most common way. Thus, it is necessary to develop effective methods to remove antibiotics from the natural aqueous environments. Metal-organic Frameworks (MOFs) - based adsorption and photocatalysis strategies have been demonstrated to be efficient, environmental and promising methods to solve antibiotic pollution and repair the environment. In this review, several strategies to improve the properties of MOFs for removal were summarized and discussed. And the removal mechanisms were also discussed. Besides, new and more reliable evaluation methods of MOFs to remove antibiotics were presented, including preferential adsorption (qp), quantum yields (QY), space time yields (SY) and figure of merit (FOM). This paper provides alternative perspectives for researchers to improve the properties of MOFs and raise analytic efficiency of antibiotic removal.

Constructing an In Situ Polymer Electrolyte and a Na-Rich Artificial SEI Layer toward Practical Solid-State Na Metal Batteries.

Sodium is one of the most promising anode candidates for the beyond-lithium-ion batteries. The development of Na metal batteries with a high energy density, high safety, and low cost is desirable to meet the requirements of both portable and stationary electrical energy storage. However, several problems caused by the unstable Na metal anode and the unsafe liquid electrolyte severely hinder their practical applications. Herein, we report a facile but effective methodology to construct an in situ polymer electrolyte and Na-rich artificial solid-electrolyte interface (SEI) layer concurrently. The obtained integrated Na metal batteries display long cycling life and admirable dynamic performance with total inhibition of dendrites, excellent contact of the cathode/polymer electrolyte, and reduction of side reactions during cycling. The modified Na metal electrode with the in situ polymer electrolyte is stable and dendrite-free in repeated plating/stripping processes with a life span of above 1000 h. Moreover, this method is compatible with different cathodes that demonstrate outstanding electrochemical performance in full cells. We believe that this approach provides a practical solution to solid-state Na metal batteries.

Cu/Au/Pt trimetallic nanoparticles coated with DNA hydrogel as target-responsive and signal-amplification material for sensitive detection of microcystin-LR.

Sensitive and reliable analytical methods for monitoring of microcystin-LR (MC-LR) are urgently necessary due to its great harm to human health and aquatic organisms. In this work, a novel Cu/Au/Pt trimetallic nanoparticles (Cu/Au/Pt TNs)-encapsulated DNA hydrogel was prepared for colorimetric detection of MC-LR. The Cu/Au/Pt TNs were captured and released with precise control by the target-responsive 3D DNA hydrogels, which combined dual advantages of the target responsive DNA hydrogel and Cu/Au/Pt TNs of enhanced peroxidase-like activity. The DNA hydrogel network was constructed by hybridizing MC-LR aptamer with two complementary DNA strands on linear polyacrylamide chains. As long as MC-LR presented, the aptamer competitively binds with the MC-LR, causing the hydrogel to dissolve and release the preloaded Cu/Au/Pt TNs which could catalyze the reaction between H2O2 and TMB to produce color changes. In view of this sensitive strategy, this Cu/Au/Pt TNs-encapsulated DNA hydrogel-based colorimetric biosensor can achieve quantitative determination of MC-LR. The results showed that as-proposed colorimetric biosensor could sensitively detect MC-LR with a linear range of 4.0-10000 ng L-1 and a detection limit of 3.0 ng L-1. This work proved that the sensor had great potential to be applied in MC-LR detection and also provided the opportunity to develop colorimetric biosensor for other targets using this target-responsive and signal-amplification strategy.

One-pot synthesized Cu/Au/Pt trimetallic nanoparticles as a novel enzyme mimic for biosensing applications.

Multimetallic nanomaterials have aroused special attention owing to the unique characteristics of chemical, optical and enhanced enzyme mimetic capabilities resulting from the synergistic effect of different metal elements. In this work, we present a facile, gentle, fast and one-pot method for preparing Cu/Au/Pt trimetallic nanoparticles (TNPs), which possess intrinsic and enhanced peroxidase-like activity as well as excellent stability, sustainable catalytic activity, and robustness to harsh environments. Kinetic analysis indicated that Cu/Au/Pt TNPs exhibited strong affinities with H2O2 and 3,3,5,5-tetramethylbenzidine (TMB) as the substrates. To investigate the feasibility of Cu/Au/Pt TNPs-based strategy in biological analysis, H2O2 was chosen as a model analyte and a sensitive and specific detection for H2O2 was acquired with a detection limit of 17 nM. By coupling with glucose oxidase (GOD), this assay could also achieve a sensitive and selective detection of glucose with a detection limit of 33 µM, indicating the versatility of the method. In view of the potential combination with diverse enzyme-related reactions, the Cu/Au/Pt TNPs-based strategy is promising as a universal platform for biosensors.

One-pot synthesized Cu/Au/Pt trimetallic nanoparticles with enhanced catalytic and plasmonic properties as a universal platform for biosensing and cancer theranostics.

Cu/Au/Pt trimetallic nanoparticles (TMNPs) with enhanced catalytic activity and intense plasmonic absorption in the NIR-I biowindow (650-950 nm) were prepared using a fast, gentle and one-pot protocol. Based on these properties and assembly of thiolated-aptamers on Cu/Au/Pt TMNPs, a universal platform was developed for applications in biosensing and theranostics.

Draft Genome Sequence of an Extracellular Protease-Producing Bacterium, Stenotrophomonas bentonitica VV6, Isolated from Arctic Seawater.

The draft genome sequence of the extracellular protease-producing bacterium Stenotrophomonas bentonitica VV6, isolated from Arctic seawater, was established. The genome size was approximately 4.365 Mb, with a G+C content of 66.54%, and it contains 3,871 predicted protein-coding sequences (CDSs) and 60 tRNAs.