Background Tuberculosis (TB) nucleic acid diagnosis urgently requires rapid, amplification-free methods to overcome limitations of quantitative real-time polymerase chain reaction (qPCR), including instrument dependency, prolonged time, and contamination risks from nucleic acid amplification. Results Here, an amplification-free electrochemiluminescence (ECL) biosensor based on peptide-templated gold nanoclusters and the clustered regularly interspaced short palindromic repeats-Cas12a system (CRISPR-Cas12a) has been constructed for the detection of Mycobacterium tuberculosis (MTB)-specific IS6110 sequences. Peptide-templated gold nanoclusters with low background and high ECL response can be used as sensitive signal probes. When CRISPR-Cas12a recognizes the target IS6110 DNA, it exhibits non-specific cleavage activity, repeatedly cleaving ferrocene-labelled DNA. This results in the restoration of ECL signals quenched by ferrocene, thereby achieving signal amplification. There is a linear relationship between the signal and the target concentration range from 10 CFU/mL to 104 CFU/mL, with a detection limit of 7 CFU/mL (S/N = 3). Clinical validation (n = 40) showed strong agreement with qPCR (κ = 0.90). Significance Critically, this amplification-free strategy eliminates the need for temperature cycling equipment, reduces detection time to 1 h, and completely avoids the risk of amplicon contamination, providing an ideal solution for rapid screening of highly infectious diseases such as TB.

Rapid on-site monitoring of aflatoxin B1 (AFB1) is critical for food and medicine safety. Herein, a multicolor plasmonic immunoassay was developed for AFB1 detection by combining competitive enzyme-linked immunosorbent assays (ELISA) with enzyme-mediated etching of gold nanobipyramids (AuNBPs). Compared with spherical gold nanoparticles, AuNBPs have sharp tips that provide stronger field enhancement; compared with gold nanorods, the double-conical tips have higher curvature, making them more sensitive to morphological changes during etching. In the competitive system, AFB1 concentration negatively regulates the amount of horseradish peroxidase (HRP) on the solid phase. HRP catalyzes TMB oxidation to trigger an oxidation-reduction etching pathway, which gradually reshapes AuNBPs from bipyramids to spheres, causing a distinct LSPR shift and multicolor change for naked-eye readout. The method shows a limit of detection of 0.67 ng/mL with a linear range of 1-20 ng/mL and performs well in real food samples. This strategy provides a simple, sensitive, and field-applicable tool for on-site AFB1 screening.

Gold nanoclusters (AuNCs) are emerging electrochemiluminescence (ECL) emitters with unique luminescence and high stability. However, conventional AuNC-based ECL systems, which rely on single-signal regulation, often exhibit weak signal responses and limited stability. To address the challenge of detecting extremely low concentrations of the Alzheimer's disease (AD) blood biomarker Aβ40, this study presents a novel ECL-based biosensing platform that leverages dual-signal regulation of AuNCs via glutathione (GSH) and H2O2 serving as ECL quenchers. Based on the peroxidase-like activity of the Aβ40-hemin complex, this system restores the ECL signal by catalyzing the consumption reaction between GSH and H2O2. Using this mechanism, a dual-regulation ECL platform was developed, demonstrating a linear detection range of 0.001-100 ng/mL for Aβ40 with a detection limit of 0.18 pg/mL (S/N = 3). This approach not only offers high sensitivity and specificity but also reduces the complexity of surface modification steps. Overall, this study advances ECL technology through dual regulation and provides a sensitive, practical, efficient, and reliable analytical method for early AD screening.

Excessive reactive oxygen species (ROS) in seminal plasma can trigger male infertility. Therefore, the development of simple and rapid ROS detection methods is urgently needed, particularly for the early self-screening of preconception couples. Herein, a gold nanobipyramid (Au NBP)-based colorimetric hydrogel for convenient and fast ROS detection is described. In the hydrogel, Au NBP is etched efficiently by ROS under the synergistic effect of Fe2+and I-, which finally causes color variations. Besides, agarose gel with the function of molecular sieve enables the separation of biomacromolecules, improving the interference resistance of the system and the stability of Au NBP. This chemical sensor can complete all the tests within 20 min, covering two detection range of 10-125 mu M at relative low H2O2 concentration and 125-1000 mu M at relative high H2O2 concentration, with the detection limits of 1.76 mu M and 12.10 mu M (S/N = 3) respectively. Furthermore, via visual observation of the color variations, it allows the initial interpretation of ROS concentration without any additional equipment. We applied this device to the detection of ROS in clinical seminal plasma samples and obtained promising results, demonstrating its potential for rapid and convenient detection in clinical applications.

The development of high-performance electrochemiluminescence (ECL) systems operating at low triggering potentials remains a critical challenge for advancing their practical applications in bioassays. In this study, a novel ECL system based on a scandium-doped gold nanocluster (Sc@AuNC) and N,N-diisopropylethylamine (DIPEA) had been designed for ultrasensitive detection of urinary Alzheimer's disease-associated neuronal thread protein (AD7c-NTP). By using DIPEA as a low-oxidation-potential coreactant and synergistic doping of Sc to promote free radical generation in the coreactant through accelerated electron transfer, the Sc@AuNC/DIPEA system achieved a 40.89-fold enhancement in ECL intensity and a higher ECL quantum yield (ΦECL) of 35.19% at 0.75 V, surpassing conventional AuNC systems. Utilizing this system, a sandwich-type ECL immunoassay (ECLIA) platform was developed, demonstrating a linear detection range of 0.001 to 100 ng/mL for AD7c-NTP with a detection limit of 0.45 pg/mL (S/N = 3). Owing to the low triggering potential, the ECL system exhibited inherent anti-interference capability by minimizing electrochemical side reactions and preserving biomolecular integrity, as evidenced by stable signals in complex urine matrices. Critically, the satisfactory recovery of clinical urine samples and high correlation with ELISA kits confirmed the utility of the constructed detection platform. This work not only advances ECL technology through coreaction acceleration but also provides a practical, scalable strategy for early Alzheimer's disease screening.

The quantitative and ultrasensitive detection of amyloid-beta peptides Aβ40 in blood is considered as a powerful strategy for early screening of Alzheimer's disease (AD). In this study, combining dual-aptamer recognition and magnetic-induced enrichment, a background-free surface-enhanced Raman scattering (SERS) platform had been developed for efficient Aβ40 detection. Au@Ag-4-ethynylaniline@Au nanoparticles (Au@Ag-4-EA@Au NPs) had been designed and modified on aptamer2 of Aβ40 to form Au@Ag@4-EA@Au-Apt2 (SERS probe). 4-EA functionalization enables the use of the Raman silent region (1800-2800 cm-1), thereby minimizing biological interference and avoiding spectral overlapping. The nanogap within gold and silver nanoshells serves as a strong plasmonic enhancer, significantly amplifying the Raman signal and improving detection sensitivity. Aptamer1 of Aβ40 was modified on a streptavidin-modified magnetic bead (SA-MB) to form SA-MB-Apt1 (capture probe). The presence of the target results in the formation of the sandwich structure of SA-MB-Apt1/Aβ40/Apt2-Au@Ag-4-EA@Au NPs. Two Aβ40-specific aptamers ensured precise biomarker recognition and quantification. Magnetic-induced assembly enriches Aβ40 molecules and generates abundant plasmonic "hot spots" through nanoparticle aggregation, resulting in signal amplification. The results demonstrate a remarkable limit of detection of 25 fM (S/N = 3), with a linear range from 10-1 to 104 pM. This SERS platform provides a robust tool for early AD diagnosis and demonstrates broad potential in clinical molecular diagnostics.

BACKGROUND:Gold nanoclusters (AuNCs) obtained by electroreduction have excellent electrochemiluminescence (ECL) properties, and its ECL intensity is regulated by the valence state. In addition, their ECL signals can be rapidly quenched by reactive oxygen species (ROS). Based on this observation, a sensitive ROS biosensor was designed based on valence regulation of AuNCs. Excessive ROS in seminal plasma can lead to male infertility, and the short half-life and instability of ROS pose a challenge for their detection. Since valence regulation can be done quickly and is very sensitive, this ECL biosensor holds promise to address this issue. RESULTS:The ECL mechanism of AuNCs and the quenching mechanism of AuNCs by ROS were explored, mainly because ROS change the valence state of AuNCs. The ECL signals of the biosensor have a linear relationship with logarithm of the target concentration in the range of 1.0 × 10-8 to 1.0 × 10-3 M and 1.0 × 10-3 to 1.0 × 10-1 M, with a detection limit of 0.75 × 10-10 M (S/N = 3). The biosensor enables rapid one-step detection of ROS and has the advantage of being stable and reusable. More notably, the results of 57 real samples showed that the biosensor can be used to accurately assess the concentration of seminal plasma ROS, guiding the monitoring of sperm quality and the diagnosis of male infertility. SIGNIFICANCE:Compared with the traditional strategy of applying AuNCs only as a luminescent body, this strategy of regulating the valence state of AuNCs to achieve sensitive and rapid detection broadens the application of AuNCs in the field of analysis. Compared with other ROS detection strategies, the one-step immediate detection method effectively avoids the inaccuracy caused by the short half-life and natural dissipation of ROS, and is expected to improve the accuracy and efficiency of clinical diagnosis.

Arginine-based carbon dots (Arg-CDs) were synthesized through a facile one-step hydrothermal method using arginine and urea as precursors. The as-prepared Arg-CDs displayed good aqueous solubility and typical excitation-dependent emission property, with the fluorescence quantum yield of 25.47 %. The fluorescence of Arg-CDs was gradually quenched by the addition of dopamine (DA) in the alkaline solution. However, under neutral and acidic condition, the fluorescence intensity of Arg-CDs remained unchanged upon the addition of DA. The sensing mechanism was probably due to the rapid oxidation of DA to dopaquinone in alkaline condition, which could quench the fluorescence of Arg-CDs through the Inner Filter Effect (IFE). Moreover, the fluorescence change of Arg-CDs was proportional to the concentration of DA under the optimized condition. Because of the linear relationship, the detection limit of dopamine was calculated to be 0.28 mu M with the detection range from 10 to 100 mu M. The interference experiment was carried out to further verify the specificity of this fluorometric method. Finally, the fluorometry based on Arg-CDs has been successfully used to the determination of DA in human serum with high accuracy and satisfactory recovery.

Surface-enhanced Raman scattering (SERS) has great potential in biological analysis due to its specificity, sensitivity, and non-invasive nature. However, effectively extracting Raman information and avoiding spectral overlapping from biological background interference remain major challenges. In this study, we developed a background-free SERS nanosensor consisting of gold nanobipyramids (Au NBPs) core-Prussian blue (PB) shell (Au NBPs@PB), for endogenous H2S detection. The PB shell degraded quickly upon contact with endogenous H2S, generating a unique Raman signal response in the Raman silent region (1800-2800 cm(-1)). By taking advantage of the high SERS-activity of Au NBPs and H2S-triggered spectral changes of PB, these SERS nanosensors effectively minimize potential biological interferences. The nanosensor exhibits a detection range of 2.0 mu M to 250 mu M and a limit of detection (LOD) of 0.34 mu M, with good reproducibility and minimal interference. We successfully applied this background-free SERS platform to monitor endogenous H2S concentrations in human serum samples with satisfied results.

The detection of specific DNA sequences and the identification of single nucleotide polymorphisms are important for disease diagnosis. Herein, by combining the high specificity of the base-stacking effect with the high reproducibility of bovine serum albumin (BSA) modified electrodes and the high loading performance of DNA nanoclews (DNA NCs), a novel sandwich-type electrochemiluminescence (ECL) biosensor is reported for the highly specific detection of HPV16 (chosen as the model target). The capture probes are loaded by BSA carrier platforms modified on the gold electrode surface to improve reproducibility. DNA NCs loaded with a large amount of Ru(phen)32+ worked as signal probes. The template probe is composed of the complementary strand of the target and two free nucleic acid anchors at the head and tail. In the presence of the target DNA, the template probes can form stacked base pairs with target, generating high base-stacking energy. This results in the shorter free anchors of template probes being able to bind to the capture and signal probes. This eventually forms a sandwich structure that allows Ru(phen)32+ to be near the electrode surface, producing an ECL signal. There is a linear relationship between the signal and the target concentration range from 10 fM to 100 pM, with a detection limit of 5.03 fM (S/N=3). Moreover, the base-stacking effect has single base recognition ability for base pairs, effectively avoiding false positive signals. The results of this strategy for clinical samples are consistent with classical methods.

16S ribosomal-RNA (16S rRNA) is often used as an ultrasensitive marker for Chlamydia trachomatis (CT) detection because of its species specificity and high copy number in CT. Robust methods for 16S rRNA detection must be developed to realize the early diagnosis of CT infections. In this work, a highly reproducible and sensitive electrochemical biosensor based on duplex-specific nuclease (DSN)-assisted target-responsive DNA hydrogels and bovine serum albumin (BSA) carrier platform for CT detection was developed. Target rRNA can trigger the DNA hydrogel response, which causes it to be repeatedly cleaved by DSN, ultimately leading to the release of a large amount of horseradish peroxidase-labelled streptavidin (SA-HRP) embedded in the hydrogel beforehand. The released SA-HRP was stably captured by the capture probes that were orderly loaded at the gold electrode with the help of a BSA layer. Then, SA-HRP catalyzed the redox reaction of 3,3',5,5'-tetramethylbenzidine and H2O2, producing a current signal that can be detected. The current signal was proportional to the concentration of CT 16S rRNA from 10 fM to 25 pM with a detection limit of 5.8 fM (S/N = 3). The signal conversion function of the DNA hydrogel avoids the instability of nonhomogeneous nucleic acid hybridization on the gold electrode surface, and combined with optimization by BSA for capture probe modification, this electrochemical biosensor is highly reproducible with a relative standard deviation of 4.3% for the detection of 10 samples of the same concentration. The proposed strategy provides a highly reproducible and sensitive detection method for the extensive screening of CT. (C) 2022 Elsevier B.V. All rights reserved.

The lack of large-scale human papillomavirus (HPV) DNA screening is a major contributor to the high incidence and mortality of cervical cancer in economically undeveloped areas. The development of sensitive, rapid, and low-cost screening techniques is urgently needed. Here, DNA nanoflowers encapsulating glucose oxidase and horseradish peroxidase (GHDFs) were synthesized by one-pot rolling circle amplification, and then the GHDFs were used as the cargo of DNA hydrogel and applied for HPV DNA detection. When target DNA was present, the DNA hydrogel cross-linking structure was disrupted, releasing GHDFs, which then catalysed the oxidation of glucose and tetramethylbenzidine in a cascade, generating a significant electrical signal. Signal intensity had a linear relationship with the logarithm of target DNA concentration in the range of 10 fM-1 nM with a detection limit of 3.76 fM. The detection time of the proposed biosensor was 25 min, which was suitable for large-scale HPV DNA screening in economically underdeveloped areas and provides a blueprint for the detection of other DNA of interest.

The application of surface-enhanced Raman scattering (SERS) in aqueous sample detection is normally limited by the low affinity between analytes and SERS-active nanoparticles. Furthermore, a tendency of uncontrollable aggregation of solute (nanoparticles or analytes) can cause poor reproducibility of the detected SERS signal. Herein, a ready-to-use plasmonic gel bead was developed for rapid and effective detection of aqueous samples with high sensitivity and reproducibility. The SERS gel bead is made of calcium alginate gel beads (CAGBs) that contain gold nanobipyramids (Au NBPs) and an aqueous sample, which can be prepared and detected within only 1 min. Au NBPs/CAGBs can generate an in-depth three-dimensional SERS-active gel network for trapping analytes and offering a uniform hotspot region, which produces a reproducible and uniform signal. Using rhodamine 6G as a model target, the proposed method exhibits excellent reproducibility with a relative standard deviation of 6.57% and a detection limit of 0.4 nM. Then, Au NBPs/CAGBs were applied to quantitatively detect serum uric acid in the range of 10-1000 mu M and a limit of detection of 0.18 mu M, and the results were strongly consistent with those of the commercial ELISA method. This work offers a low-cost and easy route for the fabrication of a versatile SERS substrate for monitoring disease-related biomarkers and point-of-care testing.

Mostly, surface-enhanced Raman scattering (SERS) sensors used the Raman characteristic bands concentrated in the Raman "fingerprint" region (500-1800 cm-1), which may result in spectral overlapping interference. The study of the response in the Raman-silent region (10-500 and 1800-2800 cm-1) can help overcome this problem. Hydrogen sulfide (H2S) gas causes a great threat to human's health, but its low concentration in the airborne species is a challenge for sensitive and selective detection. Herein, a novel low-wavenumber (10-500 cm-1) SERS sensor for H2S gas detection has been developed based on gold nano-bipyramids (Au NBPs) encapsulated by zeolitic imidazolate framework-8 (ZIF-8) (Au NBPs@ZIF-8). The sensor takes advantage of the high adsorption capacity of ZIF-8 toward H2S gas and the H2S-triggered SERS spectral changes in the low-wavenumber Raman-silent region. A clear SERS peak of Au-Br at ∼175 cm-1 generated from Au NBPs@ZIF-8 showed a decrease in the presence of H2S because of the competition of adsorption sites between Au-S and Au-Br bonds. Furthermore, Au NBPs@ZIF-8 can enrich and monitor the level of H2S gas with high efficiency and low interference. The developed sensor has a detection range of 0.2 nM to 20 mM with a limit of detection (LOD) of 0.17 nM. The developed sensor had been applied to detect the H2S gas released from the spoiled fish meat with high selectivity.

In this work, core-satellite assemblies and exonuclease assisted double amplification strategy is developed to produce surface-enhanced Raman scattering (SERS) biosensor towards ultrasensitive detection of biotoxin. In the presence of target molecules, the exonuclease III (Exo III) assisted efficient recycling amplification provides an excellent pathway for the fabrication of core-satellite SERS sensor. Briefly, the proposed strategy includes the following double amplifications: (i) Exo III induced target-related signal amplification; (ii) core-satellite assemblies assisted formation of SERS "hot-spots" induced signal amplification. To show the applicability of the suggested strategy, the detection of ochratoxin A (OTA), one of the most toxic and widely distributed biotoxin, is demonstrated as an example. The results show that the limit of detection (LOD) of OTA is 0.83 fg mL(-1) (S/N = 3). On the basis of the DNA aptamer induced specific target recognition, hence our sensing strategy is easy to be expended to the ultrasensitive detection of other targets, e.g., DNAs, RNAs, and other molecules that have corresponding DNA aptamers. (C) 2020 Elsevier B.V. All rights reserved.

A surface-enhanced Raman scattering (SERS) substrate with good flexibility and high water absorbing capacity is reported. It consists of a calcium alginate sponge incorporating gold nanoparticles. These are in close contact with the sponge without the need for amino or sulfhydryl modification. The substrate is capable of detecting the dyes crystal violet (CV) and malachite green (MG) in water directly and rapidly by immersing it into the liquid sample. Preconcentration and separation are not required. The dyes absorbed on the sponge can be detected without drying and thus the whole analytical process can be completed within 3 min. The results show that the lowest detectable concentrations are 0.1 and 0.25 μg⋅L−1 for CV and MG, respectively. This is lower than the minimum required performance limits set by the European Commission and the US EPA. Moreover, MG and CV can be simultaneously detected in liquid samples due to their different SERS bands (at 1216 and 1534 cm−1, respectively). It should be noted that the molecular structures of MG and CV are very similar. Therefore, the method has a large potential for determination of several analytes simultaneously even in complex sample metrics.

A rapid method is described for the preparation of a highly uniform and sensitive SERS substrate by an improved ‘drop-and-dry’ method. Gold nanobipyramids (Au NBPs) were prepared inside the nanoholes (nanowalls) of anodic aluminum oxide (AAO) templates with a typically 5-μm nanohole depth. The SERS substrate can be prepared by this method within 40 s and on large scale. The SERS signals obtained with this Au NBPs-AAO substrate is stronger by four-orders of magnitude compared to conventional a silicon wafer substrate. The SERS signal for dopamine (DA; measured at 1311 cm−1) is found to be enhanced by a factor of 2.2 × 108. The response to DA extends from 10 nM to 0.1 mM, and the limit of detection is 6.5 nM (at S/N = 3). The assay was applied to the determination of DA in spiked human serum.

Nicotine is highly addictive and harmful. It is one of the main active ingredients in tobacco and a major pollutant in environmental tobacco smoke. Thus, it is important to detect the nicotine content in tobacco and to monitor the nicotine content in environmental tobacco smoke. However, until present, there still has been no effective device for on-site determination of nicotine content in tobacco and environmental tobacco smoke. In this work, a portable device is fabricated for sensitive on-site evaluation of nicotine in tobacco and environmental tobacco smoke based on surface-enhanced Raman scattering (SERS). The weight of the entire device is less than 1 kg, and it uses a chargeable battery to drive both the pump and the Raman spectrometer. The total analysis time can be completed within 3-5 min. Thus, it has great potential for on-site analysis of nicotine in tobacco and environmental tobacco smoke.

In this review, we focused on the mechanisms of ordered nanoparticle assemblies and the strategies and applications of ordered assemblies for surface-enhanced spectroscopy.

The efficient extraction of targets from complex surfaces is vital for technological applications ranging from environmental pollutant monitoring to analysis of explosive traces and pesticide residues. In our present study, we proposed a proof-of-concept surface enhance Raman scattering (SERS) active substrate serving directly to the rapid extraction and detection of target molecules. The novel substrate was constructed by decorating the commercial tape with colloidal gold nanoparticles (Au NPs), which simultaneously provides SERS activity and "sticky" of adhesive. The utility of SERS tape was demonstrated by directly extracting pesticide residues in fruits and vegetables via a simple and viable "paste and peel off" approach. The obtained strong and easily distinguishable SERS signals allow us to detect various pesticide residues such as parathion-methyl, thiram, and chlorpyrifos in the real samples with complex surfaces including green vegetable, cucumber, orange, and apple.