Talanta最新文献

The growing demand for glycolate, fueled by economic development, requires the advancement of production methods. Escherichia coli (E. coli), a preferred host for glycolate production, has undergone extensive metabolic engineering to improve yield. Developing rapid and precise methods for quantifying glycolate concentration is essential for screening high-yielding strains. Here, we present the engineering of a novel circularly permuted green fluorescent protein (cpGFP)-based glycolate sensor, termed GLYCO. GLYCO exhibits high specificity (minimal interference from other metabolites), stability (no decrease in performance after 15 days at -80 °C), and ease of detection via fluorescence measurement, enabling effective in vitro glycolate quantification. GLYCO spans a quantification range from 10 μM to 1 mM, facilitating effective monitoring of glycolate production in metabolically engineered E. coli strains. This biosensor represents a significant advancement in the metabolic engineering toolkit, facilitating efficient detection and optimization of glycolate production in E. coli, with potential applications in industrial biotechnology.

Searching for new alternative to tripropylamine (TPrA) with low toxicity and high chemical stability for the tris(4,4'-dicarboxylic acid-2,2'-bipyridyl)ruthenium (II) (Ru(dcbpy)₃²⁺) based coreactant electrochemiluminescence (ECL) system is essential for widespread analytical applications. Here, nitrogen-doped graphene quantum dots (NGQDs) have been discovered to significantly amplify the ECL emission and increase the ECL efficiency of Ru(dcbpy)₃²⁺ for the first time. However, the mechanism by which NGQDs act as coreactants is not well comprehended. Therefore, various optical and electrochemical technologies were employed to investigate the ECL mechanism. It is proposed that the amino and carboxyl groups on the surface of NGQDs play crucial roles as the coreactant active sites, catalyzing the oxidation of Ru(dcbpy)₃²⁺. Based on this foundation, an "on-off-on" ECL aptasensor for the quantification of acetamiprid was developed, exhibiting a broad linear range and a detection limit of 0.056 pM. Satisfactory recoveries, ranging from 98.0 % to 101.6 %, were achieved in pakchoi samples. Consequently, NGQDs could serve as coreactants for Ru(dcbpy)₃²⁺, offering new opportunities for constructing a variety of sensors with extensive analytical applications in the ECL field.

The low sensitivity of Lateral flow assay (LFA) limits its application in rapid detection for trace targets. LFAs with nanozyme (nanozyme-LFA) as signal labels have demonstrated excellent performance in point of care testing (POCT). However, additional operational steps for substrate catalysis in nanozyme LFA are required, which makes the nanozyme-LFA operation complicated. In this work, we designed a LFA based on delayed substrate release (SGF-LFA), in which a commercialized glass fiber membrane embedded with substrate (SGF) was fixed at the sample pad. The SGF could automatically execute substrate delivery and catalysis, thus eventually achieving a one-step LFA operation for the nucleic acid detection of influenza A virus H1N1. In this SGF-LFA, 3,3 '- diaminobenzidine (DAB) was oxidized and deposited, producing a strong signal amplification under the catalysis of Au@PtNP nanozyme. The SGF-LFA could detect the nucleic acid of H1N1, with a linear range of 0.02-50 nM and a limit of detection (LOD) as low as 0.02 nM, which was 25-fold lower than that of the nanozyme-LFA before catalysis. In addition, the analytical performance was close to that of a manual operation mode of catalysis amplification. The application of SGF-LFA for detecting the H1N1 nucleic acid in serum samples obtained a recovery rate of 96 %-102.7 %, indicating that SGF-LFA has great potential in point-of-care testing.

Sensitive and accurate detection and imaging of different microRNAs (miRNAs) in cancer cells hold great promise for early disease diagnosis. Herein, a DNA tetrahedral scaffold (DTS)-corbelled autonomous-motion (AM) molecular machine based fluorescent sensing platform was designed for simultaneous detection of two types of miRNAs (miRNA-21 and miRNA-155) in HeLa cells. Locking-strand-silenced DNAzymes (P:L duplex) were firstly grafted at the loop of target-analogue-embedded double-stem hairpin substrates (TDHS) of DTS, making the sensor in a "signal off" state due to the closely distance between modified fluorophores (FAM and Cy5) with the corresponding quenchers (BHQ1 and BHQ2). The detection of miRNA-21 and miRNA-155 was mainly based on the activation of locking-strand-silenced DNAzymes, cleaving hairpin DNA into single-strand DNA segments, accompanying with the release of modified fluorophores and the signal recovery (signal on). Upon the cyclical stimulation of miRNA targets in such AM molecular machine, sensitive detection of miRNA-21 and miRNA-155 was realized in this self-feedback circuit (SFC) with the detection limit down to 38.8 aM and 27.1 aM, respectively. Moreover, the analytical performance was greatly improved for miRNAs imaging in cancer cells with enhanced tumor cell recognition ability, excellent stability in virtue of DTS, indicating a potential analytical tool in early cancer diseases diagnosis.

Accurately detecting cysteine (Cys) in vivo is crucial for diagnosing Cys-related diseases. A novel ratiometric fluorescent probe featuring dual near-infrared emission is developed in this study for the in vivo ratio imaging of Cys. The probe comprises a hemicyanine organic small-molecule dye (HCy-CYS) with specific Cys recognition capabilities covalently coupled with carbon dots (CDs) synthesized using glutathione (GSH) as the carbon source (GCDs), forming a unique composite nanofluorescent probe (GCDs@CYS). The probe undergoes a specific reaction with acrylate upon the addition of Cys, converting HCy-CYS to HCy-OH. Consequently, the GCD fluorescence intensity at 685 nm gradually decreases, whereas that of HCy-OH at 720 nm progressively increases, yielding a ratiometric fluorescence signal. Notably, both emission wavelengths of the probe exceed 650 nm, thereby effectively mitigating the interference from background signals during cellular and in vivo imaging. Furthermore, the probe demonstrates high specificity for Cys, enabling its differentiation from homocysteine and GSH. The Cys concentration and fluorescence ratiometric intensity exhibit a strong linear correlation at 10-150 μM with a detection limit of 0.95 μM. These results indicate that the ratiometric fluorescent probe can serve as a valuable tool for monitoring Cys-related physiological or pathological processes.

Anthropogenic activities have led to increased stress on our marine and other aquatic environments. There is a pressing need to monitor, measure, understand and mitigate causes of these pressures. This paper presents a novel optical head for monitoring and measuring marine based optical phenomena. The development and preliminary testing of the optical head were designed to detect optically active constituents in the marine and coastal environments. Potential applications may include the detection of Harmful Algal Blooms (HAB), which due to their production of toxins have deleterious effects on marine ecosystems, dissolved organic matter (DOM), oil spills, through the measurement of dissolved fluorescent petroleum compounds and turbidity, a key metric in marine and water quality measurements. Preliminary laboratory based results indicate that the optical head is well suited for measuring in-vivo Chlorophyll a (Chl a) fluorescence, turbidity, fluorescent dissolved organic matter (fDOM) and petroleum. For turbidity and in-vivo Chl a, analytical performance was benchmarked against off-the-shelf commercial sensors. The developed optical head demonstrates good analytical performance with certified reference standards and a very good agreement with the reference instrument.

A novel strategy for cytochrome c selective recognition assisted with cucurbit[6]uril by host-guest interaction via N-terminal epitope imprinting and reversible addition-fragmentation chain transfer (RAFT) polymerization was developed. N-terminal nonapeptide of cytochrome c (GI-9) was used as the epitope template to achieve highly selective recognition of cytochrome c. As a common supramolecule in recent years, cucurbit[6]uril can encapsulate the butyrammonium group of lysine residue to capture the peptide and improve the corresponding spatial orientation by the host-guest interaction for GI-9 or cytochrome c recognition. After cucurbit[6]uril modification and epitope immobilization, the imprinted polymer was synthesized by RAFT polymerization with 2-dodecylsulfanylcarbothioylsulfanyl-2-methylpropanoic acid as chain transfer agent. After template removal, the obtained imprinted particles showed good binding ability to GI-9 (20.28 mg g^-1, IF = 4.11) and cytochrome c (36.12 mg g^-1, IF = 3.91). With the successive addition of cucurbit[6]uril and RAFT agent, the step-by-step improvement of the IF for cytochrome c recognition further illustrated the effects of supramolecular host-guest interaction and regulation of imprinted polymer chain. The imprinted polymers showed obvious advantages for cytochrome c recognition compared to competitive proteins and had good reusability with the repeated reproduction rate 80.8 % after five cycles of adsorption and desorption. Furthermore, the selective recognition for cytochrome c in adult bovine serum proved its potentiality to be applied in practical samples. All these results demonstrated that the combination of epitope imprinting, cucurbit[6]uril host-guest interaction and RAFT strategy presented an efficient new feasible control method for protein recognition with good selectivity, stability and reusability.

Polymers and dendrimers are macromolecules, possessing unique and intriguing characteristics, that are widely applied in self-assembled functional materials, green catalysis, drug delivery and sensing devices. Traditional approaches for the structural characterization of polymers and dendrimers involve DLS, GPC, NMR, IR and TG, which provide their physiochemical features and ensemble information, whereas their unimolecular conformation and dispersion also are key features allowing to understand their transporting profile in confined ionic nanochannels. This work demonstrates the nanopore approach for the determination of charged homopolymers, neutral block copolymer and dendrimers under distinct bias potentials and pH conditions. The nanopore translocation properties reveal that the dispersion and transporting of PEI is pH-dependent, and its capture rate is much lower than that of PAA. The neutral block copolymer with longest molecular chain threads through with longest blockage duration, its highest capture rate was achieved in 0.5 M KCl at pH 5 with slow diffusion and high temporal resolution. The two generations of neutral dendrimers could also translocate under bias potentials, probably due to the ions adsorption on the dendrimers and driven by Brownian force. The TEG-81 with larger molecular size translocates with longer residence time and higher blockage ratio, as expected. Both of the dendrimers exhibit a higher blockage ratio at pH 7.4 than either acidic or alkalic condition, indicating a larger stretched conformation adopted under neutral condition. This work presents the analysis of unimolecular charged and neutral polymers and dendrimers, which will be insightful in understanding the self-assembly motion and transfer of synthetic macromolecules in confined space. It also provides a good indication for deciphering the macromolecule-nanopore interplay under electrophoretic condition.