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Obtaining stable electrochemiluminescence (ECL) emissions from a hydrophobic luminophore in aqueous solutions and designing a method without the use of an exogenous coreactant are promising for ECL biosensing. Here, a highly sensitive signal-on ECL immunoassay for the SARS-CoV-2 N protein was developed using micelles as an ECL tag. The micelles were prepared by coencapsulating the luminophore hydrophobic CdSe/ZnS quantum dots and coreactant dibenzoyl peroxide within the hydrophobic core of micelles. The ECL probe was obtained by covalently bonding a SARS-CoV-2 N protein-binding aptamer onto the micelle surface. The construction of the immunosensor was initiated by the immobilization of the anti-SARS-CoV-2 N protein antibody onto the screen-printed carbon electrode (SPCE) with a -COOH-functionalized surface. The surface functionalization of SPCEs was achieved through paste-exfoliated graphene, which was modified with a -COOH group through supramolecular-covalent scaffolds on SPCE. Upon achieving sandwich complexes on the immunosensor, an efficient ECL signal response at -1.4 V versus Ag/AgCl was obtained in phosphate buffer solution. The ECL assay was used for the sensitive determination of SARS-CoV-2 N protein with the linear range from 0.01 to 50 ng mL -1 , and the detection limit was 3.0 pg mL -1 . The immunosensor showed good reproducibility and stability, and the ECL immunoassay was used to determine the SARS-CoV-2 N protein in serum samples. The proposed approach to obtain micelles is versatile for the preparation of stable ECL luminophores by using hydrophobic materials, and the strategy provides an alternative for ECL bioassays based on the coreactant route.

3-Methylhistidine (3-MeHis) is increasingly used as an indicator of muscle protein breakdown. The development of a sensitive, simple, and non-invasive method for 3-MeHis assay is important in clinical practice. Herein, a sensitive, simple, and non-invasive electrogenerated chemiluminescence (ECL) method was proposed for the quantitation of 3-MeHis in urine by using an iridium(III) solvent complex ([Ir(dfppy) 2 (DMSO)Cl], dfppy = 2-(2,4-difluorophenyl)pyridine, Ir-DMSO) as a signal reagent. The photoluminescence (PL) and ECL responses of Ir-DMSO to 3-MeHis were studied. The ECL intensity of Ir-DMSO was enhanced in the presence of 3-MeHis because of the coordination recognition between Ir-DMSO and the imidazole group of 3-MeHis. Based on the enhancement of ECL intensity, 3-MeHis can be sensitively detected in the range of 5 to 25 μM. The detection limit was 0.4 μM. This is the first report of an ECL method for the quantitation of 3-MeHis. Further, to investigate the feasibility of the Ir-DMSO-based ECL method in practical applications, the developed ECL method was applied for 3-MeHis assay in urine samples of 28 healthy volunteers and 2 patients. The urine samples from patients hospitalized with obesity and kidney disease and healthy individuals were distinguished by the ECL responses of Ir-DMSO. The proposed ECL method based on the coordination recognition between iridium(III) solvent complex and the imidazole group of 3-MeHis allows inexpensive, fast, non-invasive, and sensitive detection of 3-MeHis in urine, which is promising for assessing large volumes of patients for routine analysis in clinical practices., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH, DE part of Springer Nature.)

Exosomes, as an emerging biomarker, have exhibited remarkable promise in early cancer diagnosis. Here, a highly sensitive, selective, and automatic electrochemiluminescence (ECL) method for the detection of cancerous exosomes was developed. Specific aptamer-(EK) 4 peptide-tagged magnetic beads (MBs-(EK) 4 -aptamer) were designed as a magnetic capture probe in which the (EK) 4 peptide was used to reduce the steric binding hindrance of cancerous exosomes with a specific aptamer. One new universal ECL signal nanoprobe (CD9 Ab-PEG@SiO 2 ϵRu(bpy) 3 2+ ) was designed and synthesized by using microporous SiO 2 nanoparticles as the carrier for loading ECL reagent Ru(bpy) 3 2+ , polyethylene glycol (PEG) layer, and anticluster of differentiation 9 antibody (CD9 Ab). A "sandwich" biocomplex was formed on the surface of the magnetic capture probe after mixing the capture probe, target exosomes, and ECL signal nanoprobe, and then it was introduced into an automated ECL analyzer for rapid and automatic ECL measurement. It was found that the designed signal nanoprobe shows a 270-fold improvement in the signal-to-noise ratio than that of the ruthenium complex-labeled CD9 antibody signal probe. The relative ECL intensity was proportional to MCF-7 exosomes as a model in the range of 10 2 to 10 4 particle/μL, with a detection limit of 11 particle/μL. Furthermore, the ECL method was employed to discriminate cancerous exosomes based on fingerprint responses using the designed multiple magnetic capture probes and the universal ECL signal nanoprobe. This work demonstrates that the utilization of a designed automated ECL tactic using the MBs-(EK) 4 -aptamer capture probe and the CD9 Ab-PEG@SiO 2 ϵRu(bpy) 3 2+ signal nanoprobe will provide a unique and robust method for the detection and discrimination of cancerous exosomes.

A highly sensitive and selective electrogenerated chemiluminescence (ECL) biosensor was developed for the determination of matrix metalloproteinase 3 (MMP-3) in serum via the target-induced cleavage of an oligopeptide. One ECL probe (named as Ir-peptide) was synthesized by covalently linking a new cyclometalated iridium(III) complex ([(3-pba) 2 Ir(bpy-COOH)](PF 6 )) (3-pba = 3-(2-pyridyl) benzaldehyde, bpy-COOH = 4'-methyl-2,2'-bipyridine-4-carboxylic acid) with an oligopeptide (CGVPLSLTMGKGGK). An ECL biosensor was fabricated by firstly casting Nafion and gold nanoparticles (AuNPs) on a glassy carbon electrode and then self-assembling both of the ECL probes, 6-mercapto-1-hexanol and zwitterionic peptide, on the electrode surface, from which the AuNPs could be used to amplify the ECL signal and Ir-peptide could serve as an ECL probe to detect the MMP-3. Thanks to the MMP-3-induced cleavage of the oligopeptide contributing to the decrease in ECL intensity and the amplification of the ECL signal using AuNPs, the ECL biosensor could selectively and sensitively quantify MMP-3 in the concentration range of 10-150 ng·mL -1 and with both a limit of quantification (26.7 ng·mL -1 ) and a limit of detection (8.0 ng·mL -1 ) via one-step recognition. In addition, the developed ECL biosensor showed good performance in the quantization of MMP-3 in serum samples, with a recovery of 92.6% ± 2.8%-105.6% ± 5.0%. An increased level of MMP-3 was found in the serum of rheumatoid arthritis patients compared with that of healthy people. This work provides a sensitive and selective biosensing method for the detection of MMP-3 in human serum, which is promising in the identification of patients with rheumatoid arthritis.

This Feature Article simply introduces principles and mechanisms of electrochemiluminescence (ECL) biosensors for the determination of biomarkers and highlights recent advances of ECL biosensors on key aspects including new ECL reagents and materials, new biological recognition elements, and emerging construction biointerfacial strategies with illustrative examples and a critical eye on pitfalls and discusses challenges and perspectives of ECL biosensors for health analysis.

This work reports a gold nanoelectrode ensembles (Au-NEE) platform taken as a disposable electrogenerated chemiluminescence (ECL) platform with immunomagnetic microbeads for ECL immunoassays for the first time. The peak-shaped voltammograms were obtained at the Au-NEE, attributed to the total diffusional overlap. The ECL intensity at Au-NEE was 12.9 folds in the Ru(bpy) 3 2+ -tri- n -propylamine (TPA) ECL system and 19.6 folds in the luminol-H 2 O 2 system, compared with that at the Au macroelectrode using the normalized active area of the electrodes, mainly attributed to the diffusion overlap of the Au-NEE and the edge effect of the individual gold nanodisks of the Au-NEE. The ECL immunoassay on the Au-NEE platform with magnetic microbeads for the determination of cancer biomarkers was developed. Carbohydrate antigen 19-9 (CA 19-9) was chosen as a model analyte while CA 19-9 antibody on the magnetic microbeads was taken as the capture probe, and ruthenium complex-labeled CA 19-9 antibody was used as the signal probe. A "sandwich" bioconjugates on the magnetic beads were transferred onto the ECL platform, and then the ECL measurements were performed in TPA solution. The developed method showed that the ECL peak intensity was directly in proportion to the concentration of CA 19-9 in the range from 0.5 to 20 U/mL with a limit of detection of 0.4 U/mL. This work demonstrates that the Au-NEE can be employed as a useful disposable ECL platform with the merits of cheapness, low nonspecific adsorption and practical application. The proposed approach will open a new avenue in the point-of-care test for the determination of protein biomarkers.

In this work, a simple electrochemiluminescence (ECL) imaging method based on the cell shield of the ECL emission was developed for the morphological and quantitative analysis of living cells under external stimulation. ECL images of MCF-7 cells cultured on or captured at the glassy carbon electrode (GCE) surface in a solution of tris(2,2'-bipyridyl)ruthenium(II)-tri- n -propylamine were recorded. Important morphological characteristics of living cells, including cell shape, cell area, average cell boundary, and junction distance between two adjacent cells, were directly obtained using the developed negative ECL imaging method. The ECL images revealed gradual morphological changes in cells on the GCE surface. During the course of H 2 O 2 stimulation of cells on the GCE surface, cells shrunk, rounded up, disengaged from surrounding cells, and finally detached from the electrode surface. During the course of electrical stimulation (0.8 V), the cells on the GCE surface exhibited aggregation as demonstrated by increases in the average cell boundary and decreases in the junction distance between two adjacent cells. Additionally, a quantitative method for the sensitive determination of MCF-7 cells with a limit of detection of 29 cells/mL was developed using the negative ECL imaging strategy. This work demonstrates that the proposed negative ECL imaging strategy is a promising approach to assess important morphological characteristics of living cells during the course of external stimulation and to obtain quantitative information on cell concentrations in solution.

This Feature simply introduces the history and mechanism of classical electrogenerated chemiluminescence (ECL) systems for the detection of biomolecules, highlights new advances and emerging fields of the ECL biosensing with recent illustrative examples, and presents the challenges and perspectives of ECL biosensing.

A simple electrochemiluminescence (ECL) immunoassay based on a proximity hybridization-regulated strategy was developed for highly sensitive and specific detection of cell surface protein and protein-overexpressing cancer cells. A biosensor was fabricated by self-assembling a thiolated capture ss-DNA3 (partially hybridize with ss-DNA1 and ss-DNA2) and blocking with 6-mercapto-1-hexanol on a gold electrode surface. Target protein was simultaneously bound by two ss-DNA-tagged antibody probes (DNA1-Ab1 and DNA2-Ab2), while DNA1 and DNA2 were brought in sufficient proximity and hybridized with capture DNA3 on the surface of the biosensor. After ECL signal reagent Ru(phen) 3 2+ was intercalated into the hybridized ds-DNAs, ECL measurement was performed in the coreactant solution. A "signal on" proximity hybridization-regulated ECL immunoassay for alpha-fetoprotein (AFP) was developed. The ECL intensity increased with the increase of AFP concentration in the range of 0.05-20.0 ng/mL with a detection limit of 6.2 pg/mL. Moreover, the developed ECL method was successfully used to detect AFP-overexpressing cancer cells (MCF-7 cancer cells as model) with a detection limit of 620 cells/mL (∼60 MCF-7 cells in 100 μL of cell suspension) and discriminate AFP expression on different types of the living cell surface. This work for the first time reports a proximity hybridization-regulated ECL immunoassay for the detection of the cell surface protein on a living cell surface with good specificity and sensitivity. This simple, specific, and sensitive strategy is greatly promising for the detection of proteins and specific cells.

A simple and sensitive electrogenerated chemiluminescence biosensor was developed to monitor matrix metalloproteinase 2 (MMP-2) by employing a specific peptide (CGPLGVRGK) as a molecular recognition substrate. Bis(2,2'-bipyridine)-4'-methyl-4-carboxybipyridine-ruthenium N-succinimidyl ester-bis(hexafluorophosphate) (Ru(bpy)2(mcbpy-O-Su-ester)(PF6)2 (Ru1) was used as ECL-emitting species and covalently labeled onto the peptide through NH2-containing lysine on the peptide via acylation reaction to form Ru1-peptide as an ECL probe. An ECL peptide-based biosensor was fabricated by self-assembling the ECL probe onto the surface of gold electrode. MMP-2 can specifically cleave the Ru1-peptide on the electrode surface, which led the partly Ru1-peptide to leave the electrode surface and resulted in the decrease of the ECL intensity obtained from the resulted electrode in 0.1 M phosphate-buffered saline (pH 7.4) containing tri-n-propylamine. The decreased ECL intensity was piecewise linear to the concentration of MMP-2 in the range from 1 to 500 ng/mL. Moreover, the ECL biosensor is successfully applied to detection of MMP-2 secreted by living cell, such as HeLa cells. Additionally, the biosensor was also applied to the evaluation of matrix metalloproteinase inhibitors. The strategy presented here is promising for other disease-related matrix metalloproteinase assay and matrix metalloproteinase inhibitor profiling with sensitivity and simplicity. Graphical Abstract Detection of MMP-2 released from living cells by ECL peptide-based biosensor.

A sensitive electrogenerated chemiluminescence (ECL) bioassay was developed for the detection of two protein kinases incorporating the peptide phosphorylation and a versatile ECL probe. Cyclic adenosine monophosphate-dependent protein kinase (PKA) and casein kinase II (CK2) were used as proof-of-concept targets while a PKA-specific peptide (CLRRASLG) and a CK2-specific peptide (CRRRADDSDDDDD) were used as the recognition substrates. Taking advantage of the ability of protein A binding with the Fc region of a variety of antibodies with high affinity, a ruthenium derivative-labeled protein A was utilized as a versatile ECL probe for bioassay of multiple protein kinases. A specific peptide substrate toward target protein kinase was first self-assembled on the surface of gold electrode and then serine in the specific peptide on the electrode was phosphorylated by target protein kinase in the presence of adenosine-5'-triphosphate. After recognition of the phosphorylated peptide by monoclonal antiphosphoserine antibody, the versatile ECL probe was specifically bound to the antiphosphoserine antibody on the electrode surface. The ECL bioassay was developed successfully in the individual detection of PKA and CK2 with detection limit of 0.005 U/mL and 0.004 U/mL, respectively. In addition, the ECL bioassay was applied to quantitative analysis of the kinase inhibitors and monitoring drug-triggered kinase activation in cell lysates. Moreover, an ECL imaging bioassay using electron-multiplying charged coupled device as detector on the gold electrode array was developed for the simultaneous detection of PKA and CK2 activity from 0.01 U/mL to 0.4 U/mL, respectively, at one time. This work demonstrates that the ingenious design and use of a versatile ECL probe are promising to simultaneous detection of multiple protein kinases and screening of kinase inhibitor.

A multi-functional eletrogenerated chemiluminescence (ECL) imaging analyzer including both a photomultiplier tube and charged coupled device as detectors has been developed. The ECL imaging analyzer can effectively work for electrochemical study, ECL intensity detection at electrode array, and ECL imaging at bipolar electrodes or electrode array. As an ECL imaging example, an ECL biosensor for visual detection of matrix metalloproteinase 7 in the range from 0.05 to 1 ng/mL is demonstrated.

A novel electrogenerated chemiluminescence (ECL) aptasensor for ultrasensitive detection of thrombin incorporating an auxiliary probe was designed by employing specific anti-thrombin aptamer as a capture probe and a ruthenium(II) complex-tagged reporter probe as an ECL probe and an auxiliary probe to assist the ECL probe close to the surface of the electrode. The ECL aptasensor was fabricated by self-assembling a thiolated capture probe on the surface of gold electrode and then hybridizing the ECL probe with the capture probe, and further self-assembling the auxiliary probe. When analyte thrombin was bound with the capture probe, the part of the dehybridized ECL probe was hybridized with the neighboring auxiliary probe, led to the tagged ruthenium(II) complex close to the electrode surface, resulted in great increase in the ECL intensity. The results showed that the increased ECL intensity was directly related to the logarithm of thrombin concentrations in the range from 5.0 × 10(-15)M to 5.0 × 10(-12)M with a detection limit of 2.0 × 10(-15)M. This work demonstrates that employing an auxiliary probe which exists nearby the capture probe can enhance the sensitivity of the ECL aptasensor. This promising strategy will be extended to the design of other biosensors for detection of other proteins and genes., (Copyright © 2014 Elsevier B.V. All rights reserved.)

A novel electrogenerated chemiluminescence peptide-based biosensor (ECL-PB) for the determination of prostate-specific antigen (PSA) was developed on the basis of target-induced cleavage of a specific peptide within Nafion film incorporated with gold nanoparticles (AuNPs) and ECL emitting species. A specific peptide (CHSSKLQK) was used as a molecular recognition element; tris(2,2'-ripyridine) dichlororuthenium(II) (Ru(bpy)3(2+)) was used as ECL emitting species, and ferrocene carboxylic acid (Fc) was employed as ECL quencher. The ECL-PB biosensor was fabricated by casting the mixture of Nafion and AuNPs onto the surface of glassy carbon electrode to form AuNPs/Nafion film, and then, Ru(bpy)3(2+) was electrostatically adsorbed into the AuNPs/Nafion film; finally, the peptide-tagged with ferrocene carboxylic acid (Fc-peptide) was self-assembled onto the surface of the AuNPs. When PSA was present, it specifically cleaved the Fc-peptide, leading the quencher to leave the electrode and resulting in the increase of the ECL intensity obtained from the resulted electrode in 0.1 M phosphate buffer saline (pH 7.4) containing tri-n-propylamine. The results showed that the increased ECL intensity was directly linear to the logarithm of the concentration of PSA in the range from 5.0 × 10(-12) to 5.0 × 10(-9) g/mL. An extremely low detection limit of 8 × 10(-13) g/mL was achieved because of the signal amplification through AuNPs and the ECL background suppression through Fc as ECL quencher. This work demonstrates that the combination of the direct transduction of peptide cleavage events with the highly sensitive ECL method is a promising strategy for the design of enzymatic cleavage-based ECL biosensors with high sensitivity and selectivity.

A novel electrogenerated chemiluminescence peptide-based (ECL-PB) biosensor for highly sensitive and selective determination of prostate-specific antigen (PSA) was developed. A helix peptide (CHSSKLQK) was served as a molecular recognition element and ruthenium bis(2,2'-bipyridine) (2,2'-bipyridine-4,4'-dicarboxylic acid)-N-hydroxysuccinimide ester (Ru(bpy)(2)(dcbpy)NHS) was used as an ECL label. The helix peptide was labeled with the ECL label at NH(2)-containing lysine and utilized as ECL probe (Ru-peptide). The ECL-PB biosensor was fabricated by immobilizing the ECL probe onto a gold electrode surface via self-assembling technique through a thiol-containing cysteine at the end of the peptide. The principle of ECL measurement is based on the specific proteolytic cleavage event of Ru-peptide on the gold electrode surface in the presence of PSA, resulting in a decrease of ECL signal. The decreased ECL intensity was directly linear to the concentration of PSA in the range from 1.0×10(-10) g/mL to 8.0×10(-9) g/mL with a detection limit of 3.8×10(-11) g/mL. This work demonstrates that the direct transduction of peptide cleavage events into an ECL signal provides a simple and sensitive method for detecting target protein., (Copyright © 2012 Elsevier B.V. All rights reserved.)

A novel label-free electrogenerated chemiluminescence (ECL) aptasensor for the determination of lysozyme is designed employing lysozyme binding aptamer (LBA) as molecular recognition element for lysozyme as a model analyte and Ru(bpy)(3)(2+) as an ECL signal compound. This ECL aptasensor was fabricated by self-assembling the thiolated LBA onto the surface of a gold electrode. Using this aptasensor, sensitive quantitative detection of lysozyme is realized on basis of the competition of lysozyme with Ru(bpy)(3)(2+) cation for the binding sites of LBA. In the presence of lysozyme, the aptamer sequence prefers to form the LBA-lysozyme complex, the less negative environment allows Ru(bpy)(3)(2+) cations to be less bound electrostatically to the LBAs on the electrode surface, in conjunction with the generation of a decreased ECL signal. The integrated ECL intensity versus the concentration of lysozyme was linear in the range from 6.4×10(-10) M to 6.4×10(-7) M. The detection limit was 1.2×10(-10) M. This work demonstrates that using the competition of target protein with an ECL signal compound Ru(bpy)(3)(2+) for binding sites of special aptamer confined on the electrode is promising approach for the design of label-free ECL aptasensors for the determination of proteins., (Copyright © 2010 Elsevier B.V. All rights reserved.)

A highly reproducible and sensitive signal-on electrogenerated chemiluminescence (ECL) biosensor based on the DNAzyme for the determination of lead ion was developed. The ECL biosensor was fabricated by covalently coupling 5'-amino-DNAzyme-tagged with ruthenium bis (2,2'-bipyridine) (2,2'-bipyridine-4,4'-dicarboxylic acid)-ethylenediamine (Ru1-17E') onto the surface of graphite electrode modified with 4-aminobenzoic acid, and then a DNA substrate with a ribonucleotide adenosine hybridized with Ru1-17E' on the electrode. Upon binding of Pb(2+) to the Ru1-17E' to form a complex which catalyzed the cleavage of the DNA substrate, the double-stranded DNA was dissociated and thus led to a high ECL signal. The signal linearly increases with the concentration of Pb(2+) in the range from 5.0 to 80 pM with a detection limit of 1.4 pM and a relative standard derivation of 2.3%. This work demonstrates that using DNAzyme tagged with ruthenium complex as an ECL probe and covalently coupling method for the fabrication of the ECL biosensor with high sensitivity, good stability and significant regeneration ability is promising approach., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Two electrogenerated chemiluminescence (ECL) aptasensors for the detection of thrombin were developed using the thrombin binding aptamer (TBA) taken as a molecular recognition element and nanomaterial as a carrier of the ECL capture/signal probe. In the "signal off" aptasensor, the thiolated capture probe (ss-DNA, 12-mer) was self-assembled on the gold nanoparticles (GNPs) which were self-assembled on the surface of gold electrode, and hybridized with six-base segment of the ss-DNA sequence (Tgt-aptamer, 21-mer) containing TBA-I (ss-DNA, 15-mer) tagged with ruthenium complex, producing a high ECL intensity. Introduction of the analyte thrombin triggered the dissociation of the Tgt-aptamer tagged with ruthenium complex from the aptasensors, led to significantly decrease in ECL intensity. The decreased ECL intensity was in proportion to the concentration of thrombin in a range from 2.7×10(-12) to 2.7×10(-9) M with a detection limit of 8×10(-13) M. In the "signal on" aptasensor, the thiolated TBA-I was self-assembled on the gold electrode for capturing thrombin onto the electrode and then the TBA-II (ss-DNA, 29-mer) labeled with single-walled carbon-nanotubes (SWNT)-ECL tag was bound with epitope of thrombin, producing a high ECL intensity. The increased ECL intensity was linearly with the concentration of thrombin from 1.0×10(-14) M to 1.0×10(-11) M with a detection limit of 3×10(-15) M. The present work demonstrates that using nanomaterial as a carrier for capture probe and signal probe is a promising way to amplify the ECL signal and to improve the sensitivity of the aptasensors., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Electrogenerated chemiluminescence (ECL) of a ruthenium complex polymer modified carbon paste electrode and its analytical applications were investigated. The ruthenium complex polymer was prepared using bis(2,2-bipyridine) (4,4-dicarboxy-2,2-bipyridine) ruthenium(II). The ECL behaviours of ruthenium complex polymer modified carbon paste electrode were investigated in the absence and presence of tripropylamine (TPA). The modified carbon paste electrode exhibited long-term stability and fine reproducibility. The ECL intensity of the modified carbon paste electrode was linear with the concentration of TPA in the range 2.0 x 10(-6)-3.8 x 10(-3) mol/L, with a detection limit (S:N = 3) of 6 x 10(-7) mol/L. It was also found that raceanisodamine could enhance the ECL intensity of the modified electrode. The ECL intensity of the modified carbon paste electrode was linear with the concentration of raceanisodamine in the range 1.1 x 10(-5)-6.0 x 10(-4) mol/L, with a detection limit (S:N = 3) of 6 x 10(-6) mol/L. This work demonstrates that the entrapment of ruthenium complex in a highly cross-linked polymer is a promising approach to construct an ECL modified electrode with long-term stability and fine reproducibility. The modified electrode designed has a potential application in the ECL detector., ((c) 2008 John Wiley & Sons, Ltd.)

A highly selective electrogenerated chemiluminescence (ECL) biosensor for the detection of target single-strand DNA (ss-DNA) was developed using hairpin DNA as the recognition element and ruthenium complex as the signal-producing compound. The ECL-based DNA biosensor was fabricated by self-assembling the ECL probe of thiolated hairpin DNA tagged with ruthenium complex on the surface of a gold electrode. In the absence of target ss-DNA, the ECL probe immobilized on the surface of the electrode was in the folded configuration in which its termini were held in close proximity to the electrode, and thus a strong ECL signal could be generated. In the presence of target ss-DNA, a stem-loop of the ECL probe on the electrode was converted into a linear double-helix configuration due to hybridization, resulting in the tag moving away from the electrode surface, which in turn decreased the ECL signal. The ECL intensity of the DNA biosensor generated a "switch off" mode, which decreased with an increase of the concentration of target DNA, and a detection limit of 9 x 10(-11) M complementary target ss-DNA was achieved. Single mismatched target ss-DNA was effectively discriminated from complementary target ss-DNA. The effect of different loop lengths of the hairpin DNA on the selectivity of the ECL DNA biosensor has been investigated. This work demonstrated that the sensitivity and specificity of an ECL DNA biosensor could be greatly improved using a hairpin DNA species which has an appropriate stem and loop length as the recognition element.

A highly sensitive homogeneous electrogenerated chemiluminescence (ECL) immunoassay for the determination of anti-digoxin antibody and digoxin hapten was developed employing Ru(bpy)(2)(dcbpy)NHS (bpy = 2,2'-bipyridyl; dcbpy = 2,2'-bipyridine-4,4'-dicarboxylic acid; NHS = N-hydroxysuccinimide ester) as an electrochemiluminescent label and bovine serum albumin (BSA) as a carrier protein. A digoxin hapten was indirectly heavily labelled with Ru(bpy)(2)(dcbpy)NHS through BSA to form Ru(bpy)(2)(dcbpy)NHS-BSA-digoxin conjugate. The ECL intensity of the immunocomplex of the conjugate with anti-digoxin antibody markedly decreased when the immunoreaction between Ru(bpy)(2)(dcbpy)NHS-BSA-digoxin conjugate and anti-digoxin antibody took place. Two formats, direct homogeneous immunoassay for anti-digoxin antibody and competitive immunoassay for digoxin, were developed to determine anti-digoxin antibody and digoxin, respectively. The anti-digoxin antibody concentration in the range 7.6 x 10(-8)-7.6 x 10(-6) g/mL was determined by direct homogeneous format. Digoxin hapten was determined throughout the range 4.0 x 10(-10)-1.0 x 10(-7) g/mL with a detection limit of 1.0 x 10(-10) g/mL by competitive format. The relative standard derivation for 6.0 x 10(-9) g/mL was 4.3%. The method has been applied to assaying digoxin in control human serum., (Copyright (c) 2006 John Wiley & Sons, Ltd.)

The electrogenerated chemiluminescence (ECL) reaction of lucigenin with isatin was investigated at a platinum electrode in a neutral aqueous solution. The ECL intensity of lucigenin at -0.65 V was greatly enhanced by isatin, and the ECL intensity was about 50 times higher than that of lucigenin without isatin. The enhanced ECL was believed to be produced by the chemiluminescence reaction between reduced lucigenin and superoxide anion that was generated by the reaction of electrochemically reduced isatin with dissolved oxygen. The conditions for the determination of isatin were optimized. Under the optimized condition, the enhanced ECL intensity vs. isatin concentration was linear in the range 4.8 x 10(-7)-1.9 x 10(-5) g/mL; with a detection limit of 3.3 x 10(-8) g/mL, and the relative standard derivation 1.0 x 10(-6) g/mL isatin was 3.8%., (Copyright 2004 John Wiley & Sons, Ltd.)