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[en] Highlights: • Accurate recovery of anisotropy from overlapping fluorescence emission. • Uses Multivariate Curve Resolution and trilinear constraints. • A simple and robust sample system for validating chemometric analysis of polarization data. - Abstract: Anisotropy resolved multidimensional emission spectroscopy (ARMES) provides valuable insights into multi-fluorophore systems like proteins that have complex overlapping emission bands. The method combines multidimensional fluorescence, anisotropy, and chemometrics to facilitate the differentiation of fluorophores with very similar emission properties. Here, we address the critical issue of standardizing the chemometric methods required to accurately extract spectral and anisotropy information from fluorophore mixtures using two standard sample sets: perylene in glycerol, and a mixture of Erythrosin B and Phloxine B with overlapping emission but different anisotropies. We show for the first time how to accurately model component anisotropy using Multivariate Curve Resolution (MCR) from data collected using total synchronous fluorescence scan (TSFS) and Excitation Emission Matrix (EEM) measurement methods. These datasets were selected to avoid the presence of inner filter effects (IFE) or Förster resonance energy transfer (FRET) that would depolarize fluorescence emission or reduce data tri-linearity. This allowed the non-trilinear TSFS data to yield accurate component anisotropy data once modelled using the correct data augmentation strategy, however, the EEM data proved to be more accurate once optimal constraints (non-negativity and correspondence among species) were employed. For perylene (S2) and Phloxine B which both have very weak anisotropy (1) and Erythrosin B which have large (>0.2) anisotropies, bilinear and trilinear EEM models built using a total tri-linearity constraint, yielded solutions without any rotational ambiguities and very accurate (±4% of real value) anisotropy values. These sample systems thus provide simple and robust test systems for validating the spectral measurement and chemometric data analysis elements of ARMES.
[en] Highlights: • A sensitive ECL immunosensor was constructed with the amplification of pL-Cys toward self-enhanced PEI-Ru(II). • The intramolecular electron transfer could efficiently amplify ECL signal owing to its benefit for electron transmission. • PL-Cys as a strongly reducing radical could react with PEI-Ru(III) to generate PEI-Ru(II)* with further enhanced ECL signal. • The prepared ECL immunosensor exhibited great advantages in simplification, rapidity, stability and selectivity. - Abstract: In this work, the luminophor tris (4,4′-dicarboxylicacid-2,2′-bipyridyl) ruthenium (II) dichloride (Ru(dcbpy)32+) was linked to coreactant polyethylenimine (PEI) with effectively shorted distance, forming a novel self-enhanced PEI-Ru(II) complex to fabricate ultrasensitive electrochemiluminescence immunosensor for carcinoembryonic antigen (CEA) detection by using polyamino acid l-cysteine (pL-Cys) as booster for further signal amplification. The light-emitting species (PEI-Ru(II)*) based on intramolecular electron transfer between PEI• and Ru(III) could efficiently amplify the ECL signal of Ru(dcbpy)32+ owing to its benefit for electron transmission. Secondly, the pL-Cys film was fabricated on the glassy carbon electrode by cyclic voltammogram. After that, the pL-Cys acted as a strongly reducing radical could react with PEI-Ru(III) to generate the excited state PEI-Ru(II)*, which further amplify the ECL signal. With the signal amplification factors, the prepared ECL immunosensor exhibited great advantages in simplification, rapidity, stability and selectivity. Furthermore, the linear range for the determination of CEA was from 0.10 pg/mL to 80 ng/mL with a correlation coefficient of 0.9942 and a detection limit of 0.045 pg/mL (S/N = 3).
[en] Highlights: • The mirror-image aptamer kissing complex (AKC) approach was demonstrated. • The scope of the AKC strategy was extended to peptide targets. • The all-L AKC improved the fluorescence anisotropy assay robustness for complex matrix analysis. • The use of methanol as cosolvent enhanced the assay sensitivity. - Abstract: The recently reported aptamer kissing complex (AKC) strategy has allowed for the development of a new kind of sandwich-like sensing tools. Currently AKC assays have been only applied to low molecular weight molecules and their functionality in complex matrices remains challenging. The objective of the present study broken down into two sub-aims; exploring the propensity to broaden the scope of detectable analytes and designing a more robust system for potential applications to realistic samples. An all L-configuration aptaswitch module derived from a hairpin spiegelmer specific to a larger target, i.e. the arginine-vasopressin (AVP) hormone, was elaborated. The target-induced AKC formation in presence of a specific mirror-image RNA hairpin (L-aptakiss) probe were analyzed by using fluorescence anisotropy. The mirror-image kissing complex was successfully formed when the L-AVP target bound to the engineered L-aptaswitch element. It was also established that the use of methanol as cosolvent significantly improved the assay sensitivity through the stabilization of the ternary complex. Finally, the capability of the mirror-image method to operate in 10-fold diluted, untreated human serum was illustrated. The current work revealed that the AKC concept can be expanded to a wider range of targets and converted to a L-configuration sensing platform especially suitable for bioanalysis purposes.
[en] Highlights: • A desorption APCI method has been optimized for the analysis of nonvolatile and volatile compounds. • Arcing, spark discharge and the erosion of the needle tip can be avoided using resistor in positive ion mode. • DC corona discharge is better than AC corona discharge for dry sample desorption and ionization. • Miniaturized sample size (0.5 µL) is important in many filed of applications. • Biological samples can be analyzed with minimum sample pretreatment. - Abstract: In this work, a simple desorption atmospheric pressure chemical ionization (SDAPCI) source is studied and optimized for analyzing a wide variety of samples such as nonvolatile, volatile and biological samples. In this ion source, the heated mass inlet was used for sample desorption, and a solid needle was used to produce a corona discharge for ionization. The utilization of any additional gas or heater is not required in SDAPCI. Due to its high sensitivity, only a small amount of sample is needed. Sample loading and the consequent mass spectrometry analysis process could be easy and fast, which was demonstrated by the analyses of different types of samples ranging from non-volatile to volatile compounds. High-throughput analysis can be performed by SDAPCI source with minimum or no sample preparation.
[en] Highlights: • Regulating MOCPs through pre-coordination with Cu and Au ions. • MOCPsCu+Au one-pot entrapped enzyme with ratio being close to 100%. • MOCPsCu+Au modified electro-biosensor presented superior enzymatic catalysis performance. - Abstract: We propose a method for regulating biomolecules immobilization performance of metal-organic coordination polymers (MOCPs) through pre-coordination for highly sensitive biosensing. 2,5-dimercapto-1,3,4-thiadiazole (DMcT) was used as organic monomers. Firstly, using CuCl2 as the source of metal ions to form short oligomers with DMcT (MOCPsCu), which can regulate the length of ligands through pre-coordination. Then exploiting NaAuCl4 as the source of Au ions to coordinate both short oligomers and biomolecules (MOCPsCu+Au), since Au ions can coordinate with both N and S atoms. Through controlling the concentration of CuCl2, oligomers with desired length could be readily obtained to prepare MOCPsCu+Au framework with controllable porosity and enzyme entrapment efficiency. Thus MOCPsCu+Au offers several advantages including improved mass transfer efficiency and biocatalytic sensitivity than conventional MOCPs using single metal ions. Glucose oxidase (GOx) was used as the representative biomolecule, the entrapment ratio of enzyme in MOCPsCu+Au case reached an extreme value of 100%. These MOCPsCu+Au biocomposites modified electrode also showed greatly enhanced biocatalytic sensitivity (127 μA cm−2 mM−1) and very low detection limit (58 nM), compared with those reported analogues. The new materials/strategy may create new avenue to regulate the performance of ligand-constructed polymers and their composites for entrapment-based applications.
[en] Highlights: • Nickel-mediated allostery of G-quadruplex is reported for the first time. • Activity of G-quadruplex DNAzyme is manipulated by nickel-mediated allostery and nickel-histidine affinity pair. • Histidine in biological fluids can be facilely and directly distinguished in the absence of additional reagents. - Abstract: Since abnormal metabolism of histidine (His) is defined as an indicator of several diseases, detection of His in biological fluids becomes increasingly urgent to us. However, due to similar structures and properties of different amino acids, selective quantification of His is difficulty, and typically needs the participation of special reagents. In this work, we report for the first time that nickel ions (Ni2+) can induce the allostery of G-quadruplex, and is thus able to manipulate the activity of G-quadruplex DNAzyme. Experimental results indicate the interaction between Ni2+ and guanine is critical to the allostery. In comparison with Ni2+-guanine interaction, Ni2+-His interaction exhibits higher affinity. Therefore, a colorimetric His biosensor is fabricated, and His can be facilely discriminated by naked eyes. Relying on the high activity of DNAzyme, His in a range of 50 nM–400 μM is determined with this method, and low detection limit (36 nM) is obtained. More importantly, His can be directly distinguished in the absence of other toxic reagents. In addition, the amount of His in serum is also measured, suggesting the applicability of this biosensor in real sample detection. Overall, this work provides an alternative way to design G-quadruplex DNAzyme-based analytical approaches.
[en] Highlights: • Formulation of DNA chimera-templated silver nanoclusters (AgNCs) was studied. • They made of ssDNA with nucleating-sequence (NC) and aptamer at different position. • Because of their location, same NC and aptamer can give turn-on or turn-off signal. • The formulation also affects selectivity and sensitivity of aptamer. - Abstract: Single strand DNA (ssDNA) chimeras consisting of a silver nanoclusters-nucleating sequence (NC) and an aptamer are widely employed to synthesize functional silver nanoclusters (AgNCs) for sensing purpose. Despite its simplicity, this chimeric-templated AgNCs often leads to undesirable turn-off effect, which may suffer from false positive signals caused by interference. In our effort to elucidate how the relative position of NC and aptamer affects the fluorescence behavior and sensing performance, we systematically formulated these NC and aptamer regions at different position in a DNA chimera. Using adenosine aptamer as a model, we tested the adenosine-induced optical response of each design. We also investigated the effect of linker region connecting NC and aptamer, as well as different NC sequence on the sensing performance. We concluded that locating NC sequence at 5′-end exhibited the best response, with immediate fluorescence enhancement observed over a wide linear range (1–2500 μM). Our experimental findings help to explain the emission behavior and sensing performance of chimeric conjugates of AgNCs, providing an important means to formulate a better aptasensor.
[en] Highlights: • Graphene grown on stainless steel mesh was synthesized and used as sorbent. • The sorptive device was applied for the extraction of PAHs. • The method is a fast, simple, and sensitive sample preparation technique. • The proposed method achieved low LODs, wide linearity, and good precision. - Abstract: In this study, graphene grown on stainless steel mesh efficiently incorporates the advanced properties of graphene in the form of an ultra-thin coating with the open geometry of the substrate, resulting in a highly sensitive and fast sorptive device capable of extracting target analytes directly from sample matrices. The synthesis of graphene on the stainless steel mesh was confirmed by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The advantages of the new microextraction device have been investigated for the determination of polycyclic aromatic hydrocarbons (PAHs) in environmental water samples in combination with gas chromatography–flame ionization detector (GC–FID). The factors affecting the extraction efficiency such as stirring speed, extraction time, ionic strength and desorption conditions, were investigated and the optimal extraction conditions were established. Under the optimal extraction condition, linearity was in the range of 0.003–100 ng mL−1 with a correlation coefficient of 0.9923–0.9985. The limits of detection (LODs) of the developed method are obtained between 1 and 3 pg mL−1. The repeatability was assessed at two concentration levels (0.01 and 10 ng mL−1) and the relative standard deviations (RSDs) obtained were 4.9–8.7%. The preparation reproducibility of sorbent device, evaluated with the RSDs, was in the range of 7.3–10.2% (n = 5) in one batch, and 8.3–11.0% (n = 5) among different batches. Finally, the proposed method is applied for the determination of the target PAHs in the real water samples.
[en] Highlights: • Ambient ionization platform provided direct rapid fingerprinting of solid lignin. • Spray-based ionization methods generated ions with wider m/z range than DART. • Robust data visualization tools permitted direct inspection of complex lignin data sets. • Modified Kendrick mass defect filtering enabled lignin subunit separation. - Abstract: Ambient ionization techniques are typically used to analyze samples in their native states with minimal or no sample pretreatment prior to mass spectrometric (MS) analysis. Desorption electrospray ionization (DESI) and direct analysis in real time (DART) were systematically investigated in this work for direct solid analysis of depolymerized lignin samples, with the goal of rapidly fingerprinting these samples, to efficiently characterize the subunits of this renewable energy source. High resolution MS was required for enhanced selectivity in this study due to the inherent structural complexity of lignin. DESI provided results across a broader mass range (up to m/z 700) than DART and also ionized saturated compounds of low oxygen-to-carbon (O/C) ratios and low double bond equivalents (DBE). While DART detected the same core lignin monomeric and dimeric compounds as seen with DESI and electrospray ionization (ESI), results were restricted to a narrower mass range to m/z 500, due to thermal degradation and losses of methoxy groups. In contrast to DESI and ESI, the DART spectra were nearly void of saturated components. On a structural level, the core lignin compounds were visually fingerprinted and ionization method performances critically assessed by employing simplified Kendrick-based data mining approaches. A novel simplified data visualization approach was developed in this work based on modified Kendrick mass defect (KMD) filtering for lignin subunits and plotting the mass defect values against the degree of unsaturation. Direct visualization of monomeric, dimeric and trimeric lignin species was simplified by the KMD separation plots, easily allowing the core lignin compounds to be visually identified and compared. Modified KMD bases, namely methoxy and phenol bases, which represent monomer-specific lignin constituents, were successfully used to classify and group the complex mixture of lignin species. Further separation of methoxy-related lignin species was successfully achieved by employing the more specific phenolic KMD base.
[en] Highlights: • In-situ generation of CdS on TiO2 NRAs with high photo-to-current conversion efficiency was used. • Fe3O4@PDA conjugate was used as the label carrier to immobilize Ab2 and HRP. • Fe3O4 NPs and HRP can synergistically accelerate the oxidation of 4-CN to produce BCP. • An ultrasensitive PEC immunosensor was constructed for MC-LR detection. - Abstract: An ultrasensitive photoelectrochemical (PEC) immunoassay based on multiple signal amplification strategy was fabricated for the detection of microcystin-LR (MC-LR). The CdS/TiO2 nanorod arrays (CdS/TiO2 NRAs) modified FTO electrode, which can weaken the self-oxidation by photogenerated holes of CdS nanoparticles, and limit the recombination of electron-hole pairs and broaden optical absorption of TiO2 NRAs, was used as a visible-light driven material to immobilize antigens. Then, Fe3O4 nanoparticles/polydopamine (Fe3O4@PDA) was used as the carrier to load secondary antibody (Ab2) and horseradish peroxidase (HRP), where Fe3O4 nanoparticles and HRP can synergistically accelerate the oxidation of 4-chloro-1-naphthol (4-CN) by H2O2 to produce biocatalytic precipitation (BCP) on the surface of modified electrode. Due to the catalytic activity of Fe3O4 nanoparticles and HRP, the nonproductive absorption of HRP and the steric hindrance by BCP, the photocurrent change was amplified. The proposed PEC immunosensor can detect MC-LR in a range of 0.005–500 μg/L with a detection limit of 0.001 μg/L. Meanwhile, the PEC immunosensor exhibited high sensitivity, good stability, acceptable selectivity and reproducibility, indicating its potential application in environmental monitoring.