Electrogenerated chemiluminescence from tris(2,2'-bipyridyl)ruthenium(II) immobilized in titania-perfluorosulfonated ionomer composite films

Electrochemical behavior and electrogenerated chemiluminescence (ECL) of tris(2,2'-bipyridyl)ruthenium(II) (Ru(bpy)3(2+)) immobilized in sol-gel-derived titania TiO2)-Nafion composite films coated on a glassy carbon electrode have been investigated. The electroactivity of Ru(bpy)3(2+) ion exchanged into the composite films and its ECL behavior were strongly dependent upon the amount of Nafion incorporated into the TiO2-Nafion composite films. The ECL sensor of Ru(bpy)32+ immobilized in a TiO2-Nafion composite with 50% Nafion content showed the maximum ECL intensities for both tripropylamine (TPA) and sodium oxalate in 0.05 M phosphate buffer solution at pH 7. Detection limits were 0.1 microM for TPA and 1.0 microM for oxalate (S/N = 3) with a linear range of 3 orders of magnitude in concentration. The present ECL sensor showed improved ECL sensitivity and long-term stability, as compared to the ECL sensors based on pure Nafion films. The present Ru(bpy)3(2+) ECL sensor based on TiO2-Nafion (50%) composite films was applied as an HPLC detector for the determination of erythromycin in human urine samples. The present Ru(bpy)3(2+) ECL sensor was stable in the mobile phase containing a high content of organic solvent (30%, v/v), in contrast to a pure Nafion-based Ru(bpy)3(2+) ECL sensor. The detection limit for erythromycin was 1.0 microM, with a linear range of 3 orders of magnitude in concentration.     

Method for effective immobilization of Ru(bpy)(3)2+ on an electrode surface for solid-state electrochemiluminescene detection

A novel method for effective immobilization of Ru(bpy)3(2+) on an electrode surface is developed. The whole process involves two steps: the electrostatic interactions between citrate-capped gold nanoparticles (AuNPs) and Ru(bpy)3Cl2 in aqueous medium were used to fabricate Ru(bpy)(3)2+-AuNP aggregates (Ru-AuNPs) first, and then the Au-S interactions between as-formed Ru-AuNPs and sulfhydryl groups were used to effectively immobilize the Ru-AuNPs on a sulfhydryl-derivated indium tin oxide (ITO) electrode surface. As-prepared ITO electrode shows excellent stability, and the ECL active species Ru(bpy)3(2+) contained therein exhibit excellent ECL behaviors.     

The effects of nonionic surfactants on the tris(2,2'-bipyridyl)ruthenium(II)--tripropylamine electrochemiluminescence system

The electrochemistry and electrogenerated chemiluminescence (ECL) of Ru(bpy)3(2+) (bpy = 2,2'-bipyridyl) were studied in the presence of the nonionic surfactants Triton X-100, Thesit, and Nonidet P40. The anodic oxidation of Ru(bpy)3(2+) produces ECL in the presence of tri-n-propylamine in both aqueous and surfactant solutions. Increases in both ECL efficiency (> or =8-fold) and duration of the ECL signal were observed in surfactant media. A shift to lower energies of the Ru(bpy)3(2+) ECL emission by approximately 8 nm was also observed. The one-electron oxidation of Ru(bpy)3(2+) to Ru(bpy)3(3t) occurs at + 1.03 V vs Ag/AgCl in aqueous buffered (0.2 M potassium phosphate) solution as found by square wave voltammetry. This potential did not shift in surfactant systems, indicating that the redshifts in ECL emission are due to stabilization of ligand pi* orbitals in the metal-to-ligand charge-transfer excited state. These results are consistent with hydrophobic interactions between Ru(bpy)3(2+) and the nonionic surfactants.     

Electrochemiluminescence sensor based on partial sulfonation of polystyrene with carbon nanotubes

Herein, homogenously partial sulfonation of polystyrene (PSP) was performed. An effective electrochemiluminescence (ECL) sensor based on PSP with carbon nanotube (CNTs) composite film was developed. Cyclic voltammetry and electrochemical impendence spectroscopy were applied to characterize this composite film. The PSP was used as an immobilization matrix to entrap the ECL reagent Ru(bpy)3(2+) due to the electrostatic interactions between sulfonic acid groups and Ru(bpy)3(2+) cations. The introduction of CNTs into PSP acted not only as a conducting pathway to accelerate the electron transfer but also as a proper matrix to immobilize Ru(bpy)3(2+) on the electrode by hydrophobic interaction. Furthermore, the results indicated the ECL intensity produced at this composite film was over 3-fold compared with that of the pure PSP film due to the electrocatalytic activity of the CNTs. Such a sensor was verified by the sensitive determinations of 2-(dibutylamino)ethanol and tripropylamine.     

Surfactant chain length effects on the light emission of tris(2,2'-bipyridyl)ruthenium(II)/ tripropylamine electrogenerated chemiluminescence

The effects of nonionic surfactant chain length on the properties of tris(2,2'-bipyridyl)ruthenium(II) (Ru(bpy)3(2+) where bpy = 2,2'-bipyridine) electrochemiluminescence (ECL) have been investigated. The electrochemistry, photophysics, and ECL of Ru(bpy)3(2+) in the presence of a series of nonionic surfactants are reported (Triton X-100, 114, 165, 405, 305, and 705-70). These surfactants differ in the number of poly(ethylene oxide) units incorporated into the surfactant molecule. The anodic oxidation of Ru(bpy)3(2+) produces ECL in the presence of tri-n-propylamine (TPrA) in aqueous surfactant solution. Increases in ECL efficiency (> or = 5-fold) and TPrA oxidation current (> or = 2-fold) have been observed in surfactant media. Slight decreases in ECL intensity are observed as the chain length of the nonionic surfactant increases. The data supports adsorption of surfactant on the electrode surface, thus facilitating TPrA and Ru(bpy)3(2+) oxidation and leading to higher ECL efficiencies.     

Electrogenerated chemiluminescence from R(bpy)3(2+) ion-exchanged in carbon nanotube/perfluorosulfonated ionomer composite films

The electrochemistry and electrogenerated chemiluminescence (ECL) of ruthenium(II) tris(bipyridine) (Ru(bpy)(3)(2+)) ion-exchanged in carbon nanotube (CNT)/Nafion composite films were investigated with tripropylamine (TPA) as a coreactant at a glassy carbon (GC) electrode. The major goal of this work was to investigate and develop new materials and immobilization approaches for the fabrication of ECL-based sensors with improved sensitivity, reactivity, and long-term stability. Ru(bpy)(3)(2+) could be strongly incorporated into Nafion film, but the rate of charge transfer was relative slow and its stability was also problematic. The interfusion of CNT in Nafion resulted in a high peak current of Ru(bpy)(3)(2+) and high ECL intensity. The results indicated that the composite film had more open structures and a larger surface area allowing faster diffusion of Ru(bpy)(3)(2+) and that the CNT could adsorb Ru(bpy)(3)(2+) and also acted as conducting pathways to connect Ru(bpy)(3)(2+) sites to the electrode. In the present work, the sensitivity of the ECL system at the CNT/Nafion film-modified electrodes was more than 2 orders of magnitude higher than that observed at a silica/Nafion composite film-modified electrode and 3 orders of magnitude higher than that at pure Nafion films. The CNT/Nafion composite film-modified GC electrodes also exhibited long-term stability.     

Electrochemiluminescent detection based on solid-phase extraction at tris(2,2'-bipyridyl)ruthenium(II)-modified ceramic carbon electrode

A sensitive electrochemiluminescent detection scheme by solid-phase extraction at Ru(bpy)3(2+)-modified ceramic carbon electrodes (CCEs) was developed. The as-prepared Ru(bpy)3(2+)-modified CCEs show much better long-term stability than other Nafion-based Ru(bpy)3(2+)-modified electrodes and enjoy the inherent advantages of CCEs. The log-log calibration plot for dioxopromethazine is linear from 1.0 x 10(-9) to 1.0 x 10(-4) mol L(-1) using the new detection scheme. The detection limit is 6.6 x 10(-10) mol L(-1) at a signal-to-noise ratio of 3. The new scheme improves the sensitivity by approximately 3 orders of magnitude, which is the most sensitive Ru(bpy)3(2+) ECL method. The scheme allows the detection of dioxopromethazine in a urine sample within 3 min. Since Ru(bpy)3(2+) ECL is a powerful technique for determination of numerous amine-containing substances, the new detection scheme holds great promise in measurement of free concentrations, investigation of protein-drug interactions and DNA-drug interactions, pharmaceutical analysis, and so on.     

Electrogenerated chemiluminescence of tris (2,2'-bipyridyl)ruthenium(II) ion-exchanged in nafion-silica composite films

The voltammetry and electrogenerated chemiluminescence (ECL) of tris(2,2'-bipyridyl)ruthenium(II) (Ru(bpy)3 2+) ion-exchanged in Nafion and Nafion-silica composite materials have been investigated. The major goal of this work was to investigate and develop new materials and immobilization approaches for the fabrication of ECL-based sensors with improved reactivity and long-term stability. Nation-silica composite materials with varying contents of Nation (53-100 wt% relative to silica) were prepared via the two-step acid/base hydrolysis and condensation of tetramethoxysilane. The Nafion doped sols were spin cast on glassy carbon electrodes, dried, and then ion-exchanged with Ru(bpy)3 2+. The shapes of the cyclic voltammetric curves and the amount of Ru(bpy)3 2+ exchanged into the films strongly depends on the amount of Nafion incorporated into the hybrid sol. Nafion-silica films with a low content of Nafion ion-exchanged less Ru(bpy)3 2+ and exhibited tail-shaped voltammetry at 100 mV/s. The ECL of immobilized Ru(bpy)3 2+ in the presence of either tripropylamine or sodium oxalate in pH 5 acetate buffer was also strongly dependent on the amount of Nafion introduced into the composite with greater ECL observed for the Nafion-silica films relative to pure Nafion.     

Electrogenerated chemiluminescence sensor for formaldehyde based on Ru(bpy)32+-doped silica nanoparticles modified Au electrode

A novel and sensitive electrogenerated chemiluminescence (ECL) sensor for formaldehyde was developed with the amine-functionalized Ru(bpy)₃²⁺-doped silica nanoparticles (Ru-DSNPs) as ECL emitter. Ru(bpy)₃²⁺ doped on the silica nanoparticle can maintain its electrochemical activities, which made silica nano-beads a excellent carrier of Ru(bpy)₃²⁺ species. The uniform Ru-DSNPs (about 75 nm) were conjugated with Au electrode using mercaptoacetic acid as the intermediate to fabricate an ECL sensor for formaldehyde. The ECL analytical performances of this ECL sensor for formaldehyde based on its enhancement ECL emission of Ru(bpy)₃²⁺ were investigated in details. Under the optimum condition, the ECL intensity was linear with the formaldehyde concentration in the range of 1.0 × 10^? 8 mol/L to 1.0 × 10^? 6 mol/L. The detection limit was 6.0 × 10^? 9 mol/L (S/N = 3). This approach offered obvious advantages of being simpler, faster, and more stable compared with other sensors, and possessed great potential for formaldehyde detection which could be applied to determine directly the formaldehyde in real samples without pre-separation.     

Electrochemiluminescent detection of metal cations using a ruthenium(II) bipyridyl complex containing a crown ether moiety

The effects of metal ions on the electrochemiluminescence (ECL) properties of (bpy)2Ru(AZA-bpy) (bpy = 2,2'-bipyridine; AZA-bpy = 4-(N-aza-18-crown-6-methyl-2,2'-bipyridine) have been investigated. The electrochemistry, photophysics and ECL of Ru(bpy)3(2+) in the presence of Pb2+, Hg2+, Cu2+, and K+ are reported. The anodic oxidation of Ru(bpy)3(2+) produces ECL in the presence of tri-n-propylamine (TPrA) in 50:50 (v/v) CH3CN:H2O solution. Increases in ECL efficiency (photons generated per redox event) up to 20-fold that depend on both the concentration and nature of the metal ion have been observed, making this an interesting system for electrochemiluminescence metal ion sensing.     

A two-channel microfluidic sensor that uses anodic electrogenerated chemiluminescence as a photonic reporter of cathodic redox reactions

This paper describes a new approach for sensing electrochemically active substrates in microfluidic systems. This two-electrode sensor relies on electrochemical detection at one electrode and electrogenerated chemiluminescent (ECL) reporting at the other. Each microfabricated indium tin oxide electrode is located in a separate microfluidic channel, but the channels are connected downstream of the electrodes to maintain a complete electrical circuit. Because of laminar flow, there is no bulk mixing of the fluids in the detecting and reporting channels. This approach allows the ECL reaction to be physically and chemically decoupled from the sensing channel of the device, which greatly expands the number of analytes that can be detected. However, because the cathode and anode are connected, electron-transfer processes occurring at the sensing electrode are electrically coupled to the ECL reaction. Charge balance permits the ECL light output to be quantitatively correlated to electrochemical reductions at the cathode. The system is used to detect Fe(CN)6(3-), Ru(NH3)6(3+), and benzyl viologen and report their presence via Ru(bpy)3(2+) (bpy = bipyridine) luminescence. Each different redox target initiates ECL at a unique potential bias related to its standard redox potential. The influence of the concentrations of Ru(bpy)3(2+) and the target analytes is discussed.     

Fabrication of chemiluminescence sensor based on conducting polymer@SiO2/Nafion composite film

Tris(2,2'-bipyridyl)ruthenium (II) (Ru(bpy)2+) electrogerated chemiluminescence (ECL) sensor was fabricated by immobilization of Ru(bpy)2+ complex on conducting polymer@SiO2/Nafion composite film on surface of glassy carbon electrode. The conducting polymer@SiO2 nanocomposites were prepared by coating polyaniline (PANI), polypyrrole (PPy), and polythiophene (PTh) on the surface of the SiO2 sphere. The conducting polymer@SiO2 nanocomposite was characterized by scanning electron microscopy (SEM), Transmission electron microscopy (TEM), and atomic force microscopy (AFM). The sensitivity and reproducibility of the prepared ECL sensor to tripropylamine (TPA) was evaluated. As a result, the PPy@SiO2 composite electrode exhibited high sensitivity and good reproducibility compared to that obtained with PANI@SiO2 and PTh@SiO2 composite electrodes because of the strong interaction between PPy@SiO2 and Ru(bpy)2+ complex.     

Electrochemiluminescence of Tris(2,2'-bipyridine)ruthenium in Water at Carbon Microelectrodes

Electrochemiluminescence (ECL) of Ru(bpy)(3)(2+) in water only, without any added electrolyte or reducing agents, has been obtained at carbon interdigitated microelectrode arrays (C-IDAs) of 2 μm width and spacing. In a generation/collection biasing mode, ECL can be clearly seen with the naked eye in normal room lighting at concentrations greater than 1 mM. Using a conventional photomultiplier tube (PMT), a detection limit of 10(-)(7) M Ru(bpy)(3)(2+) has been achieved for an electrode area of 0.25 mm(2). In comparison, the ECL intensity produced at Pt-IDA of the same geometry, under identical experimental conditions, was more than 300 times less. The ECL obtained at C-IDAs is attributed to the annihilation reaction of the reduced and oxidized forms of the Ru(bpy)(3)(2+) made possible due to the small electrode spacing.     

Enhanced electrogenerated chemiluminescence in the presence of fluorinated alcohols

The electrochemistry, UV-vis absorption, photoluminescence (PL), and coreactant electrogenerated chemiluminescence (ECL) of Ru(bpy)3(2+) (where bpy=2,2'-bipyridine) have been obtained in a series of hydroxylic solvents. The solvents included fluorinated and nonfluorinated alcohols and alcohol/water mixtures. Tri-n-propylamine was used as the oxidative-reductive ECL coreactant. Blue shifts of up to 30 nm in PL emission wavelength maximums are observed compared to a Ru(bpy)3(2+)/H2O standard due to interactions of the polar excited state (i.e., *Ru(bpy)3(2+)) with the solvent media. For example, Ru(bpy)3(2+) in water has an emission maximum of 599 nm while in the more polar hexafluoropropanol and trifluoroethanol it is 562 and 571 nm, respectively. ECL spectra are similar to PL spectra, indicating the same excited state is formed in both experiments. The difference between the electrochemically reversible oxidation (Ru(bpy)3(2+/3+)) and first reduction (Ru(bpy)2(2+/1+)) correlates well with the energy gap observed in the luminescence experiments. Although the ECL is linear in all solvents with [Ru(bpy)3(2+)] ranging from 100 to 0.1 nm, little correlation between the polarity of the solvent and the ECL efficiency (phiecl=number of photons per redox event) was observed. However, dramatic increases in phiecl ranging from 6- to 270-fold were seen in mixed alcohol/water solutions.     

A multichannel microfluidic sensor that detects anodic redox reactions indirectly using anodic electrogenerated chemiluminescence

Here, we describe a new approach for detecting redoxactive targets by electrochemical oxidation and reporting their presence by electrogenerated chemiluminescence (ECL) based on electrochemical oxidation of Ru(bpy)3(2+) (bpy = 2,2'-bipyridine) and tripropylamine (TPA). This new strategy, which complements our previous reports of using ECL to signal the presence of targets undergoing electrochemical reduction, takes advantage of many of the attractive attributes of microfluidic-based electrochemical cells. These attributes include close proximity of multiple flow channels and electrodes, ability to move reagents through channels under laminar flow conditions, and the capacity to precisely place device components relative to one another using photolithography. Specifically, the microfluidic electrochemical sensor described here consists of three channels. The analyte and ECL reporting cocktail flow through separate channels, but they share a common anode. The cathode resides in a channel containing a sacrificial reductant. In this configuration, the target analyte competes with Ru(bpy)3(2+) and TPA to provide electrons for the reductant. Accordingly, in this competitive assay approach, the presence of the analyte is signaled as a lowering of the ECL intensity. In this report, the device performance characteristics are reported, and the detection of both ferrocyanide and dopamine is demonstrated at micromolar concentrations.     

Chemiluminescence detection using regenerable tris(2,2'-bipyridyl)ruthenium(II) immobilized in Nafion.

The development of a detection method based on the electrogenerated chemiluminescence of tris(2,2'-bipyridine)ruthenium(II), (Ru(bpy)3(2+], immobilized in a Nafion film coated on an electrode is discussed. Control of the electrode potential controls creation of the reactive reagent Ru(bpy)3(3+) which reacts with certain analytes to yield chemiluminescence emission of intensity proportional to the analyte concentration. The reaction results in Ru(bpy)3(3+) being converted to Ru(bpy)3(2+), which then is recycled to Ru(bpy)3(3+) again at the electrode. This sensor has been used in flow injection to determine oxalate, alkylamines, and NADH. Detection limits are 1 microM, 10 nM, and 1 microM, respectively, with working ranges extending over 4 decades in concentration. Sensitivity is constant over the wide pH range from 3 to 10. With oxalate, and to a small extent with amines, emission intensities increase with increasing ionic strength; this was shown to be a phenomenon related to the Nafion film and not to the chemiluminescence reaction. Emission intensities increase with temperature. The sensor remains stable for several days with suitable storage conditions. Significant amounts of Ru(bpy)3(3+) are shown to be capable of storage within the film.     

Electrogenerated chemiluminescence. 67. Dependence of light emission of the tris(2,2')bipyridylruthenium(II)/tripropylamine system on electrode surface hydrophobicity

We describe the effect of electrode surface hydrophobicity on the electrochemical behavior and electrogenerated chemiluminescence (ECL) of Ru(bpy)3(2+) (bpy = 2,2'-bipyridyl)/tripropylamine (TPrA) system. Gold and platinum electrodes were modified with different thiol monolayers. The hydrophobicity of the electrode surfaces changed with different terminal groups of the thiol molecules. The oxidation rate of TPrA was found to be much larger at the modified electrode with a more hydrophobic surface. The adsorption of neutral TPrA species on this kind of surface was assumed to contribute to the faster anodic kinetics. Due to the rapid generation of the highly reducing radical, TPrA., ECL intensity increased significantly at more hydrophobic electrodes. This electrode surface effect in the ECL analytical system allows one to improve the detection sensitivity at low concentrations of Ru(bpy)3(2+). The surfactant effect on the ECL process was also examined and discussed based on the change of electrode hydrophobicity by the adsorption of surfactant species.     

Multilabeling biomolecules at a single site. 1. Synthesis and characterization of a dendritic label for electrochemiluminescence assays

Ultrasensitive bioanalytical assays are of great value for early detection of human diseases and pathogens. The sensitivities of immunoassays and DNA probing can be enhanced by multilabeling the biorecognition partner used for affinity-based assays. However, the bioreactivity of biomolecules is affected by a high degree of multilabeling at multiple functional sites. It is proposed that dendritic scaffoldings be used to link multiple signal-generating units to a single site with potentially minimum impact on the bioaffinity. A prototype label, a zeroth-generation dendron, bearing three [Ru(bpy)(3)](2+) units for electrochemiluminescence (ECL) assays was synthesized and characterized preliminarily by spectroscopic, electrochemical, and ECL methods. No evidence of interaction between the neighboring [Ru(bpy)(3)](2+) units in the label molecule was found from these characterizations. Both the photoluminescence and ECL of the prototype label have features very similar to those of mononuclear [Ru(bpy)(3)](2+) compounds. Labeling a model protein with a triad of [Ru(bpy)(3)](2+) at one NH(2) position was demonstrated. The results reported here provide support to applying the proposed multilabeling strategy to affinity-based bioanalytical assays.     

Carbon nanotube microwell array for sensitive electrochemiluminescent detection of cancer biomarker proteins

This paper describes fabrication of a novel electrochemiluminescence (ECL) immunosensor array featuring capture-antibody-decorated single-wall carbon nanotube (SWCNT) forests residing in the bottoms of 10-μL wells with hydrophobic polymer walls. Silica nanoparticles containing [Ru(bpy)(3)](2+) and secondary antibodies (RuBPY-silica-Ab(2)) are employed in this system for highly sensitive two-analyte detection. Antibodies to prostate specific antigen (PSA) and interleukin-6 (IL-6) were attached to the same RuBPY-silica-Ab(2) particle. The array was fabricated by forming the wells on a conductive pyrolytic graphite chip (1 in. × 1 in.) with a single connection to a potentiostat to achieve ECL. The sandwich immunoassay protocol employs antibodies attached to SWCNTs in the wells to capture analyte proteins. Then RuBPY-silica-Ab(2) is added to bind to the captured proteins. ECL is initiated in the microwells by electrochemical oxidation of tripropyl amine (TprA), which generates excited state [Ru(bpy)(3)](2+) in the 100-nm particles, and is measured with a charge-coupled device (CCD) camera. Separation of the analytical spots by the hydrophobic wall barriers enabled simultaneous immunoassays for two proteins in a single sample without cross-contamination. The detection limit (DL) for PSA was 1 pg mL(-1) and for IL-6 was 0.25 pg mL(-1) (IL-6) in serum. Array determinations of PSA and IL-6 in patient serum were well-correlated with single-protein ELISAs. These microwell SWCNT immunoarrays provide a simple, sensitive approach to the detection of two or more proteins.     

Green electrochemiluminescence from ortho-metalated tris(2-phenylpyridine)iridium(III)

The electrochemiluminescence (ECL) of Ir(ppy)3 (ppy = 2-phenylpyridine) is reported in acetonitrile (CH3CN), mixed CH3CN/H20 (50:50 v/v), and aqueous (0.1 M KH2PO4) solutions with tri-n-propylamine as an oxidative-reductive coreactant. ECL efficiencies (phi(ecl), photons emitted per redox event) of 0.00092 in aqueous, 0.0044 in mixed, and 0.33 in CH3CN solutions for Ir(ppy)3 were obtained using Ru(bpy)3(2+) (bpy = 2,2'-bipyridine) as a relative standard (phi(ecl) = 1). Photoluminescence (PL) efficiencies of 0.039, 0.050, and 0.069 were obtained in aqueous, mixed, and acetonitrile solutions, respectively, compared to Ru(bpy)3(2+) (phi(em) = 0.042). The ECL spectra were identical to photoluminescence spectra (lambda(max) approximately equal to 517 nm), indicating formation of the same metal-to-ligand (MLCT) excited states in both ECL and PL. The ECL is linear over several orders of magnitude in mixed and acetonitrile solution with theoretical detection limits (blank plus three times the standard deviation of the noise) of 1.23 nM in CH3CN and 0.23 microM in CH3CN/ H20 (50:50 v/v).