Carbon dots combined with phytosphingosine inhibit acid-induced demineralization of hydroxyapatite in vitro.

OBJECTIVES: To study the effects of carbon dots (CDs), in combination with phytosphingosine (PHS), against acid-induced demineralization of hydroxyapatite in vitro. METHODS: CDs were generated from citric acid and urea by microwave heating. Transmission electron microscope (TEM), FT-IR, and fluorescence intensity were used to characterize the CDs. A hydroxyapatite (HAp) model was used to investigate the protective effects of CDs, PHS, and their combinations with and without a salivary pellicle against acid-induced demineralization in vitro. Ca2+ release as a parameter to evaluate the inhibition of demineralization was measured by capillary electrophoresis. The interactions between CDs, PHS, and HAp discs were investigated using a fluorescence detector. RESULTS: Uniform-sized CDs were synthesized, showing typical optical characteristics. CDs exhibited no inhibition of acid-induced demineralization in vitro, in contrast to PHS. Notably, a pre-coating of CDs increased the protective effects of PHS against acid-induced demineralization, which was not disturbed by the presence of a salivary pellicle and Tween 20. Scanning electron microscope (SEM) confirmed the binding and layers formed of both CDs and PHS to the HAp surfaces. Based on fluorescence spectra CDs binding to HAp seemed to be dependent on Ca2+ and PO43- interactions. CONCLUSIONS: CDs combined with PHS showed protective effects against acid-induced demineralization of HAp discs in vitro.

A ratiometric SERS sensor with one signal probe for ultrasensitive and quantitative monitoring of serum xanthine.

Xanthine can be converted into uric acid, and a high concentration of xanthine in the human body can cause many diseases. Therefore, it is important to develop a sensitive, simple, and reliable approach for measuring xanthine in biological liquids. Hence, a ratiometric surface-enhanced Raman spectroscopy (SERS) sensing strategy with one signal probe was exploited for reliable, sensitive, and quantitative monitoring of serum xanthine. 3-Mercaptophenylboronic acid (3-MPBA) was used as a typical reference with a Raman peak at 996 cm-1. First, 3-MPBA was bound to gold nanoflowers@silica (GNFs@Si) through Au-S bonds. Xanthine oxidase (XOD) catalyzed the oxidation of xanthine into H2O2 on GNFs@Si. Afterward, the obtained H2O2 further reduced 3-MPBA to 3-hydroxythiophenol (3-HTP) accompanied by the emergence of a new Raman peak at 883 cm-1. Meanwhile, the Raman intensity at 996 cm-1 remained constant. Therefore, the ratio of I883/I996 increased with the increasing of xanthine concentration, thus realizing quantitative detection of xanthine. As a result, a ratiometric SERS sensor for the detection of xanthine was proposed with a detection limit of 5.7 nM for xanthine. The novel ratiometric SERS sensor provides a new direction for analyzing other biomolecules with high sensitivity and reliability.

Ultrasensitive detection of thiram based on surface-enhanced Raman scattering via Au@Ag@Ag core/shell/shell bimetallic nanorods.

We developed a highly sensitive and stable SERS-active substrate of Au@Ag@Ag core/shell/shell nanorods, formed by encapsulating Au nanorods (Au NRs) into a bilayer silver shell with Raman reporter molecules (4-mercaptobenzoic acid (4-MBA) and thiram) in the shell-shell gap. The core/shell/shell nanostructures demonstrated a high SERS enhancement and easy assembly. The important role of the bilayer silver shell in boosting the SERS intensity and detection sensitivity was revealed by comparing the performances of the Au@Ag@4-MBA@Ag NRs, Au@Ag@4-MBA NRs, and Au@4-MBA NRs. The obtained Au@Ag@4-MBA@Ag NRs exhibited a significantly promoted SERS intensity, which could reach around 2.6 times and 240 times that of the Au@Ag@4-MBA NRs and Au@4-MBA NRs, where the enhancement factor was found to be strongly correlated with the shell thickness. The controllable plasma properties and SERS effect of the Au@Ag@4-MBA@Ag NRs could be optimized by adjusting the dose of silver nitrate. The SERS substrate comprising core/shell/shell nanorods was highly reproducible and stable (retaining 83% SERS intensity after one month). Moreover, the highly sensitive detection of the pesticide thiram with a detection limit as low as 1.74 × 10-9 M was achieved by taking advantage of the great SERS response of the core/shell/shell nanostructures, which was 1-2 orders of magnitude lower than for other SERS substrates. The developed SERS substrate could be readily extended to embed other target analytes into the core/shell/shell nanostructure for novel and sensitive detection. This study could enable fresh approaches toward next-generation ultrasensitive analyte detection.

The applications of carbon dots in oral health: A scoping review.

OBJECTIVES: This scoping review aims to provide an overview of the research and potential applications of carbon dots (CDs) for oral health purposes. DESIGN: Systematic literature searches were performed on PubMed and Web of Science databases (up to February 2023). Two co-authors selected the published works independently and extracted the data in accordance with the PRISMA statement. Studies with the application of CDs for oral health purposes were included. RESULTS: Among 152 articles, 19 articles were finally selected. Eight studies investigated the anti-microbial effects of CDs against, for example, oral pathogens, eight studies explored the applicability of CDs in relation to oral cancer, and three studies investigated CDs in relation to cell differentiation and tissue regeneration in oral health. The studies showed the promising potential of CDs in oral health, particularly for inducing bacterial killing by increasing reactive oxygen species, killing oral cancer cells via photodynamic therapeutic effects, and inducing dental pulp and periodontal bone regeneration. CONCLUSION: The findings show that CDs have the potential to be utilized in the future for various oral health purposes. Besides, these results underline the broad-spectrum applicability of CDs, crossing the borders of oral health.

Carbon quantum dots as immune modulatory therapy in a Sjögren's syndrome mouse model.

OBJECTIVES: The objective of the study was to evaluate the therapeutic effects of carbon quantum dots (CQDs) in immunomodulation on non-obese diabetic (NOD) mice, as the model for Sjögren's syndrome (SS). METHODS: Carbon quantum dots were generated from Setaria viridis via a hydrothermal process. Their toxic effects were tested by cell viability and blood chemistry analysis, meanwhile therapeutic effects were investigated in NOD mice in the aspects of saliva flow, histology, and immune cell distribution. RESULTS: Carbon quantum dots, with rich surface chemistry and unique optical properties, showed non-cytotoxicity in vitro or no damage in vivo. Intravenously applied CQDs alleviated inflammation in the submandibular glands in NOD mice after 6-week treatments. The inflammatory area index and focus score were significantly decreased in CQD-treated mice. Besides, the levels of anti-SSA and anti-SSB were decreased in the presence of CQDs. The stimulated saliva flow rates and weight of submandibular glands were significantly increased in CQD-treated mice by reducing the apoptosis of cells. The CD3+ and CD4+ T cells distributed around the ducts of submandibular glands were significantly decreased, while the percentage of Foxp3+ cells was higher in CQD-treated mice than that in the control group. CONCLUSIONS: Our findings suggest that CQDs may ameliorate the dysregulated immune processes in NOD mice.

In vitro and in vivo toxicological evaluation of carbon quantum dots originating from Spinacia oleracea.

Food-derived carbon quantum dots (CQDs) can relatively easily be synthesized and chemically manipulated for a broad spectrum of biomedical applications. However, their toxicity may hinder their actual use. Here, Spinacia oleracea-derived CQDs i.e., CQD-1 and CQD-2, were synthesized by means of different shredding methods and followed by a microwave-assisted hydrothermal approach. Subsequently, these CQDs were analyzed in vitro and in an in vivo mice model to test their biocompatibility and potential use as bioimaging agents and for activation of osteogenic differentiation. When comparing CQD-1 and CQD-2, it was found that CQD-1 exhibited 7.6 times higher photoluminescent (PL) emission intensity around 411 nm compared to CQD-2. Besides, it was found that the size distribution of CQD-1 was 2.05 ± 0.08 nm, compared with 2.14 ± 0.04 nm for CQD-2. Upon exposure to human bone marrow-derived mesenchymal stem cells (hBMSCs) in vitro, CQD-1 was endocytosed into the cytoplasm and significantly increased the differentiation of hBMSCs up to 10 µg mL-1 after 7 and 14 days. Apparently, the presence of relatively low doses of CQD-1 showed virtually no toxic or histological effects in the major organs in vivo. In contrast, high doses of CQD-1 (1 mg mL-1) caused cell death in vitro ranging from 35% on day 1 to 80% on day 3 post-exposure, and activated the apoptotic machinery and increased lymphocyte aggregates in the liver tissue. In conclusion, S. oleracea-derived CQDs have the potential for biomedical applications in bioimaging and activation of stem cells osteogenic differentiation. Therefore, it is postulated that CQD-1 from S. oleracea remains potential prospective material at appropriate doses and specifications.

Identification and Characterization of MUC5B Binding Peptides by Phage Display.

OBJECTIVES: MUC5B plays a multifactorial role in oral health. As a consequence, decreased MUC5B output leads to impaired salivary functions and xerostomia. Synthetic combinatorial technologies have been used to develop functional peptide libraries by phage display e.g. for therapeutic purposes. In this light, our primary aim was to identify peptide sequences with specific selectivity for salivary MUC5B in vitro using phage display. Our secondary aims were to analyze their effect on salivary spinnbarkeit in situ and their effect on acid-induced demineralization in vitro. METHODS: MUC5B binding phages were selected by phage display. Peptide affinity to MUC5B was evaluated using MUC5B coated hydroxyapatite (HA) granules. The MUC5B binding peptides (MBPs) were then examined for their effects on salivary spinnbarkeit and protective effect on acid-induced demineralization in vitro. A competitive ELISA was performed to identify the binding epitope on MUC5B using F2, a MUC5B specific antibody. RESULTS: MBP-12 and MBP-14 displayed the highest affinity to MUC5B. MBP-12 mildly stabilized the spinnbarkeit of serous saliva after overnight incubation and of mucous saliva at all timepoints tested. The addition of MBP-12 to a pellicle of unstimulated saliva on HA discs showed no additive protective effect against acid-induced demineralization. Epitope characterization suggested sulfo-Lewisa SO3-3Gal_1-3GlcNAc (galactose residue) as MBP-12 binding site on MUC5B. CONCLUSIONS: The use of phage display in generating MBPs was successful. Characterization of the MBPs revealed a mild effect on spinnbarkeit in case of mucous saliva. Possibly, combinatorial peptide libraries might contribute to the development of novel formulations to treat xerostomia.

Signal amplification surface-enhanced Raman scattering immunosorbent assay of human chorionic gonadotrophin based on repeated enzyme biocatalytic precipitation.

A novel surface-enhanced Raman scattering (SERS) immunoassay method based on tyramine signal amplification (TSA) technology triggering the formation of enzyme repeats on an enzyme-linked immunosorbent assay (ELISA) was designed for highly sensitive detection of human chorionic gonadotropin (hCG) using enzymatic biocatalytic precipitation toward o-phenylenediamine (OPD). Initially, a horseradish peroxidase (HRP)-labeled hCG antibody was fixed by the double antibody sandwich method, and then a tyramine-HRP conjugate was used to form HRP repeats by triggering the immobilized HRP on ELISA with the aid of H2O2. In the presence of the target hCG, the HRP repeats carried by the sandwich immune complex catalyzed the oxidation of OPD to produce product molecules with different structures, resulting in changes in the SERS fingerprint spectrum. The analytical performance of the SERS immunoassay was studied in detail using SERS spectral characterization. Under the optimum conditions, the immunosensor displayed a working range from 1 IU L-1 to 16 IU L-1 with a detection limit (LOD) of 0.17 IU L-1 relative to the target hCG. Compared to the traditional SERS immunosensor, a higher detection sensitivity can be obtained. Therefore, this work provides a new strategy for hCG detection and inspiration for the construction of sensitive and efficient immunosensors.

Direct preparation of solid carbon dots by pyrolysis of collagen waste and their applications in fluorescent sensing and imaging.

The fluorescent carbon dots (CDs) have found their extensive applications in sensing, bioimaging, and photoelectronic devices. In general terms, the synthesis of CDs is straight-forward, though their subsequent purification can be laborious. Therefore, there is a need for easier ways to generate solid CDs with a high conversion yield. Herein, we used collagen waste as a carbon source in producing solid CDs through a calcination procedure without additional chemical decomposition treatment of the raw material. Considering a mass of acid has destroyed the original protein macromolecules into the assembled structure with amino acids and peptide chains in the commercial extraction procedure of collagen product. The residual tissues were assembled with weak intermolecular interactions, which would easily undergo dehydration, polymerization, and carbonization during the heat treatment to produce solid CDs directly. The calcination parameters were surveyed to give the highest conversion yield at 78%, which occurred at 300°C for 2 h. N and S atomic doping CDs (N-CDs and S-CDs) were synthesized at a similar process except for immersion of the collagen waste in sulfuric acid or nitric acid in advance. Further experiments suggested the prepared CDs can serve as an excellent sensor platform for Fe3+ in an acid medium with high anti-interference. The cytotoxicity assays confirmed the biosafety and biocompatibility of the CDs, suggesting potential applications in bioimaging. This work provides a new avenue for preparing solid CDs with high conversion yield.

GPCR/endocytosis/ERK signaling/S2R is involved in the regulation of the internalization, mitochondria-targeting and -activating properties of human salivary histatin 1.

Human salivary histatin 1 (Hst1) exhibits a series of cell-activating properties, such as promoting cell spreading, migration, and metabolic activity. We recently have shown that fluorescently labeled Hst1 (F-Hst1) targets and activates mitochondria, presenting an important molecular mechanism. However, its regulating signaling pathways remain to be elucidated. We investigated the influence of specific inhibitors of G protein-coupled receptors (GPCR), endocytosis pathways, extracellular signal-regulated kinases 1/2 (ERK1/2) signaling, p38 signaling, mitochondrial respiration and Na+/K+-ATPase activity on the uptake, mitochondria-targeting and -activating properties of F-Hst1. We performed a siRNA knockdown (KD) to assess the effect of Sigma-2 receptor (S2R) /Transmembrane Protein 97 (TMEM97)-a recently identified target protein of Hst1. We also adopted live cell imaging to monitor the whole intracellular trafficking process of F-Hst1. Our results showed that the inhibition of cellular respiration hindered the internalization of F-Hst1. The inhibitors of GPCR, ERK1/2, phagocytosis, and clathrin-mediated endocytosis (CME) as well as siRNA KD of S2R/TMEM97 significantly reduced the uptake, which was accompanied by the nullification of the promoting effect of F-Hst1 on cell metabolic activity. Only the inhibitor of CME and KD of S2R/TMEM97 significantly compromised the mitochondria-targeting of Hst1. We further showed the intracellular trafficking and targeting process of F-Hst1, in which early endosome plays an important role. Overall, phagocytosis, CME, GPCR, ERK signaling, and S2R/TMEM97 are involved in the internalization of Hst1, while only CME and S2R/TMEM97 are critical for its subcellular targeting. The inhibition of either internalization or mitochondria-targeting of Hst1 could significantly compromise its mitochondria-activating property.

Horseradish peroxidase-repeat assay based on tyramine signal amplification for highly sensitive H2O2 detection by surface-enhanced Raman scattering.

A new and simple surface-enhanced Raman scattering (SERS) biosensor based on the tyramine signal amplification (TSA)-triggered formation of horseradish peroxidase (HRP) repeats on a gold sensing chip was designed for the highly sensitive detection of hydrogen peroxide (H2O2). Initially, gold wafers were functionalized with HRP as sensing chips. Then, the HRP immobilized on the chips triggers the TSA reaction to transform the tyramine-HRP conjugate into a tyramine-HRP repeat array. With the aid of the target H2O2, the HRP repeats catalyze the oxidation of o-phenylenediamine (OPD) and produce an enzyme catalytic product with a different chemical structure, thus altering the fingerprint of the SERS spectra from that of OPD. H2O2 can be quantitatively analyzed according to the change in SERS signal intensity. On the basis of the TSA strategy, the proposed method allows the detection of H2O2 with a limit of detection (LOD) of 0.8 × 10-8 M. The as-prepared SERS sensor can achieve high-sensitivity H2O2 detection with a small amount of sample for each analysis. Therefore, this sensor exhibits significant potential for application in bioanalysis.

Recent advances in catalytic hairpin assembly signal amplification-based sensing strategies for microRNA detection.

Accumulative evidences have indicated that abnormal expression of microRNAs (miRNAs) is closely associated with many health disorders, making them be regarded as potentialbiomarkers for early clinical diagnosis. Therefore, it is extremely necessary to develop a highly sensitive, specific and reliable approach for miRNA analysis. Catalytic hairpin assembly (CHA) signal amplification is an enzyme-free toehold-mediated strand displacement method, exhibiting significant potential in improving the sensitivity of miRNA detection strategies. In this review, we first describe the potential of miRNAs as disease biomarkers and therapeutics, and summarize the latest advances in CHA signal amplification-based sensing strategies for miRNA monitoring. We describe the characteristics and mechanism of CHA signal amplification and classify the CHA-based miRNA sensing strategies into several categories based on the "signal conversion substance", including fluorophores, enzymes, nanomaterials, and nucleotide sequences. Sensing performance, limit of detection, merits and disadvantages of these miRNA sensing strategies are discussed. Moreover, the current challenges and prospects are also presented.

First Report of Anthracnose on Jerusalem Cherry Caused by Colletotrichum liaoningense in Shandong, China.

Jerusalem cherry (Solanum pseudocapsicum), which belongs to the genus Solanum and the family Solanaceae, possesses high ornamental value and is widely cultivated as an indoor ornament due to its bright red berries at maturity (Xu et al., 2018). In September 2019, leaf spot was detected on jerusalem cherry plants in Yuxiu Park, Shizhong district, Jinan, Shandong Province. Field surveys were done in a 1/15 ha park. Disease incidence was estimated at approximately 18% across the survey area. Foliar symptoms began as small white spots. As the disease progressed, lesions expanded and merged, and developed into large irregular white spots, with pale grey edge. At last, lesions were densely distributed throughout the leaves. To isolate the pathogen, twenty leaf tissues (5 × 5 mm) were cut from the border between diseased and healthy tissue, surface disinfected in 75% alcohol for 15 s, soaked in 0.1% mercuric chloride for 1 min, washed with sterile distilled water three times, and cultured on potato dextrose agar (PDA) at 25°C. Nineteen fungal isolates were obtained and were single-spored to obtain pure cultures. The colony of LCL7, a representative isolate, on PDA was initially white to orange, but turned black after 3 to 4 days incubation with black conidial masses. Conidia were single-celled, hyaline, straight, cylindrical, apex obtuse, and ranged from 13.4 to 18.3 × 3.2 to 4.9 µm (n = 50) （Diao et al., 2017）. To validate the species identification, rDNA internal transcribed spacer (ITS) region (White et al., 1990), and the partial sequences of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), actin (ACT), ß-tubulin (TUB2), and chitin synthase (CHS-1) (Damm et al., 2019; He et al., 2019), were amplified and sequenced. The ITS, GAPDH, ACT, TUB2, and CHS-1 sequences of isolate LCL7 were submitted to GenBank (MW221320, MW227217, MW227218, MW227219, and MW266988, respectively). ITS, ACT, TUB2, and CHS-1 BLAST showed 99-100% homology with sequences of Colletotrichum liaoningense (ITS, 100% to MH636504; ACT, 100% to MH622582; TUB2, 99.56% to MH622714, CHS-1, 99.33% to MH622446, respectively), although GAPDH showed 93.98% homology with sequence MH681383 (234/249bp). Neighbor-joining tree based on concatenated sequences of the five genes was constructed using MEGA7.0. The results showed the isolate was closely related to C. liaoningense. Based on morphological and molecular characteristics, the isolate LCL7 was identified as C. liaoningense. Pathogenicity tests were performed by spraying a conidial suspension (1 × 105 conidia/mL) on ten two-year-old healthy jerusalem cherry plants. Ten other plants with sterile water served as controls. All samples were incubated in a growth chamber at 25±2°C and transparent plastic bags to keep relative humidity high for 2 days. All inoculated plants showed symptoms similar to those observed in the field after 21 days, but no disease occurred on control plants. The same fungus was successfully reisolated from inoculated leaves and reidentified based on morphology and molecular characteristics, and the fungus was not isolated from the control plants, thus confirming Koch's postulates. Pathogenicity tests were repeated twice. C. liaoningense can cause anthracnose in chili and mango in China (Diao et al., 2017; Li et al., 2019).To our knowledge, this is the first report of anthracnose on jerusalem cherry caused by C. liaoningense in China, which influences ornamental value and reduces market value. Identification of the causes of the disease will help develop effective strategies for managing this disease.

Macrophage-derived small extracellular vesicles promote biomimetic mineralized collagen-mediated endogenous bone regeneration.

Macrophages play an important role in material-related immune responses and bone formation, but the functionality of macrophage-derived extracellular vesicles (EVs) in material-mediated bone regeneration is still unclear. Here, we evaluated intracellular communication through small extracellular vesicles (sEVs) and its effects on endogenous bone regeneration mediated by biomimetic intrafibrillarly mineralized collagen (IMC). After implantation in the bone defect area, IMC generated more neobone and recruited more mesenchymal stem cells (MSCs) than did extrafibrillarly mineralized collagen (EMC). More CD63+CD90+ and CD63+CD163+ cells were detected in the defect area in the IMC group than in the EMC group. To determine the functional roles of sEVs, extracellular vesicles from macrophages cultured on different mineralized collagen were isolated, and they showed no morphological differences. However, macrophage-derived sEVs in the IMC group showed an enhanced Young's modulus and exerted beneficial effects on the osteogenic differentiation of bone marrow MSCs by increasing the expression of the osteoblastic differentiation markers BMP2, BGLAP, COL1, and OSX and calcium nodule formation. Mechanistically, sEVs from IMC-treated macrophages facilitated MSC osteogenesis through the BMP2/Smad5 pathway, and blocking sEV secretion with GW4869 significantly impaired MSC proliferative, immunomodulative and osteogenic potential. Taken together, these findings show that macrophage-derived sEVs may serve as an emerging functional tool in biomaterial-mediated endogenous bone regeneration.

Anion-Regulated Synthesis of ZnO 1D Necklace-Like Nanostructures with High Photocatalytic Activity.

One-dimensional (1D) nanomaterials with specific architectures have received increasing attention for both scientific and technological interests for their applications in catalysis, sensing, and energy conversion, etc. However, the development of an operable and simple method for the fabrication of 1D nanostructures remains a challenge. In this work, we developed an "anion-regulated morphology" strategy, in which anions could regulate the dimensionally-restricted anisotropic growth of ZnO nanomaterials by adjusting the surface energy of different growth facets. ZnO 1D necklace-like nanostructures (NNS) could be prepared through a hydrothermal treatment of zinc acetate and urea mixture together with a subsequent calcination procedure at 400 °C. While replacing the acetate ions to nitrate, sulfate, and chlorion ions produced ZnO nanoflowers, nanosheets and hexagonal nanoplates, respectively. Density functional theory calculations were carried out to explain the mechanism behind the anions-regulating anisotropic crystal growth. The specified ZnO 1D NNS offered improved electron transport while the grain surface could supply enlarged specific surface area, thus providing advanced photocatalytic ability in the following photodegradation of methyl orange (MO). Among the four photocatalysts with different morphologies, ZnO 1D NNS, possessing the highest catalytic activity, degraded 57.29% MO in the photocatalytic reaction, which was 2 times, 10 times and 17 times higher than nanoflowers, nanosheets and hexagonal nanoplates, respectively. Our work provides new ideas for the construction and application of ZnO 1D nanomaterials.

"Signal-on" SERS sensing platform for highly sensitive and selective Pb2+ detection based on catalytic hairpin assembly.

In this work, a surface-enhanced Raman scattering (SERS) sensing strategy was proposed for the analysis of lead ion (Pb2+) with high sensitivity and specificity based on the high specificity of the catalytic nucleic acids (DNAzymes) to Pb2+ and the catalytic hairpin assembly (CHA) amplification. First, the Pb2+-DNAzyme initiated the reaction by target Pb2+ and released a linear DNA (rS1). Second, the hairpin DNA 1 (H1) was immobilized on the SERS substrate surface via Ag-S bond. Then, the rS1 could cyclically trigger the allosteric effects of CHA amplification and the H1 was opened and then the R6G-labeled hairpin probe 2 (H2) hybridized with H1 to form H1-H2 double-stranded and the released rS1 could initiate the next cycle of CHA reaction. This process made the Raman tag of R6G close to the surface of SERS substrate, and the intensity of SERS signal from R6G labels increase with the increase of concentration of target Pb2+. Benefiting from outstanding performances of the Pb2+-specific DNAzymes and enzyme-free CHA amplification system, this biosensor exhibits good sensitivity for Pb2+ with a limit of detection of 0.42 pM. More importantly, this developed detection platform could be employed for reliable analysis of Pb2+ in real environment system.

Lengthening the aptamer to hybridize with a stem-loop DNA assistant probe for the electrochemical detection of kanamycin with improved sensitivity.

By adding 6 thymines to lengthen the parent aptamer combined with the change of "on" and "off" induced by the target for an assistant stem-loop DNA probe (ASP-SLP-MB), a new folding-type electrochemical kanamycin (Kana) aptamer-engineering dual-probe-based sensor (sensor d) was developed. By purposefully reducing the background current and increasing the electron transfer efficiency of methylene blue (MB), the sensor obtained significantly enhanced detection sensitivity compared with non-aptamer-engineering one-probe-based sensor (sensor a). Such efficacy was validated by a big decrease from 530.6 to 210.2 nA for the background current signal and from 360 to 0.3 nM for the detection limit. In addition to the improved sensitivity, the sensor also exhibited good selectivity, anti-fouling detection performance, and potential quantitative analysis ability, showing a feasible potential practical analytical application in real-life complicated samples, for example, milk and serum. The released results prove that the aptamer-engineering method is effective in improving the analytical performance of folding-type sensors and provides a methodological guidance for the design and fabrication of other high-performance folding-type aptasensors. Graphical abstract.

A simple enzyme-free SERS sensor for the rapid and sensitive detection of hydrogen peroxide in food.

A simple enzyme-free method based on surface-enhanced Raman scattering (SERS) was developed for the first time to detect H2O2 in food by etching a self-assembled film of silver nanoparticles (Ag NPs) on a glass substrate. H2O2 is able to oxidize Ag NPs to yield Ag+ ions; this process reduces the size of the Ag NPs and ultimately leads to a decrease in the SERS signal of the Raman probe. The intensities of the SERS spectra were strongly correlated with H2O2 concentration, which indicated that the Ag NP self-assembled SERS sensor can be successfully used for the quantitative analysis of H2O2. The main advantage of this SERS sensor is that it can directly detect H2O2 without introducing complex enzymatic reactions. This easy-to-operate and fast-response detection technology has great potential for the sensitive detection of H2O2 in food.

Ratiometric SERS biosensor for sensitive and reproducible detection of microRNA based on mismatched catalytic hairpin assembly.

MicroRNAs (miRNAs) serve as significant regulators in a variety of diseases and have been emerging as a class of promising biomarkers for early cancer diagnosis. Herein, an enzyme-free surface-enhanced Raman scattering (SERS) platform was proposed for sensitive and reliable detection of target miRNA-21 using a corrective internal standard (IS)-based ratiometric SERS probe coupled with mismatched catalytic hairpin assembly (CHA) amplification. The 4-aminothiophenol (4-ATP) was used as IS molecule and modified on the surface of silver nanoparticles decorated silicon wafer. In principle, the presence of miRNA-21 could cyclically trigger the allosteric effects of mismatched CHA amplification and the 3'-R6G labeled hairpin probe 1 (H1) was opened. Then, the hairpin probe 2 (H2) hybridized with H1 to form H1-H2 complex and the released miRNA-21 was free to participate in the next cycle of CHA reaction. Meanwhile, the H1-H2 complex could hybridize with the capture DNA on the SERS chip, making the Raman tag of R6G close to the surface of SERS substrate, and the intensity of SERS signal from R6G labels increase while that from 4-ATP remain relatively unchanged. Benefiting from outstanding performances of the ratiometric SERS strategy and enzyme-free CHA amplification system, this platform exhibits sensitivity toward miRNA-21 with a limit of detection of 3.5 fM and a broad dynamic range from 10 fM to 100 nM. More importantly, this proposed method presents an excellent reliable SERS analysis with the correction of IS. The developed strategy holds a potential alternative tool for miRNA detection in biomedical research and early clinical diagnosis.