Search

Conventional ELISA methods have been widely used for disease diagnosis, yet the sensitivity and accuracy are still to be improved. Based on our label-free AuNPs-mediated plasmonic sensor, we developed an ultrasensitive plasmonic ELISA by integrating the plasmonic sensor and cascade amplification. In principle, alkaline phosphatase (ALP)-conjugated antibodies captured targets, and the ALP hydrolyzed 2-phospho-ascorbic acid (AA2P) into ascorbic acid (AA). Then, the released AA changed Fe3+ into Fe2+, which further triggered Fe2+-catalyzed H 2 O 2 -preventing aggregation of AuNPs by oxidizing Cysteine (FeHOAuC). Finally, the target content was negatively related with the AuNPs aggregation and the subsequent color variation. As a proof-of-concept, this novel plasmonic ELISA exhibited an ultralow LOD (0.1 ng/mL, around 4.4 pM) and a detection range (0.25–8.0 ng/mL) on detecting neutrophil gelatinase-associated lipocalin (NGAL) within 120 min. The sensitivity was superior to those of conventional ELISA and plasmonic ELISA. In summary, our study provided a novel and sensitive plasmonic ELISA for proteinic biomarker diagnosis. [Display omitted] • A label-free plasmonic ELISA was fabricated by cysteine-induced AuNPs clustering. • The detection was triply amplified by enzymatic catalysis and our nanosensor platform. • The plasmonic ELISA specifically sensed proteins with an ultralow LOD (4.4 pM). • The plasmonic ELISA was feasible to detect NGAL for serum sample of disease mouse. [ABSTRACT FROM AUTHOR]

Label-free gold nanoparticles (AuNPs) plasmonic sensors are limited in the biological assay for weak expansibility. Here, a novel H 2 O 2 -sensing AuNPs plasmonic sensor was proposed using 3, 3′, 5, 5′-tetramethylbenzidine (TMB) redox catalysis of H 2 O 2 -indcued cysteine oxidation for AuNPs coloration (TreHOAuC), and the application of TreHOAuC was examined by combining H 2 O 2 -generating oxidase. In the TreHOAuC platform, TMB is transformed into oxidized TMB (oxTMB) by horseradish peroxidase in the presence of H 2 O 2 , and oxTMB is restored into TMB after oxidizing cysteine into cystine. The redox cycling of TMB boosts H 2 O 2 -induced cysteine oxidation, and the cysteine oxidation is visualized by AuNPs. Critical parameters of the TreHOAuC platform were optimized such as concentration, time, pH, and buffer. Using the TreHOAuC system, H 2 O 2 can be specifically sensed by naked eyes within 2 min, and the detection limit was less than 0.5 μM. As a proof-in-conception, a cholesterol oxidase-paired TreHOAuC system was used to detect cholesterol in serum samples, indicating a linear range of 2–40 μM and great preciseness. In summary, our proposed TreHOAuC method provided a novel and more comprehensive application for biological sensing. [Display omitted] • A label-free plasmonic sensor was fabricated by cysteine-induced AuNPs clustering. • H 2 O 2 initiated cysteine oxidation was catalyzed by HRP-assisted TMB redox cycle. • TreHOAuC system rapidly and specifically senses H 2 O 2 within 2 min (LOD: 0.5 μM). • TreHOAuC system assays more molecules by coupling with H 2 O 2 -generating oxidases. [ABSTRACT FROM AUTHOR]

Highlights • UCNPs-MoS 2 nanoassemblies were developed to detect reactive oxygen species (ROS) with a fast response time. • The nanoprobe had been successfully applied in the imaging of exogenous and endogenous ROS in living cells. • The nanoprobe was employed in in vivo visualization of ROS level in zebrafish. Abstract Nanoassemblies for bioimaging in living cells have attracted much attention. In this work, an efficient and sensitive turn-on fluorescent detection platform for reactive oxygen species (ROS) in living cells has been constructed using the nanoassemblies of upconversion nanoparticles (UCNPs)-MoS 2 nanoflakes. In this nanostructure, the positively charged UCNPs could assemble with negatively charged poly(acrylic acid) modified MoS 2 nanoflakes by electrostatic force, which led to the remarkable fluorescence quenching of UCNPs owing to the excellent quenching ability of MoS 2 nanoflakes. In contrast, in the presence of ROS, the fluorescence recovery of UCNPs is successfully realized owing to the decomposition of MoS 2 nanoflakes through the chemical reaction with ROS. The ROS-mediated disassembly of the nanoarchitecture display excellent fluorescent response toward ROS. Furthermore, the disassembly of UCNPs-MoS 2 nanoflakes has been successfully used for ROS bioimaging in cells and zebrafish. [ABSTRACT FROM AUTHOR]

• An ultrasensitive strategy is proposed for tumor-specific exosomal protein assay. • A single MB is manipulated as the reaction carrier for the enrichment of fluorescence signals. • TdT-initated DNA extension on the MB is applied for signal amplification. • AuNPs is used as the carrier loaded with DNA aptamers to further amplify the detection signals. • The proposed method is applied for direct detection of Exo-EpCAM from plasma microsamples. Tumor-derived exosomes carry lots of tumor-specific proteins, attracting more attention as ideal biomarkers for non-invasive diagnosis. However, due to the low concentration of exosomes in biological samples, the development of an ultrasensitive method for tumor-specific exosomal protein detection is highly desired. Here, a sensitive Single Microbead (MB)-based Aptasensor coupled with Terminal deoxynucleotidyl transferase (TdT)-triggered signal amplification (SMAT) is reported to detect tumor-specific exosomal proteins. In the SMAT detection system, a single MB is used as the reaction carrier and firstly modified by the antibodies of generic membrane biomarkers (anti-CD63 and anti-CD81) of exosomes for efficient enrichment of exosomes in samples. Then, the DNA aptamers immobilized on gold nanoparticles (AuNPs) are applied to specifically bind to the tumor-specific proteins on exosome surfaces. Subsequently, the DNA aptamers can be efficiently extended by TdT to generate a long poly(dT) tail. By hybridization with lots of fluorescein-labeled poly(dA) 25 (FAM-poly(dA) 25) probes, large amounts of fluorescence molecules will be highly enriched on one MB and in situ monitored with fluorescence imaging. With this strategy, exosomal EpCAM protein (Exo-EpCAM) has been sensitively detected from the samples containing 55 particles/μL tumor-derived exosomes. Furthermore, the proposed method could be applied for detection of Exo-EpCAM from plasma microsamples, indicating great application prospect in future for clinical diagnosis. [ABSTRACT FROM AUTHOR]

• The high-quality hydrophobic and hydrophily red-QD and green-QD were synthesized for dQDs-FLISA. • Cross-reactivity among Ag-Ab can be avoided by the specific probe adding sequence. • The simultaneous and quantitative detection was developed in one sample-well. • The assay has a wide analytical range, a low LOD, and good recovery rates. Simultaneous detections of different but clinically relevant biomarkers are of extremely importance in biomedicine. Due to their unique photophysical properties, quantum dots (QDs) are ideally suited for highly sensitive multiplexed determination. Herein, a dual quantum dots-based fluorescence-linked immunosorbent assay (dQDs-FLISA) for simultaneous and quantitative detection of inflammation biomarkers (i.e. serum amyloid A (SAA) and C-reactive protein (CRP)) has been established. After being coated by amphiphilic oligomers (polymaleic acid n-hexadecanol ester, PMAH), the red-QD and green-QD with high quantum yields were used as fluorescence probes to couple SAA and CRP antibodies, respectively. Cross-reactivity among the SAA and CRP's antibodies and antigens have been carefully examined by interference experiments, and it was successfully avoided by the specific probe adding sequence. Therefore, the assay provided a broad linear analytical range, including SAA quantitative range of 10–1,000 ng mL−1 with linear correlation (R2) of 0.992, and CRP quantitative range of 10–1,000 ng mL−1 with R2 of 0.998. The limit of detections (LODs) for SAA and CRP using dQDs-FLISA were 2.39 ng mL−1 and 6.37 ng mL−1, respectively. The accuracy of the assay has been confirmed with recoveries of 92.13%–101.85%. More importantly, the assay results showed good specificity, the QD-antibody probe could couple corresponding antigen (CRP or SAA) high-efficiently and it could be out of interference of other antigens and substances in serum. Given its good performance, the proposed dQDs-FLISA method offers great potential for simultaneous and quantitative detection of other biomarkers in in vitro diagnostic (IVD). [ABSTRACT FROM AUTHOR]

Colloidal quantum dots (QDs) have been widely used as fluorescent labeling materials in the field of in vitro diagnostics (IVD). In spite of the significant development of direct single-signal response fluorescent immunosensors, it is still a challenge to obtain the high accuracy and sensitivity of quantitative results of these sensors due to the influence of several factors unrelated to the analysis. Herein, we present a delicate design and synthesis of dual-color QDs bilayer nanobeads (red and green QDs nanobeads, R&G-QD nanobeads), achieving the precisely control of growth of the silicon layer with different proportions of ammonia, and inhibiting fluorescence resonance energy transfer (FRET) successfully. The dual-color QDs bilayer nanobeads have realized dual-channel detection (the acquisition of two emission wavelengths) of the target, and avoided the problems of low analytical accuracy and imprecise sensitivity of single-signal response. A proof-of-concept demonstration of a fluorescence-linked immunosorbent assay (FLISA) platform is conducted for the quantitative detection of prostate-specific antigen (PSA) with a sensitivity of 49 pg/mL and a linear range of 0.2–800 ng/mL. The dual-color QDs bilayer nanobeads FLISA platform with high sensitivity and accuracy holds great potential for early diagnosis of cancer or other diseases and provides a new approach for IVD. [Display omitted] • Dual-color QDs bilayer nanobeads were designed to synthesize by layer-by-layer assembly method. • Controlled growth of alternate red and green QD layers on pre-synthesized SiO 2 beads can suppress energy transfer. • Such dual-color QDs nanobeads deliver dual single calibration and can avoid errors caused from the single signal response. • A proof-of-concept demonstration of a FLISA platform is conducted for detecting PSA with high sensitivity and accuracy. [ABSTRACT FROM AUTHOR]

• The nanoflower-shaped Bi 2 S 3 had a high photoelectric activity and large specific surface area. • Toehold-mediated strand displacement reaction without enzyme was used to recycle target for signal amplification. • TSDR strategy improved the sensitivity and reaction efficiency of the biosensor. Herein, a photoelectrochemical (PEC) biosensor we proposed was used to detect let-7a via studying the photocurrent response change resulted from the change of the toehold-mediated strand displacement reaction (TSDR) between the AuNPs-H2 (a conjugate of gold nanoparticles and DNA strand H2) and let-7a. Without the participation of let-7a, after the alkaline phosphatase (ALP) were immobilized at Bi 2 S 3 surface through modified on the AuNPs-H2, ALP converted the ascorbic acid 2-phosphate (AAP) to produce ascorbic acid (AA), AA could be catalytically oxidized to provide electrons, resulting in the enhancement of photocurrent signal on the Bi 2 S 3. In contrast, the conjugate of ALP@AuNPs-H2 (a conjugate of ALP and AuNPs-H2) were compelled away from Bi 2 S 3 surface when the let-7a participated in the process and hybridized with H1. The fuel DNA assisted the recycling of the let-7a via another TSDR, the recovered let-7a could participate in the next enzyme-free cycle. The PEC biosensor not only exploited TSDR signal amplification strategy, but also achieved good linearity in the range of 0.01 nM-1000 nM with a low detection limit of 6.7 pM. Due to the excellent performance, we believe it will offer more opportunities for bioanalysis and clinical biomedicine. [ABSTRACT FROM AUTHOR]

Nerve agents such as Sarin, Soman and Tabun are all very toxic organophosphorus (OP) compounds capable of being used as chemical weapons. Unfortunately, the ability to rapidly sense OPCs is still an ongoing challenging. In this report, we designed and synthesized a fast-response fluorescent probe ( 1 ) for the detection of OP nerve agent simulants with high selectivity and sensitivity. The probe contains an electron donor-to-acceptor system in which a BODIPY moiety acts as a reporter group and the hydroxyl group as a reaction site for nerve agent simulants. The electron deficient nitrile group was introduced to lower the background fluorescence of the probe. A significant enhancement of fluorescence intensities at 531 nm was observed upon gradual addition of nerve agent simulants on excitation at 480 nm. The emission intensity rapidly reached its maximum within only 540 s on adding 6 equiv. of nerve agent simulants with a quantum yield of the reaction product of 0.808, 81-fold larger than the probe itself (Ф = 0.01). A color change in following a paper test experiment verified its portable application. [ABSTRACT FROM AUTHOR]