Facile Preparation of WO3−x Dots with Remarkably Low Toxicity and Uncompromised Activity as Co-reactants for Clinical Diagnosis by Electrochemiluminescence

The exceptional nature of WO_3−x dots has inspired widespread interest, but it is still a significant challenge to synthesize high-quality WO_3−x dots without using unstable reactants, expensive equipment, and complex synthetic processes. Herein, the synthesis of ligand-free WO_3−x dots is reported that are highly dispersible and rich in oxygen vacancies by a simple but straightforward exfoliation of bulk WS₂ and a mild follow-up chemical conversion. Surprisingly, the WO_3−x dots emerged as co-reactants for the electrochemiluminescence (ECL) of Ru(bpy)₃²⁺ with a comparable ECL efficiency to the well-known Ru(bpy)₃²⁺/tripropylamine (TPrA) system. Moreover, compared to TPrA, whose toxicity remains a critical issue of concern, the WO_3−x dots were ca. 300-fold less toxic. The potency of WO_3−x dots was further explored in the detection of circulating tumor cells (CTCs) with the most competitive limit of detection so far.     

Electrogenerated Chemiluminescence Sensor Based on Tris(2,2′‐bipyridine)ruthenium(II)‐Immobilized Natural Clay and Ionic Liquid

A novel electrogenerated chemiluminescence (ECL) sensor based on natural clay and ionic liquid was fabricated. Tris(2,2′‐bipyridine)ruthenium(II) (Ru(bpy)₃²⁺) was immobilized on natural clay surface through simple adsorption. An ECL sensor was prepared by mixing Ru(bpy)₃²⁺‐incorporated clay, graphite powder and an ionic liquid (1‐butyl‐3‐methylimidazolium hexafluorophosphate) as the binder. The electrochemical behavior and ECL of the immobilized Ru(bpy)₃²⁺ was investigated. It was observed that the ECL of immobilized Ru(bpy)₃²⁺ was activated by the ionic liquid. The proposed ECL sensor showed high sensitivity to tri‐n‐propylamine (TPrA) and the detection limit was found to be 20 pM. In addition, the ECL sensor displayed good stability for TPrA detection and long‐term storage stability.     

Inhibition of Ru Complex Electrochemiluminescence by Phenols and Anilines

The effect of 30 phenols and anilines on typical Ru complex electrochemiluminescence (ECL) was systematically investigated under different conditions. It was found that all the tested compounds showed an ECL inhibiting signal. The magnitude of ECL inhibition was related to the position of the substituting group in the benzene ring and decreased in the following order: meta‐>ortho‐>para‐. The oxidation potential of the tested compounds, the ECL spectra and UV‐visible absorption spectra of Ru(bpy) /tripropylamine (TPrA) in the presence of phenols and anilines, and the direct ECL between Ru(bpy) and phenols/aniline were studied. The mechanism of ECL inhibition has been proposed due to energy transfer from the excited state Ru(bpy) to a quinone or ketone or their polymer formed by electro‐oxidation of phenols and anilines. The potential of analytical application was explored by use of the inhibited ECL. The results demonstrate that numerous compounds are detectable with the detection limits in the range of 10^?8–10^?9 mol/L for Ru(bpy) /TPrA system and in the range of 10^?6–10^?7 mol/L for Ru(bpy) /C₂O system, respectively.     

Amine‐Rich Nitrogen‐Doped Carbon Nanodots as a Platform for Self‐Enhancing Electrochemiluminescence

Amine‐rich nitrogen‐doped carbon nanodots (NCNDs) have been successfully used as co‐reactant in electrochemiluminescence (ECL) processes. Primary or tertiary amino groups on NCNDs have been studied as co‐reactant sites for Ru(bpy)₃²⁺ ECL, showing their eligibility as powerful alternatives to tripropylamine (TPrA). We also report the synthesis and ECL behavior of a new covalently linked hybrid of NCNDs and Ru(bpy)₃²⁺. Notably, the NCNDs in the hybrid act both as carrier for ECL labels and as co‐reactant for ECL generation. As a result, the hybrid shows a higher ECL emission as compared to the combination of the individual components, suggesting the self‐enhancing ECL of the ruthenium complex due to an intramolecular electron transfer process.     

Electrochemiluminescence of Tris(2,2′‐bipyridine)ruthenium with Various Co‐reactants under Ultrasound Irradiation

Electrochemiluminescence (ECL) of tris(2,2′‐bipyridine)ruthenium, Ru(bpy)₃²⁺ in the presence of various co‐reactants, such as tripropylamine (TPA), oxalate ion (C₂O₄^2?), ascorbic acid (H₂A) and dehydroascorbic acid (DHA), were investigated under ultrasound irradiation. In sono‐ECL experiments, an indium‐thin‐oxide (ITO) was used as working electrode, and a titanium tipped sonic horn probe (diameter 2 mm) which operated at a frequency of 20 kHz was set in the front of the ITO electrode. Under the ultrasound irradiation, ECL signals were found to be significantly enhanced when TPA and C₂O₄^2? were used as co‐reactants, only slightly enhanced in Ru(bpy)₃²⁺/DHA system, but total quenched in Ru(bpy)₃²⁺/H₂A system. The difference of Ru(bpy)₃²⁺ ECL behaviors for various co‐reactant could to be due to the different kinetics of catalytic reactions associated in ECL schemes. ECL quenching effect observed in Ru(bpy)₃²⁺/H₂A system was suggested to be due to electron transfer (ET) route between the excited state *Ru(bpy)₃²⁺ and ascorbate anion HA^? diffused from the bulk solution, where the diffusional HA^? species served as electron donor. The effect becomes more pronounced upon sonication because the effective collision frequency between *Ru(bpy)₃²⁺ and HA^? would be significantly increased by the enhanced mass transport effect of ultrasound.     

Formation of [Ru(bpy)3]2+‐Containing Microstructures Induced by Electrostatic Assembly and Their Application in Solid‐State Detection of Electrochemiluminescence

Tris(2,2′‐bipyridine)ruthenium(II) ([Ru(bpy)₃]²⁺) is one of the most extensively studied and used electrochemiluminescent (ECL) compounds owing to its superior properties, which include high sensitivity and stability under moderate conditions in aqueous solution. In this paper we present a simple method for the preparation of [Ru(bpy)₃]²⁺‐containing microstructures based on electrostatic assembly. The formation of such microstructures occurs in a single process by direct mixing of aqueous solutions of [Ru(bpy)₃]Cl₂ and K₃[Fe(CN)₆] at room temperature. The electrostatic interactions between [Ru(bpy)₃]²⁺ cations and [Fe(CN)₆]^3? anions cause them to assemble into the resulting microstructures. Both the molar ratio and concentration of reactants were found to have strong influences on the formation of these microstructures. Most importantly, the resulting [Ru(bpy)₃]²⁺‐containing microstructures exhibit excellent ECL behavior and, therefore, hold great promise for solid‐state ECL detection in capillary electrophoresis (CE) or CE microchips.     

Highly Sensitive and Selective Detection of Pb(II) by NH2−SiO2/Ru(bpy)32+−UiO66 based Solid‐state ECL Sensor

In this study, a novel electrochemiluminescence (ECL) sensor for highly sensitive and selective detection of Pb(II) was developed based on Ru(bpy)₃²⁺ encapsulated UiO66 metal‐organic‐framework (Ru(bpy)₃²⁺?UiO66 MOF) and ?NH₂ group functionalized silica (NH₂?SiO₂). The NH₂?SiO₂ with large surface area provided an excellent platform for the ECL sensor. As numerous exposed carboxyl groups were present on UiO66 backbone, the Ru(bpy)₃²⁺?UiO66 could be steadily immobilized to NH₂?SiO₂ by forming amide bonds. Meanwhile, the introduced UiO66 MOF which used for the encapsulation of Ru(bpy)₃²⁺, significantly enhanced the ECL efficiency of the proposed sensor, as it possessed a large specific surface area and porosity for the loading of Ru(bpy)₃²⁺. Moreover, a high quenching effect on ECL intensity was obtained in the presence of Pb(II) in the electrolyte. Under the optimal conditions, the quenched ECL intensity showed a good linear relationship within Pb(II) concentration in the range from 1.0×10^?6 to 1.0×10² μM, with a detection limit of 1.0×10^?7 μM (S/N=3). The proposed sensor for Pb(II) detection was simple in operation, rapid in testing, stable in signal, and showed a good anti‐interference ability to some other metal ions. Besides, its application for detecting Pb(II) in a real sample was also investigated here. This work provides a potential platform for metal ions detection in environmental monitoring field.     

A Solid‐State Electrochemiluminescence Immunosensor Based on MWCNTs‐Nafion and Ru(bpy)32+/Nano‐Pt Nanocomposites for Detection of α‐Fetoprotein

A approach was successfully employed for constructing a solid‐state electrochemiluminescence (ECL) immunosensor by layer‐by‐layer self‐assembly of multiwall carbon nanotubes (MWCNTs)‐Nafion composite film, Ru(bpy)₃²⁺/nano‐Pt aggregates (Ru‐PtNPs) and Pt nanoparticles (PtNPs). The influence of Pt nanoparticles on the ECL intensity was quantitatively evaluated by calculating the electroactive surface area of different electrodes with or without PtNPs to immobilize Ru(bpy)₃²⁺. The principle of ECL detection for target α‐fetoprotein antigen (AFP) was based on the increment of resistance after immunoreaction, which led to a decrease in ECL intensity. The linear response range was 0.01–10 ng mL^?1 with the detection limit of 3.3 pg mL^?1. The immunosensor exhibited advantages of simple preparation and operation, high sensitivity and good selectivity.     

Flow injection analysis of gallic acid with inhibited electrochemiluminescence detection

A flow injection (FI)–electrochemiluminescent (ECL) method has been developed for the determination of gallic acid, based on an inhibition effect on the Ru(bpy)₃²⁺/tri-n-propylamine (TPrA) ECL system in pH 8.0 phosphate buffer solution. The method is simple and convenient with a determination limit of 9.0×10^–9 mol/L and a dynamic concentration range of 2×10^–8–2×10^–5 mol/L. The relative standard deviation (RSD) was 1.0% for 1.0×10^–6 mol/L gallic acid (n=11). It was successfully applied to the determination of gallic acid in Chinese proprietary medicine—Jianming Yanhou Pian. The inhibition mechanism proposed for the quenching effect of the gallic acid on the Ru(bpy)₃²⁺/TPrA ECL system was the interaction of electrogenerated Ru(bpy)₃^2+* and o-benzoquinone derivative at the electrode surface. The ECL emission spectra and UV-visible absorption spectra were applied to confirm the mechanism.     

Quenching Behavior of the Electrochemiluminescence of Ru(bpy)32+/TPrA System by Phenols on a Smartphone-Based Sensor

Phenolic compounds such as vanillic and p-coumaric acids are pollutants of major concern in the agro-industrial processing, thereby their effective detection in the industrial environment is essential to reduce exposure. Herein, we present the quenching effect of these compounds on the electrochemiluminescence (ECL) of the Ru(bpy)₃²⁺/TPrA (TPrA=tri-n-propylamine) system at a disposable screen-printed carbon electrode. Transient ECL profiles are obtained from multiple video frames following 1.2 V application by a smartphone-based ECL sensor. A wide range of detection was achieved using the sensor with limit of detection of 0.26 μM and 0.68 μM for vanillic and p-coumaric acids, respectively. The estimated quenching constants determined that the quenching efficiency of vanillic acid is at least two-fold that of p-coumaric acid under the current detection conditions. The present ECL quenching approach provided an effective method to detect phenolic compounds using a low-cost, portable smartphone-based ECL sensor.     

A Novel Electrogenerated Chemiluminescence (ECL) Sensor Based on Ru(bpy)32+‐Doped Titania Nanoparticles Dispersed in Nafion on Glassy Carbon Electrode

A novel electrogenerated chemiluminescence (ECL) sensor based on Ru(bpy)₃²⁺‐doped titania (RuDT) nanoparticles dispersed in a perfluorosulfonated ionomer (Nafion) on a glassy carbon electrode (GCE) was developed in this paper. The electroactive component‐Ru(bpy)₃²⁺ was entrapped within the titania nanoparticles by the inverse microemulsion polymerization process that produced spherical sensors in the size region of 38±3 nm. The RuDT nanoparticles were characterized by electrochemical, transmission electron and scanning microscopy technology. The Ru(bpy)₃²⁺ encapsulation interior of the titania nanoparticles maintains its ECL efficiency and also reduces Ru(bpy)₃²⁺ leaching from the titania matrix when immersed in water due to the electrostatic interaction. This is the first attempt to prepare the RuDT nanoparticles and extend the application of electroactive component‐doped nanoparticles into the field of ECL. Since a large amount of Ru(bpy)₃²⁺ was immobilized three‐dimensionally on the electrode, the Ru(bpy)₃²⁺ ECL signal could be enhanced greatly, which finally resulted in the increased sensitivity. The ECL analytical performance of this ECL sensor for tripropylamine (TPA) was investigated in detail. This sensor shows a detection limit of 1 nmol/L for TPA. Furthermore, the present ECL sensor displays outstanding long‐term stability.     

Electochemiluminescence of CdTe/CdS Quantum Dots with Triproprylamine as Coreactant in Aqueous Solution at a Lower Potential and Its Application for Highly Sensitive and Selective Detection of Cu2+

Anodic electrochemiluminescence (ECL) of 3‐mercaptopropionic acid (MPA)‐ capped CdTe/CdS core‐shell quantum dots (QDs) with tripropylamine (TPrA) as the co‐reactant were studied in aqueous (Tris buffer) solution for the first time. The results suggest that the oxidation of TPrA at a glassy carbon electrode (GCE) surface participated in the ECL of QDs, and the onset potential and the intensity of ECL of CdTe/CdS QDs were affected seriously by TPrA, as the co‐reactant, in Tris buffer solution. The onset potential of ECL in this new system was about +0.5 V (vs. Ag/AgCl) and the ECL intensity greatly enhanced when TPrA was present. Various influencing factors, such as the electrolyte, pH, QDs concentration, potential range and scan rates on the ECL were studied. Based on the selective quenching by Cu²⁺ to the light emission from CdTe/CdS QDs/TPrA system, a highly sensitive and selective method for the determination of Cu²⁺ was developed. At the optimal conditions, the relative ECL intensity, I₀/I, was proportional to the concentration of Cu²⁺ from 14 nM to 0.21 μM with the detection limit of 6.1 nM based on the signal‐to‐noise ratio of 3. The possible ECL mechanism of QDs and the quenching mechanism of ECL were proposed.     

Electrochemiluminescence Distance and Reactivity of Coreactants Determine the Sensitivity of Bead-Based Immunoassays

Herein we report the study of electrochemiluminescence (ECL) generation by tris(2,2′-bipyridyl)ruthenium (Ru(bpy)₃²⁺) and five tertiary amine coreactants. The ECL distance and lifetime of coreactant radical cations were measured by ECL self-interference spectroscopy. And the reactivity of coreactants was quantitatively evaluated in terms of integrated ECL intensity. By statistical analysis of ECL images of single Ru(bpy)₃²⁺-labeled microbeads, we propose that ECL distance and reactivity of coreactant codetermine the emission intensity and thus the sensitivity of immunoassay. 2,2-bis(hydroxymethyl)-2,2′,2′′-nitrilotriethanol (BIS-TRIS) can well balance ECL distance-reactivity trade-off and enhance the sensitivity by 236 % compared with tri-n-propylamine (TPrA) in the bead-based immunoassay of carcinoembryonic antigen. The study brings an insightful understanding of ECL generation in bead-based immunoassay and a way of maximizing the analytical sensitivity from the aspect of coreactant.     

Quality control of PET radiopharmaceuticals by high‐performance liquid chromatography with tris(2,2′‐bipyridyl)ruthenium(II) electrogenerated chemiluminescence detection

A highly sensitive reversed‐phase liquid chromatographic (HPLC) method was investigated to analyze a range of positron emission tomography (PET) radiopharmaceuticals using electrogenerated chemiluminescence (ECL) detection. ECL is based on the reaction of PET molecules with tris(2,2′‐bipyridyl)ruthenium(III) [Ru(bpy)₃³⁺], which is generated through the on‐line electro‐oxidation of Ru(bpy)₃²⁺. In 21 different radiopharmaceuticals studied, 18 compounds could be detected with detection limits (signal‐to‐noise ratio = 3) of 0.12–72 ng/mL per 20 μL injection. Sufficient reproducibility and linearity were obtained for the quantitative determination of PET molecules in pharmaceutical fluid. This method could be successfully applied to quality control tests of PET radiopharmaceuticals with ultra‐high specific radioactivity. Copyright © 2009 John Wiley & Sons, Ltd.     

Study on Electrochemiluminesce of Ru(bpy3)2+ Immobilized in a Titania Sol‐Gel Membrane

The immobilization of tris(2,2′‐bipyridyl)ruthenium(II), Ru(bpy)₃²⁺, at a glassy carbon electrode was achieved by entrapping the Ru(bpy)₃²⁺ in a vapor deposited titania sol‐gel membrane. The electrogenerated chemiluminescence (ECL) of the immobilized Ru(bpy)₃²⁺ was studied. The Ru(bpy)₃²⁺ modified electrode showed a fast ECL response to both oxalate and proline. The ECL intensity was linearly related to concentrations of oxalate and proline over the ranges from 20 to 700 μmol L^?1 and 20 to 600 μmol L^?1, respectively. The detection limits for oxalate and proline at 3σ were 5.0 μmol L^?1 and 4.0 μmol L^?1, respectively. This electrode possessed good precision and stability for oxalate and proline determinations. The electrogenerated chemiluminescence mechanism of proline system was discussed. This work provided a new way for the immobilization of Ru(bpy)₃²⁺ and the application of titania sol‐gel membrane in electrogenerated chemiluminescence.     

Control of Excitation and Quenching in Multi‐colour Electrogenerated Chemiluminescence Systems through Choice of Co‐reactant

We demonstrate a new approach to manipulate the selective emission in mixed electrogenerated chemiluminescence (ECL) systems, where subtle changes in co‐reactant properties are exploited to control the relative electron‐transfer processes of excitation and quenching. Two closely related tertiary‐amine co‐reactants, tri‐n‐propylamine and N,N‐diisopropylethylamine, generate remarkably different emission profiles: one provides distinct green and red ECL from [Ir(ppy)₃] (ppy=2‐phenylpyridinato‐C2,N) and a [Ru(bpy)₃]²⁺ (bpy=2,2′‐bipyridine) derivative at different applied potentials, whereas the other generates both emissions simultaneously across a wide potential range. These phenomena can be rationalized through the relative exergonicities of electron‐transfer quenching of the excited states, in conjunction with the change in concentration of the quenchers over the applied potential range.     

Characterization of the low-oxidation-potential electrogenerated chemiluminescence of tris(2,2′-bipyridine)ruthenium(II) with tri-n-propylamine as coreactant

The electrogenerated chemiluminescence (ECL) of the Ru(bpy)₃²⁺ (bpy, 2,2′-bipyridine)/tri-n-propylamine (TPrA) system can be produced at an oxidation-potential well before the oxidation of Ru(bpy)₃²⁺. Here, we describe the unique features of the low-oxidation-potential (LOP) ECL. The LOP ECL exhibited strong dependence on solution pH with the maximum emission at pH  7.7. Compared with the conventional ECL, the LOP ECL was much more significantly diminished at high pH (>10), probably due to the short lifetime of TPrA cation radical which is a crucial intermediate for the LOP emission. It was also found that the preceding deprotonation step played an important role in TPrA oxidation at neutral pH and would remarkably influence the emission intensity. As excess intermediate radicals were produced upon rapid TPrA oxidation, only 5 mM TPrA was needed to achieve the maximum LOP ECL intensity in detecting trace Ru(bpy)₃²⁺ (<1 μM) and the LOP ECL response to Ru(bpy)₃²⁺ concentration was linear. Compared with the conventional Ru(bpy)₃²⁺/TPrA ECL, the LOP ECL technique not only produces higher emission intensity at lower oxidation-potential, but also significantly reduces the amount of the coreactant.     

Tris (2,2′-bipyridyl)ruthenium(II) electrogenerated chemiluminescence in analytical science

Ru(bpy) ₃ ²⁺ electrogenerated chemiluminescence (CL) has rapidly gained importance as a sensitive and selective detection method in analytical science. The Ru(bpy) ₃ ²⁺ ECL is observed when Ru(bpy) ₃ ³⁺ reacts with Ru(bpy) ₃ ⁺ and yields an excited state Ru(bpy) ₃ ^2+* . ECL emission can also be obtained when a variety of oxidants and reductants react with the reduced or oxidized forms of Ru(bpy) ₃ ²⁺ . Either the reductant or the oxidant can be treated as an analyte. The Ru(bpy) ₃ ²⁺ ECL is used as a detection method for the determination of oxalate and a variety of amine-containing analytes without derivatization in flowing streams such as flow injection and HPLC. When the ECL format is used as a detector for HPLC, unstable post-column reagent addition can often be eliminated and, the problems of both sample dilution and band broadening can be avoided because the Ru(bpy) ₃ ³⁺ species are generatedin situ in the reaction/observation flow cell. Since NADH is sensitively detected with the Ru(bpy) ₃ ²⁺ ECL, many clinically important analytes can be detected by coupling them to dehydrogenase enzymes that utilize -nicotinamide adenine cofactors to convert NAD⁺ to NADH. Ru(bpy) ₃ ²⁺ -derivatives are used as CL labels for immunoassay and PCR assay with Ru(bpy) ₃ ²⁺ /tripropylamine ECL system. The Ru(bpy) ₃ ²⁺ ECL label can be sensitively determined at subpicomolar concentrations, along with an extremely wide dynamic range of greater than six orders of magnitude. Furthermore, it can eliminate disposal and lifetime problems inherent in radio immunoassays. In this paper, basic principles of the Ru(bpy) ₃ ²⁺ ECL are discussed. In addition, analytical applications of the Ru(bpy) ₃ ²⁺ ECL are illustrated with examples.     

Enhanced Electrochemiluminescence from a Stoichiometric Ruthenium(II)–Iridium(III) Complex Soft Salt

Electrochemiluminescence (ECL) and electrochemistry are reported for a heterometallic soft salt, [Ru(dtbubpy)₃][Ir(ppy)₂(CN)₂]₂ ( [Ir][Ru][Ir] ), consisting of a 2:1 ratio of complementary charged Ru and Ir complexes possessing two different emission colors. The [Ru]²⁺ and [Ir]^? moieties in the [Ir][Ru][Ir] greatly reduce the energy required to produce ECL. Though ECL intensity in the annihilation path was enhanced 18× relative to that of [Ru(bpy)₃]²⁺, ECL in the co‐reactant path with tri‐n‐propylamine was enhanced a further 4×. Spooling spectroscopy gives insight into ECL mechanisms: the unique light emission at 634 nm is due to the [Ru]²⁺* excited state and no [Ir]^?* was generated in either route. Overall, the soft salt system is anticipated to be attractive and suitable for the development of efficient and low‐energy‐cost ECL detection systems.     

A Sandwich‐type Electrochemiluminescence Aptasensor for Thrombin Based on Functional Co‐polymer Electrode Using Ru(bpy)32+ Doped Nanocomposites as Signal‐amplifying Tags

Herein, a signal‐on sandwich‐type electrochemiluminescence (ECL) aptasensor for the detection of thrombin (TB) was proposed. The graphene (GR) doped thionine (TH) was electropolymerized synchronously on the bare glassy carbon electrode (GCE) to form co‐polymer (PTG) electrode. The gold nanoparticles (AuNPs) were decorated on the surface of the PTG by in‐situ electrodeposition, and the functional co‐polymer (PTG‐AuNPs) electrode was utilized as sensing interface. Then, TB binding aptamer I (TBA I) as capture probes were modified on the PTG‐AuNPs electrode to capture TB, and Ru(bpy)₃²⁺/silver nanoparticles doped silica core‐shell nanocomposites‐labeled TB binding aptamer II (RuAg/SiO₂NPs@TBA II) were used as signal probes to further bind TB, resulting in a sandwich structure. With the assistant of silica shell and AgNPs, the enrichment and luminous efficiency of Ru(bpy)₃²⁺ were significantly improved. Under the synergy of PTG‐AuNPs and RuAg/SiO₂NPs, the ECL signal was dramatically increased. The proposed ECL aptasensor displayed a wide linear range from 2 fM to 2 pM with the detection limit of 1 fM, which is comparable or better than that in reported ECL aptasensors for TB using Ru(bpy)₃²⁺ and its derivatives as the luminescent substance. The excellent sensitivity makes the proposed aptasensor a promising potential in pharmaceutical and clinical analysis.