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Electrochemical aptamer-based (EAB) sensors are a molecular measurement platform that enables the continuous, real-time measurement of a wide range of drugs and biomarkers in situ in the living body. EAB sensors are fabricated by depositing a thiol-modified, target-binding aptamer on the surface of a gold electrode, followed by backfilling with an alkanethiol to form a self-assembled monolayer. And while the majority of previously described EAB sensors have employed hydroxyl-terminated monolayers, a handful of studies have shown that altering the monolayer headgroup can strongly affect sensor performance. Here, using 4 different EAB sensors, we show that the mixed monolayers composed of mixtures of 6-carbon hydroxyl-terminated thiols and varying amounts of either 6- or 8-carbon, carboxylate-terminated thiols lead to improved EAB sensor performance. Specifically, the use of such mixed monolayers enhances the signal gain (the relative change in the signal seen upon target addition) for all tested sensors, often by several fold, both in buffer and whole blood at room temperature or physiological temperatures. Moreover, these improvements in gain are achieved without significant changes in the aptamer affinity or the stability of the resulting sensors. In addition to proving a ready means of improving EAB sensor performance, these results suggest that exploration of the chemistry of the electrode surface employed in such sensors could prove to be a fruitful means of advancing this unique in vivo sensing technology.

Cryo-EM structure determination of protein-free RNAs has remained difficult with most attempts yielding low to moderate resolution and lacking nucleotide-level detail. These difficulties are compounded for small RNAs as cryo-EM is inherently more difficult for lower molecular weight macromolecules. Here we present a strategy for fusing small RNAs to a group II intron that yields high resolution structures of the appended RNA. We demonstrate this technology by determining the structures of the 86-nucleotide (nt) thiamine pyrophosphate (TPP) riboswitch aptamer domain and the recently described 210-nt raiA bacterial non-coding RNA involved in sporulation and biofilm formation. In the case of the TPP riboswitch aptamer domain, the scaffolding approach allowed visualization of the riboswitch ligand binding pocket at 2.5 Å resolution. We also determined the structure of the ligand-free apo state and observe that the aptamer domain of the riboswitch adopts an open Y-shaped conformation in the absence of ligand. Using this scaffold approach, we determined the structure of raiA at 2.5 Å in the core. Our versatile scaffolding strategy enables efficient RNA structure determination for a broad range of small to moderate-sized RNAs, which were previously intractable for high-resolution cryo-EM studies.

RNA sequences encode structures that impact protein production and other cellular processes. Misfolded RNAs can also potentiate disease, but a complete picture is lacking. To establish more comprehensive and accurate RNA structure-function relationships, new methods are needed to interrogate RNA in native environments. Existing tools rely primarily on electrophiles that are constitutively on or triggered by UV light, often resulting in high background. Here we describe an alternative, chemically triggered approach to cross-link RNAs using bioorthogonal cyclopropenones (CpOs). These reagents selectively react with phosphines to provide ketenes─electrophiles that can trap neighboring nucleophiles to forge covalent cross-links. As a proof-of-concept, we conjugated a CpO motif to thiazole orange (TO-1). TO-1-CpO bound selectively to a model RNA aptamer (Mango) with nanomolar affinity, as confirmed by fluorescence turn-on. After phosphine administration, covalent cross-links were formed between the CpO and RNA. Cross-linking was both time and dose dependent. We further applied the chemically triggered tools to model RNAs under biologically relevant conditions. Collectively, this work expands the toolkit of probes for studying RNA and its native conformations.

Condensation of RNA and proteins is central to cellular functions, and the ability to program it would be valuable in synthetic biology and synthetic cell science. Here we introduce a modular platform for engineering synthetic RNA condensates from tailor-made, branched RNA nanostructures that fold and assemble co-transcriptionally. Up to three orthogonal condensates can form simultaneously and selectively accumulate fluorophores through embedded fluorescent light-up aptamers. The RNA condensates can be expressed within synthetic cells to produce membrane-less organelles with a controlled number and relative size, and showing the ability to capture proteins using selective protein-binding aptamers. The affinity between otherwise orthogonal nanostructures can be modulated by introducing dedicated linker constructs, enabling the production of bi-phasic RNA condensates with a prescribed degree of interphase mixing and diverse morphologies. The in situ expression of programmable RNA condensates could underpin the spatial organization of functionalities in both biological and synthetic cells.

Recent discoveries in biology have highlighted the importance of protein and RNA-based condensates as an alternative to classical membrane-bound organelles. Here, we demonstrate the design of pure RNA condensates from nanostructured, star-shaped RNA motifs. We generate condensates using two different RNA nanostar architectures: multi-stranded nanostars whose binding interactions are programmed via linear overhangs, and single-stranded nanostars whose interactions are programmed via kissing loops. Through systematic sequence design, we demonstrate that both architectures can produce orthogonal (distinct and immiscible) condensates, which can be individually tracked via fluorogenic aptamers. We also show that aptamers make it possible to recruit peptides and proteins to the condensates with high specificity. Successful co-transcriptional formation of condensates from single-stranded nanostars suggests that they may be genetically encoded and produced in living cells. We provide a library of orthogonal RNA condensates that can be modularly customized and offer a route toward creating systems of functional artificial organelles for the task of compartmentalizing molecules and biochemical reactions.

A molecular beacon is an oligonucleotide hybridization probe that can report the presence of specific nucleic acids in homogeneous solutions. Using an aptamer has allowed an aptamer-based molecular beacon–aptamer beacon to be developed, which has shown advantages of simplicity, rapidity, and sensitivity in imaging and sensing non-nucleic acid substances. However, due to requirement for a deliberate DNA hairpin structure for the preparation of a molecular beacon, not any given aptamer is suitable for designing an aptamer beacon probe. This paper provides a general design strategy for the preparation of an aptamer beacon probe, which theoretically can be used for any given aptamer. Through coupling an aptamer and a short complementary DNA into one DNA molecule via a rational poly thymidine (T) linker, novel molecular beacon probes are successfully prepared and used for the detection of targets (aflatoxin B1 and ochratoxin A). The working mechanism of this aptamer beacon probe is based on intramolecular hybridization/dehybridization, which is more efficient than commonly aptasensor strategies based on intermolecular reactions. This aptamer beacon probe shows advantages of low background, a signal-on response, a large signal change, as well as simplicity and rapidity of analysis, which have promising application potential. [ABSTRACT FROM AUTHOR]

The overexpression of Human Epidermal Growth Factor Receptor 2 (HER2) protein is specifically related to tumor cell proliferation in breast cancers. Its presence in biological serum samples indicates presence or progression of cancer, becoming a promise biomarker. However, their detection needs a simple and high accuracy platform. In this study, we report the develop and optimization of a simple highly sensitive electrochemical platform for HER2. Gold electrode surface was modified with a self-assembled monolayer composed by DNA aptamer, 6-(ferrocenyl) hexanethiol and 6-mercapto-1-hexanethiol. Electrochemical impedance spectroscopy was used to quantify the changes in capacitance on the interface due to the presence ferrocene, whether acting as a redox charge or its behavior under different HER2 concentration in PBS and undiluted human serum. As a result, the approach allows detection of HER2 with a limit of detection of 3.61 pg/mL, 12.28 nF sensitivity per decade and a linear range from 1 pM to 1 M in serum. This electrochemical aptasensor can be applied to different arrays for aptamer screening and has a significant importance to interaction study of biological systems. [ABSTRACT FROM AUTHOR]

The development of aptamers has been almost exclusively performed based on the SELEX method since their inception. While this method represents a powerful means of harnessing the in vitro evolution of sequences that bind to a given target, there are significant constraints in the design. The most significant constraint has been the reliance on counter selection on off-targets to drive specificity. Counter selection has not been as effective at driving aptamer specificity as the presence of immune tolerance, the capacity of the immune system to remove antibodies that bind to host targets, is for antibody development. This deficiency has constrained the commercial efficacy of aptamers to date. These limitations have been addressed with our design of a novel platform for aptamer identification. This new design is based on what we refer to as a Neomer library with sixteen random nucleotides interspersed with fixed sequences. The fixed sequences are designed to minimize the potential for hybridization, such that secondary structure is driven by the random nucleotides. The use of sixteen random nucleotides reduces the possible library sequences to 4.29 × 109. This enables the application of the same sequences to either the same target or different targets while maintaining a high level of structural diversity. In effect, this introduces the capacity for reproducibility in aptamer selection and an in-silico approach to replicating immune tolerance. We provide here an overview of the new method and a description of the performance of aptamers selected for interleukin 6 developed using this approach. [ABSTRACT FROM AUTHOR]

DNA-based therapies are often limited by challenges such as stability, long-term integration, low transfection efficiency, and insufficient targeted DNA delivery. This review focuses on recent progress in the design of non-viral delivery systems for enhancing targeted DNA delivery and modulation of therapeutic efficiency. Cellular uptake and intracellular trafficking mechanisms play a crucial role in optimizing gene delivery efficiency. There are two main strategies employed to improve the efficiency of gene delivery vectors: (i) explore different administration routes (e.g., mucosal, intravenous, intramuscular, subcutaneous, intradermal, intratumoural, and intraocular) that best facilitates optimal uptake into the targeted cells and organs and (ii) modify the delivery vectors with cell-specific ligands (e.g., natural ligands, antibodies, peptides, carbohydrates, or aptamers) that enable targeted uptake to specific cells with higher specificity and improved biodistribution. We describe how recent progress in employing these DNA delivery strategies is advancing the field and increasing the clinical translation and ultimate clinical application of DNA therapies. [Display omitted] [ABSTRACT FROM AUTHOR]

A nanozyme-based aptasensor combines the unique properties of nanozymes with the specificity of aptamers for the detection of various biomolecules. Nanozymes are nanomaterials that possess enzyme-like properties, demonstrating substantial potential for enhancing the sensing capabilities of biosensors. In recent years, the incorporation of nanozymes into biosensors has opened new avenues for the detection of tumor biomarkers. The unique attributes of nanozymes and aptamers lead to biosensors characterized by high sensitivity, specificity, reproducibility and accuracy in analytical performance. This article reviews the research progress of nanozyme-based aptasensors in tumor biomarker detection over the past decade. We categorize these sensors based on their sensing modes and target types, and examine the properties and applications of the nanozymes employed in these devices, providing a thorough discussion of the strengths and weaknesses associated with each sensor type. Finally, the review highlights the strengths and challenges associated with nanozyme-based biosensors and envisions future developments and applications in this field. The objective is to provide insights for improving biosensor performance in tumor biomarker detection, thereby contributing to advancements in precision cancer diagnosis and treatment. [ABSTRACT FROM AUTHOR]

Early detection of biomarkers has a crucial role in preventing blindness caused by diabetic retinopathy (DR). In this work, we developed a rapid, highly sensitive method for the simultaneous detection of DR-related biomarkers (miR-21 and miR-152) based on an Au nano-hexagon (AuNHs) integrated microarray chip combined with a competitive recognition strategy. The nucleic acid aptamer-modified AuNHs array served as capture substrate, while Au nano-trisoctahedrons (AuNTs) with complementary strands of nucleic acid aptamers mounted on their surface and Raman signaling molecules served as SERS probes (AuNTs@4-MBA@H1 and AuNTs@DTNB@H2). SERS probes carrying Raman signaling molecules (4-MBA and DTNB) were displaced by the target miRNAs in the experiments because they had a higher binding affinity to nucleic acid aptamers than their structural analogs (H1 and H2). This approach led to a notable decrease in the Raman signal. The competitive recognition strategy was straightforward and useful, and required only a one-step dropwise reaction to detect both miRNAs simultaneously and effectively. The microarray chip designed accordingly had the advantages of sensitive, rapid and high-throughput detection, and the detection time was only 8 min. The limits of detection (LOD) for miR-21 and miR-152 were as low as 2.76 × 10−14 M and 1.94 × 10−14 M, respectively. The utility of the method in complex systems was verified by detecting miRNAs in the serum of DR rats, and the accuracy of the method was verified against the detection results of a RT-qPCR method. This method provides a new idea and manner for the detection of miRNAs in serum, which is crucial for the early diagnosis of DR. [ABSTRACT FROM AUTHOR]

An aptasensor was developed based on surface-enhanced Raman spectroscopy (SERS) to detect alpha-fetoprotein (AFP) using a mirror aptamer. AFP triggered and altered SERS signaling molecules through the release recognition mechanism as they are released. Two signaling molecules, cyanine 3 (Cy3) and 2-mercaptobenzimidazole-5-carboxylic acid (MBIA), were used to quantify AFP. The results showed that AFP could be reliably and sensitively detected in the concentration range 0.0038–100 ng/mL (R2 = 0.9907), with a minimum detection limit of 3.8 pg/mL. The sensor can be used for AFP detection in cancer patient serum samples. This quantitative SERS technology provides a promising tool for biomedical research and clinical diagnosis. [ABSTRACT FROM AUTHOR]

A self-powered dual-electrode aptasensor was developed for the detection of tumor marker carcinoembryonic antigen (CEA). The composite BiVO4/ZnIn2S4, which is capable of forming a Z-scheme heterojunction, was chosen as the photoanode, and the AuNP/CuBi2O4 complex was chosen as the photocathode in photoelectrochemical (PEC) detection. The experiments showed that the constructed self-powered dual-electrode system had a good photoelectric response to white light, and the photocurrent signal of the photocathode was significantly enhanced under the influence of the photoanode. The introduction of G-quadruplex/hemin DNAzyme through aptamer recognition resulted in the generation of insoluble species through catalytic reaction, which led to a significant decrease in the photocurrent, and achieved a highly sensitive detection of the target CEA with a detection limit of 0.021 pg/mL. At the same time, the developed photocathode detection exhibited good selectivity and could effectively avoid interference, making it suitable for real sample analysis. It is expected that this study can facilitate the application of PEC sensors in point-of-care (POC) diagnosis for life analysis. Self-powered dual-photoelectrode photoelectrochemical aptasensor amplified by hemin/ G-quadruplex-based DNAzyme [ABSTRACT FROM AUTHOR]

An innovative colorimetric sensing strategy was developed for the detection of glucose by the integration of glucose aptamer, glucose oxidase (GOx), and horseradish peroxidase (HRP), termed aptamer proximal enzyme cascade reactions (APECR). In the presence of glucose, aptamer binding enables GOx to catalyze glucose oxidation into H2O2 efficiently. Subsequently, the adjacent HRP catalyzes the oxidation of the peroxidase substrate, 2,2′-biazobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS2−), utilizing the generated H2O2, resulting in a distinct color change. In comparison to the free enzymes and the HRP-GOx system, APECR exhibited higher colorimetric signal. This approach achieved glucose detection within three minutes, which was significantly faster than previous methods. This method showed good sensitivity and selectivity with a limit of detection of 0.013 mM. Moreover, the practical utility of this strategy was verified by achieving rapid detection of glucose in clinical serum samples. Hence, the developed strategy has the advantages of simple operation and rapid analysis time for the detection of glucose in human serum. [ABSTRACT FROM AUTHOR]

We present a lateral flow aptasensor for the visual detection of alpha-fetoprotein (AFP) in human serum. Leveraging the precise molecular recognition capabilities of aptamers and the distinct optical features of gold nanoparticles, a model system utilizing AFP as the target analyte, along with a pair of aptamer probes, is implemented to establish proof-of-concept on standard lateral flow test strips. It is the first report of an antibody-free lateral flow assay using aptamers as recognition probes for the detection of AFP. The analysis circumvents the numerous incubation and washing steps that are typically involved in most current aptamer-based protein assays. Qualitative analysis involves observing color changes in the test area, while quantitative data are obtained by measuring the optical response in the test zone using a portable strip reader. The biosensor exhibits a linear detection range for AFP concentrations between 10 and 100 ng/mL, with a minimum detection limit of 10 ng/mL. Additionally, it has been successfully applied to detect AFP in human serum samples. The use of aptamer-functionalized gold nanoparticle probes in a lateral flow assay offers great promise for point-of-care applications and fast, on-site detection. [ABSTRACT FROM AUTHOR]

Exosomes carry diverse tumor-associated molecular information that can reflect real-time tumor progression, making them a promising tool for liquid biopsy. However, traditional methods for exosome isolation and detection often rely on large, expensive equipment and are time-consuming, limiting their practical applicability in clinical settings. Microfluidic technology offers a versatile platform for exosome analysis, with advantages such as seamless integration, portability and reduced sample volumes. Aptamers, which are single-stranded oligonucleotides with high affinity and specificity for target molecules, have been frequently employed in the development of aptamer-based microfluidics for the isolation, signal amplification, and quantitative detection of exosomes. This review summarizes recent advances in aptamer-based microfluidic strategies for exosome analysis, including (1) strategies for on-chip exosome capture mediated by aptamers combined with nanomaterials or nanointerfaces; (2) aptamer-based on-chip signal amplification techniques, such as enzyme-free hybridization chain reaction (HCR), rolling circle amplification (RCA), and DNA machine-assisted amplification; and (3) various aptamer-assisted detection methods, such as fluorescence, electrochemistry, surface-enhanced Raman scattering (SERS), and magnetism. The limitations and advantages of these methods are also summarized. Finally, future challenges and directions for the clinical analysis of exosomes based on aptamer-based microfluidics are discussed. [ABSTRACT FROM AUTHOR]

The epidermal growth factor receptor (EGFR) is one of the key oncomarkers in glioblastoma (GB) biomedical research. High levels of EGFR expression and mutations have been found in many GB patients, making the EGFR an attractive target for therapeutic treatment. The EGFRvIII mutant is the most studied, it is not found in normal cells and is positively associated with tumor cell aggressiveness and poor patient prognosis, not to mention there is a possibility of it being a tumor stem cell marker. Some anti-EGFR DNA aptamers have already been selected, including the aptamer U2. The goal of this study was to construct a more stable derivative of the aptamer U2, while not ruining its functional potential toward cell cultures from GB patients. A multiloop motif in a putative secondary structure of the aptamer U2 was taken as a key feature to design a novel minimal aptamer, Gol1, using molecular dynamics simulations for predicted 3D models. It turned out that the aptamer Gol1 has a similar putative secondary structure, with G-C base pairs providing its stability. The anti-proliferative activities of the aptamer Gol1 were assessed using patient-derived GB continuous cell cultures, G01 and BU881, with different abundances of EGFR and EGFRvIII. The transcriptome data for the cell culture G01, after aptamer Gol1 treatment, revealed significant changes in gene expression; it induced the transcription of genes associated with neurogenesis and cell differentiation, and it decreased the transcription of genes mediating key nuclear processes. There were significant changes in the gene transcription of key pro-oncogenic signaling pathways mediated by the EGFR. Therefore, the aptamer Gol1 could potentially be an efficient molecule for translation into biomedicine, in order to develop targeted therapy for GB patients. [ABSTRACT FROM AUTHOR]

AbstractKanamycin (KANA) plays a key role in the treatment of bacterial infections and has been widely used in animal husbandry. However, its overuse causes antibiotic residues in animal-derived foods. Determination methods for KANA are urgently needed for food safety. Most of the developed fluorescent aptamer sensors for detecting KANA use parental aptamer (kana-Apt) as recognition unit. However, excessive bases tend to form secondary structures and lead to high background or nonspecific signals. In this study, two fluorescent sensors based on one (kana1-Apt) and two (kana1/kana2-Apt) split fragments were developed for KANA detection. The LODs of the kana1-Apt/ThT system and kana1/kana2-Apt/ThT systems were 4.88 nM and 4.53 nM, respectively. In addition, satisfactory recoveries of the kana1-Apt/ThT system and kana1/kana2-Apt/ThT system were obtained in the detection of KANA in milk, which were 97.6%-104.5% and 98.4%-105.9%, respectively. Moreover, the results indicated that the kana1-Apt fragment plays a critical role in recognition. In conclusion, the results of the present study provide a novel strategy for molecular detection based on split aptamers. [ABSTRACT FROM AUTHOR]

The article "Discovery of a Novel Cerebrospinal Fluid-Specific DNA Aptamer" published in the Journal of Neurological Surgery. Part B. Skull Base discusses the development of a DNA aptamer for detecting cerebrospinal fluid (CSF) leaks. Through Systematic Evolution of Ligands by EXponential enrichment (SELEX), two aptamers (C2 and C3) with high selectivity for CSF-specific biomarkers were identified. These aptamers show promise for integration into an electrochemical platform for point-of-care diagnostic devices to detect CSF leaks. [Extracted from the article]

Metal–organic frameworks (MOFs), as a class of crystalline organic–inorganic hybrid materials, have demonstrated excellent performance in many fields, resulting in attracting more and more attention. Recently, MOFs have been widely used as functional materials to immobilize aptamers via various interaction forces. The MOF-based aptasensors exhibit outstanding sensing performance, including high sensitivity, excellent selectivity, and good designability, by utilizing the advantages of MOFs and aptamers. Developing MOF-based aptasensors is gradually becoming one of the important research hotspots. Herein, a comprehensive review of aptasensors based on MOFs is provided to summarize recent advancements in this field. Some representative examples are discussed in detail. Furthermore, opportunities and challenges in this field are proposed in this review. [ABSTRACT FROM AUTHOR]

[Display omitted] Multifunctional micelles that permit both diagnosis and treatment present enormous advantage and potential for precision medicine. However, the inherent complexities and structural instability of these systems often cause unsatisfactory targeting and therapeutic performances. Herein, by ingenious design of a 2,5-bis(2-thienyl)pyrrole (SNS) modifier to covalently link with AS1411 aptamer and lipid segment, a simple strategy is proposed for one-step enzymatic preparation of interlocked aptamer-micelle (IApM) under bio-friendly conditions. The interlocked poly(SNS) skeleton in IApM can not only stabilize the micelle structure but also enhance near-infrared (NIR) absorption ability, thus further enhancing cellular internalization and photothermal therapy. In addition, the multivalent AS1411 aptamers tethered in the hydrophilic shell can simultaneously increase the specific binding affinity of DNA micelles and induce nucleus-targeted accumulation for DNA damage-triggered apoptosis. This DNA micelle achieves "best of both worlds" with enhanced biostability for cellular internalization and improved NIR photothermal conversion efficiency for nucleus-targeted therapy, which provides a promising formulation strategy for precision cancer treatment. [ABSTRACT FROM AUTHOR]

Flavokavain B (FKB), a hepatotoxic chalcone from Piper methysticum (kava), has raised safety concerns due to its role in disrupting redox homeostasis and inducing apoptosis in hepatocytes. Conventional chromatographic methods for FKB detection, while sensitive, are costly and impractical for field applications. In this work, DNA aptamers were selected using the library-immobilized method and high-throughput sequencing. Three families of aptamers were obtained, and the best one named FKB-S showed a dissociation constant (KD) of 280 nM using microscale thermophoresis. To demonstrate its practical utility, a rapid and label-free colorimetric aptasensor was developed based on aptamer-induced gold nanoparticle aggregation. This assay achieved a detection limit of 150 nM (43.46 ng/mL) and provided results within 10 min. Compared to traditional chromatographic methods, the aptasensor offers a simple, cost-effective, and equipment-free approach for on-site FKB detection, making it a promising tool for the quality control and safety monitoring of kava-based products in diverse environments. [ABSTRACT FROM AUTHOR]

Real-time monitoring of therapeutic drugs is crucial for treatment management and pharmacokinetic studies. We present the optimization and affinity tuning of split-aptamer sandwich assay for real-time monitoring of the narrow therapeutic window drug vancomycin, using surface plasmon resonance (SPR). To achieve reversible, label-free sensing of small molecules by SPR, we adapted a vancomycin binding aptamer in a sandwich assay format through the split-aptamer approach. By evaluating multiple split sites within the secondary structure of the original aptamer, we identified position 27 (P27) as optimal for preserving target affinity, ensuring reversibility, and maximizing sensitivity. The assay demonstrated robust performance under physiologically relevant ranges of pH and divalent cations, and the specific ternary complex formation of the aptamer split segments with the analyte was confirmed by circular dichroism spectroscopy. Subsequently, we engineered a series of split-aptamer pairs with increasing complementarity in the stem regions, improving both the affinity and limit of detection up to 10-fold, as compared to the primary P27 pair. The kinetics of the engineered split-aptamer pairs were evaluated, revealing fast association and dissociation rates, confirming improved affinity and detection limits across variants. Most importantly, the reversibility of the assay, essential for real-time monitoring, was maintained in all pairs. Finally, the assay was further validated in complex biological matrices, including the cerebrospinal fluid from dogs and diluted plasma from rats, demonstrating functionality in biological environments and stability exceeding 9 hours. Our study paves the way for applications of split-aptamers in real-time monitoring of small molecules, with potential implications for in vivo therapeutic drug monitoring and pharmacokinetic studies. [ABSTRACT FROM AUTHOR]

S-Adenosyl- L -methionine (SAM) is crucial for methylation and tightly controlled in cells. We examined SAM-III riboswitch response to 17 SAM analogues and used a Spinach/SAM aptasensor to monitor their enzymatic formation in situ. Most SAM analogues were recognized, unless they featured an ortho-substituted benzyl ring, indicating potential regulatory effects via SAM riboswitches. [ABSTRACT FROM AUTHOR]

Development of rapid and simple detection method holds great significance for the prevention and control of Staphylococcus aureus. Herein, the SELEX method was utilized to screen new aptamers for S. aureus. The double aptamers combined recognition target strategy were adopted to enhance specificity, and magnetic bead target enrichment and the rolling circle amplification strategy were employed to improve sensitivity, thereby achieving a highly specific and highly sensitive visual detection of S. aureus. This method can specifically identify S. aureus and has high detection sensitivity, with a minimum detection limit of 10 CFU/mL. It also shows good repeatability, with a total time consumption of less than 9 h and can be used for the visual detection of actual samples. [ABSTRACT FROM AUTHOR]

Pseudomonas plecoglossicida is a common bacterial pathogen in aquaculture, often leading to visceral white spot disease in large yellow croakers (Pseudosciaena crocea). Previous studies have found that certain aptamers show an efficient antibacterial effect against this pathogen. In this study, we analyzed the transcriptome of P. plecoglossicida to get insights into the antibacterial and inhibitions mechanisms following exposure to the aptamer B4. The results showed seven differentially expressed genes (DEGs) associated with the antibacterial effect of the aptamer, namely sad gene encoding aldehyde dehydrogenase, the paaB gene of phenylacetyl coenzyme A cyclooxygenase, the metN1 gene of ABC transporter proteins, two transposase genes with different positions but identical sequences involved in cutting and splicing DNA sequences, and two hypothetical protein genes with unknown functions. Gene Ontology (GO) analysis showed that the DEGs were mainly involved in DNA-mediated translocation, phenylacetic acid catabolism, growth hormone catabolism, polyamine transporter ATPase activity, betaine aldehyde dehydrogenase activity, ABC transporter protein complex, and other related pathways. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the metabolic pathway of niacin and niacinamide mediated through the sad gene was the most significant and relevant, followed by the metabolism of phenylalanine, alanine, aspartic acid and glutamic acid. Real-time quantitative PCR validation showed that the changes in the DEGs were consistent with the transcriptome analysis. These results suggest that the antibacterial aptamer B4 may inhibit P. plecoglossicida by blocking the synthesis of essential nucleic acids and proteins through the modulation of these DEGs and inhibiting their metabolic pathways. [ABSTRACT FROM AUTHOR]

Background and purpose: Aptamers, a new category of molecular probes, are overthrowing antibodies in molecular diagnostics. However, there are serious problems with using aptamers for this application including poor or non-specific binding in vivo conditions. Systematic evolution of aptamers is achieved through various approaches including CE-SELEX and Cell-SELEX, each suffering its inevitable weaknesses. The shortcomings of negative selection and the lengthy procedure are Cell-SELEX's main problems, while CE-SELEX is deprived of native targets. Here, we introduced a kind of hybrid CE-Cell-SELEX, named CEC hybrid-SELEX, for addressing these limitations in creating aptamer probes detecting human aspartate β-hydroxylase (ASPH), which is a well-established tumor biomarker, in cancer diagnostic investigations. Experimental approach: In our approach, the selected oligomer pool from the last cycle of CE-SELEX was sequenced and then subjected to 3 additional rounds of Cell-SELEX which provides native ASPH (CEC hybrid-SELEX). High-throughput sequencing was applied to achieve a comprehensive sight of the enriched pools. Further confirmatory investigations on oligomers with higher copy numbers were performed using flow cytometry. Findings/Results: Three selected oligomers, AP-CEC 1, AP-CEC 2, and AP-CEC 3, showing Kd values of 43.09 nM, 34.85 nM, and 35.92 nM, respectively, were achieved based on the affinity assessment of the ASPH-expressing cells. Conclusion and implications: Our research suggested that CEC hybrid-SELEX could help recognize which oligomers from CE-SELEX are more capable of binding native ASPH in vivo. [ABSTRACT FROM AUTHOR]

RNA nanoparticles, derived from the packaging RNA three-way junction motif (pRNA-3WJ) of the bacteriophage phi29 DNA packaging motor, have been demonstrated to be thermodynamically and chemically stable, with promise as a nanodelivery system. Background/Objectives: A previous study showed that RNA nanoparticles with antiangiogenic aptamers (anti-vascular endothelial growth factor (VEGF) and anti-angiopoietin-2 (Ang2) aptamers) inhibited cell proliferation via WST-1 assay. To further investigate the antiangiogenic potential of these RNA nanoparticles, a modified three-dimensional (3D) spheroid sprouting assay model of human umbilical vein endothelial cells was utilized in the present study. Methods: Three groups of RNA nanoparticles were evaluated, namely, pRNA-3WJ series, RNA square series (polygon-type RNA nanoparticles), and 8WJ series (multiple-way junction RNA nanoparticles), which were conjugated with a single anti-VEGF, the combination of one anti-VEGF and one anti-Ang2, or multiple anti-VEGF aptamers. The core scaffold RNA nanoparticles (without aptamers) were used as the references, and bevacizumab was used as the positive control. Results: The results demonstrated the inhibition effects of the RNA nanoparticles on endothelial cell tube formation at 67 nM in a 3D spheroid sprouting model. The results in the 3D spheroid sprouting assay are consistent with those of the WST-1 proliferation assays. Conclusions: Among the RNA nanoparticles evaluated, 3WJ-3VEGF and SQR-VEGF-Ang2 had inhibition effects equivalent to bevacizumab and were promising for anti-angiogenesis treatment. [ABSTRACT FROM AUTHOR]

Background/Objectives: Traditional paclitaxel therapy often results in significant side effects due to its non-specific targeting of cancer cells. Peptide aptamer–paclitaxel conjugates present a promising alternative by covalently attaching paclitaxel to a versatile peptide aptamer via a linker. Compared to antibody–paclitaxel conjugates, peptide aptamer–paclitaxel conjugates offer several advantages, including a smaller size, lower immunogenicity, improved tissue penetration, and easier engineering. Methods: This review provides an in-depth analysis of the multifunctional peptide aptamers in these conjugates, emphasizing their structural features, therapeutic efficacy, and challenges in clinical applications. Results: This analysis highlights the potential of peptide aptamer–paclitaxel conjugates as a novel and effective approach for targeted cancer therapy. By harnessing the unique properties of peptide aptamers, these conjugates demonstrate significant promise in improving drug delivery efficiency while reducing the adverse effects associated with traditional paclitaxel therapy. Conclusions: The incorporation of peptide aptamers into paclitaxel conjugates offers a promising pathway for developing more efficient and targeted cancer therapies. However, further research and clinical studies are essential to fully unlock the therapeutic potential of these innovative conjugates and enhance patient outcomes. [ABSTRACT FROM AUTHOR]