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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)2Ir(bpy-COOH)](PF6)) (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. [ABSTRACT FROM AUTHOR]

A sensitive electrogenerated chemiluminescence peptide-based (ECL-PB) biosensing method for the determination of protein was developed by employing peptideintegrating Ru[(bpy).sub.3.sup.2+](bpy=2,2'-bipyridine)-functionalized gold nanoparticles (Ru[(bpy).sub.3.sup.2+]-AuNPs-peptide) as nanoprobe. Cardiac troponin I (cTnI), a reliable clinical biomarker for the detection of cardiac injury, was chosen as target protein, while a specific binding peptide (CFYSHSFHENWPS) was used as molecular recognition element. AuNPs were firstly functionalized with Ru[(bpy).sub.3.sup.2+] through electrostatic interactions between citrate-capped AuNPs and Ru[(bpy).sub.3.sup.2+] to form Ru[(bpy).sub.3.sup.2+]-AuNPs aggregates and then functionalized with peptide through Au-S bounds to form Ru[(bpy).sub.3.sup.2+]-AuNPs-peptide nanoprobe. AuNPs not only can capture numerous signalgenerating molecules, resulting in high ECL intensity but also can capture a significant amount of the peptide, providing poly binding motif. The specific capture peptide was self-assembled on the surface of a gold electrode and then incubated with the target cTnI and Ru[(bpy).sub.3.sup.2+]-AuNPs-peptide successively. A sandwich-type peptide/cTnI/Ru[(bpy).sub.3.sup.2+]-AuNPs-peptide conjugate was formed on the surface of the electrode and an ECL signal was obtained in the presence oftri-n-propylamine. The novel biosensing method facilitates the sensitive detection of cTnI in the range from 3.0 x [10.sup.-12] g m[L.sup.-1] to 7.0x [10.sup.-11] g m[L.sup.-1] with a low detection limit of 0.5 pg m[L.sup.-1]. This work provides a promising strategy for the determination of proteins with simplicity, high sensitivity, and selectivity. Keywords Electrogenerated chemiluminescence * Biosensing * Peptide * Cardiac troponin I * Gold nanoparticles, Introduction Cardiac troponin I (cTnI), a part of the troponin complex present in cardiac muscle tissues, is a reliable biomarker of cardiac muscle tissue injury and is widely used in [...]

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. [ABSTRACT FROM AUTHOR]

Abstract Electrogenerated chemiluminescence (electrochemiluminescence, ECL) generates species at electrode surfaces, which undergoes electron-transfer reactions and forms excited states to emit light. It has become a very powerful analytical technique and has been widely used in such as clinical testing, biowarfare agent detection, and pharmaceutical analysis. This review focuses on the current trends of molecular recognition-based biosensing methods for pharmaceutical analysis since 2010. It introduces a background of ECL and presents the recent ECL developments in ECL immunoassay (ECLIA), immunosensors, enzyme-based biosensors, aptamer-based biosensors, and molecularly imprinted polymers (MIP)-based sensors. At last, the future perspective for these analytical methods is briefly discussed. [ABSTRACT FROM AUTHOR]

The synthesis and characterization of compounds as electrogenerated chemiluminescence (ECL) reagents are crucial in fundamental research for ECL biosensors. In this work, a new cyclometalated iridium(III) complex, [(bt)2Ir(dmphen)](PF6), was synthesized by using 2-phenylbenzothiazole (bt) as the [ABSTRACT FROM AUTHOR]

A screen-printed carbon nanotube (CNTs) based disposable third generation biosensor for hydrogen peroxide (HO) was constructed by mixing CNTs with a cellulose acetate binder on an epoxy substrate. The surface was covered with a layer consisting of horseradish peroxidase (HRP) crosslinked to bovine serum albumin with glutaraldehyde. The CNTs acted as both the electrode material and charge promoter to facilitate the direct electron transfer between immobilized HRP and the modified electrode. At a working potential of −300 mV (vs. Ag/AgCl), the biosensor displays an excellent electrocatalytic response to the reduction of HO without the aid of a mediator. It has a fast (3 s) amperometric response and a linear calibration in the concentration range 0.005-0.1 mmol L, with a 0.85 μM detection limit (at an S/N of 3). The biosensor can be easily produced, is stable and reproducible. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

We describe a highly sensitive electrochemiluminescence (ECL) based method for the determination of the human immunodeficiency virus-1 (HIV-1) gene. A long-range self-assembled double strand DNA (ds-DNA) is used as a carrier, and the ruthenium complex Ru(phen) as an ECL indicator for signal amplification. The thiolated ss-DNA serving as a capture probe is firstly self-assembled on the surface of a gold electrode. After the target HIV-1 gene is completely hybridized with the capture probe, two previously hybridized auxiliary probes are hybridized with the target HIV-1 gene to form long-range supersandwich ds-DNA polymers on the surface of the electrode. Finally, the ECL indicator is intercalated into the supersandwich ds-DNA grooves. This results in a strongly increased ECL in tripropylamine solution because a large fraction of the intercalator is intercalated into supersandwich ds-DNA. The results showed that the increased ECL intensity is directly related to the logarithm of the concentration of the HIV-1 gene in the range from 0.1 pM to 0.1 nM, with a detection limit of 0.022 pM and using only 10 μL of analyte samples. The method can effectively discriminate target HIV-1 gene (a perfectly matched ss-DNA) from a 2-base mismatched ss-DNA. This work demonstrates that the high sensitivity and selectivity of an ECL DNA biosensor can be largely improved by using supersandwich ds-DNA along with ECL indicators. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

novel third generation biosensor for hydrogen peroxide (HP) was constructed by crosslinking horseradish peroxidase (HRP) onto an electrode modified with multi-wall carbon nanotubes (MWNTs). Glutaraldehyde and bovine serum albumin were used to cross-link HRP and the MWNTs. Scanning electron microscopy revealed that HRP was evenly distributed on the surface of the nanotubes. Factors influencing the performance of the biosensor were studied in detail. The biosensor exhibited a fast amperometric response (2.0 s) to HP which is linear in the concentration range from 9.5 mM to 0.95 μM. The detection of limit is 0.4 μM. The sensor exhibits good stability and durability (on testing 0.1 mM solution of HP over a period of 10 days), with a long-term relative standard deviation of <5%. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

Abstract: An amperometric third-generation hydrogen peroxide biosensor was designed by immobilizing hemoglobin (Hb) on a glassy carbon electrode modified with multiwall carbon nanotubes (MWNT). The direct electron transfer of the Hb immobilized on the MWNT-modified electrode was observed. The formal potential of the immobilized Hb was −0.241V versus Ag/AgCl (3M NaCl) and the heterogeneous electron transfer rate constant was 0.58s−1 in a 0.20M acetate buffer solution (pH 5.4). The immobilized Hb exhibited excellent electrocatalytic activity to reduce hydrogen peroxide in the absence and presence of oxygen, which facilitated designing an amperometric third-generation biosensor for hydrogen peroxide. In the presence of oxygen, the response to hydrogen peroxide of the designed biosensor at a potential of −0.35V was linear in the concentration range from 6.0×10−6 to 6.0×10−3 M, and the detection limit was 1.2×10−6 M. The relative standard deviation was 2.4% for nine successive assays at 1.0×10−5 M hydrogen peroxide. The designed biosensor was applied to the determination of hydrogen peroxide in pharmaceutical injections with high sensitivity and rapid response. [Copyright &y& Elsevier]

An amperometric glucose biosensor based on a coimmobilized electron transfer mediator (osmium-complex) layer, an enzyme (glucose oxidase) layer and a permeable pre-oxidation layer on a glassy carbon electrode was devised. The composition of the pre-oxidation layer was optimized. The permeable cellulose acetate membrane employing lead oxide as an interference-removing agent provided favorable permeability and was insusceptible to interference of ascorbic acid and uric acid. The biosensor exhibited high selectivity and a fast response and allowed a high working potential of +0.55V vs. SCE. The amperometric response to glucose shows a linear relation in the range from 0 to 10 mM with a correlation coefficient of 0.9967. The biosensor was applied to the determination of glucose in human serum. [ABSTRACT FROM AUTHOR]

An electrogenerated chemiluminescence (ECL) biosensor array for the individual and simultaneous detection of multiple biomarkers has been developed at first time on the basis of the aptamer-based biosensor incorporating a versatile signal probe. As principle-of-proof, the acute myocardial infarction (AMI) biomarkers, myoglobin(Myo), cardiac troponin I(cTnI) and cardiac troponin T(cTnT) were chosen as model analytes. The biosensor array was fabricated by self-assembling thiolated specific ssDNA aptamers on the surface of gold electrodes, respectively. After each target analyte was bound to the capture probes and then to each corresponding biotinylated antibody, and finally to the versatile ECL signal probe, ECL signals were recorded using PMT or CCD as a detector. For PMT model, the developed method showed extremely low detection limits for 0.30 ng/mL cTnT, 31 pg/mL Myo, 0.79 pg/mL cTnI. For ECL image model, a biosensor array containing three target biosensors and a control biosensor was found to be highly sensitive, no cross-talking, and accurate towards simultaneous detection of the AMI biomarkers. This work demonstrates that the aptamer-based biosensor array with a versatile signal probe is a promising for ECL simultaneous detection of multi-biomarkers. [ABSTRACT FROM AUTHOR]

Highly selective bioassays are extremely important in biological and chemical sensing. Herein, a selective biosensor incorporating sensitive electrogenerated chemiluminescence (ECL) is reported for simultaneous detection of two matrix metalloproteinases (MMPs) that are related to pathological development and progression of cancer. An Ir complex ((dfppy) 2 Ir(dcbpy)PF 6 , Ir1) and a Ru complex ((Ru(bpy) 2 (mcbpy-O-Su-ester)(PF 6 ) 2 , Ru1) were employed as potential resolved ECL labels. It was found that the Ru1 and Ir1 in the presence of tripropylamine could produce two well-separated strong ECL emissions at +0.9 V and +1.4 V vs Ag/AgCl, respectively, with a large difference of ECL peak potential (∼0.5 V) at the gold electrode, which provided an access for potential-resolved detection of dual targets. A multiplexed biosensor was simply fabricated by co-immobilizing the Ir1 and Ru1 labeled peptides on a gold electrode via self-assembling technique. It was facile to determine MMP-2 and MMP-7 in a single run with detection limit as low as 5 ng/mL and 10 pg/mL, respectively. The advantages of the multiplexed ECL biosensor were further exemplified by analyzing two MMPs released from living HeLa, K562, HCT116 and macrophage cells. The multiplexity of the biosensor will open a new door for facile and rapid detection of multiple biomarkers. [ABSTRACT FROM AUTHOR]

This work reports the performance enhancement strategies on magnetic beads (MBs)-based electrochemiluminescence (ECL) platforms by using double magnetic field actuation of the ECL magnetic microbiosensors (MMbiosensors) for highly sensitive determination of cancer biomarker and exosomes. To obtain the high sensitivity and reproducibility of the ECL MMbiosensors, a series of strategies have been developed including replacing a conventional photomultiplier tube (PMT) with a diamagnetic PMT, replacing the stacked ring-disc magnets with circular-disc magnets lain-in glassy carbon electrode, adding a pre-concentration process of MBs using external magnet actuation. For fundamental research, the ECL MBs taken as the substitute of ECL MMbiosensors were prepared by binding biotinylated DNA tagged with Ru(bpy) 3 2+ derivative (Ru1) to streptavidin-coated MB(MB@SA) were which showed that the developed strategies can enhance 45-fold sensitivity. Importantly, the developed MBs-based ECL platform was estimated by determination of prostate specific antigen (PSA) and exosomes. For PSA, MB@SA•biotin-Ab1(PSA) was taken as the capture probe and Ru1-labeled Ab2 (PSA) was done as ECL probe, while for exosomes, MB@SA•biotin-aptamer (CD63) was taken as the capture probe and Ru1-labeled Ab (CD9) was done as the ECL probe. The experiment results showed that the developed strategies can enhance 33-fold sensitivity of ECL MMbiosensors for PSA and exosomes. The detection limit is 0.28 ng mL−1 for PSA and 4.9 × 102 particle mL-1 for exosomes. This work demonstrated that a series of proposed magnetic field actuation strategies greatly increase the sensitivity of the ECL MMbiosensors. The developed strategies can be expanded to MBs-based ECL and electrochemical biosensors for clinical analysis with greater sensitivity. [Display omitted] • Magnetic field greatly decreases photomultiplier tube response to ECL emission. • Circular-disc magnet contributed uniform distribution of magnetic microbeads. • Magnetic actuation of magnetic microbeads enhances the sensitivity. • High performance of ECL biosensor achieved by magnetic actuation. • Highly sensitive MBs-based ECL biosensor was developed for exosomes. [ABSTRACT FROM AUTHOR]

A sensitive electrogenerated chemiluminescence (ECL) aptasensor was developed for the detection of human Burkitt’s lymphoma Ramos cells based on the cells-specific oligonucleotide TD05 served as a molecular recognition element and ruthenium(II) complex (Ru1) labeled polyamidoamine dendrimer (Dend) as a signal probe. Dend was used as platform to assemble substantial Ru1 and TD 05 to form Ru1-Dend-SH-(CH 2 ) 6 -oligo(ethylene oxide) 6 -TD05 bioconjugate which was used as the signal probe. Dend was not only used as platform to assemble substantial Ru1 but also used as a coreactant of Ru1 to amplify the ECL signal. The biosensor platform was fabricated by covalently coupling the 52-mer amino-substituted oligonucleotide probes with carboxyl groups on single wall carbon nanotubes-coated glassy carbon electrode. With the sandwich assay, Ramos cells were selectively captured on the surface of the biosensor based on the specific affinity between aptamers and mIgM on the target cells surface, and then, the signal probe was bound with the captured cells. In the presence of tri- n -propylamine, the increased ECL intensity of the biosensor was logarithmically proportion to the concentration of Ramos cells over a range from 1.0 × 10 2 to 1.0 × 10 4 cells/mL with a detection limit of 55 cells/mL. It was found that the sensitivity of the biosensor increased 4.3-fold by comparison with that without Dend. And also, the ECL biosensor showed satisfied selectivity against the tested control cancer cells. [ABSTRACT FROM AUTHOR]

An electrochemical lectin-based biosensor array employing lectin as molecular recognition elements was developed for sensitive detection and discrimination of carcinoembryonic antigen (CEA) based on dual signal amplification of gold nanoparticles and enzymatic catalysis. The electrochemical biosensor was fabricated by self-assembly of cysteamine on the surface of gold nanoparticle-modified screen-printed carbon electrodes and subsequently covalently coupling lectins on the surface of the cysteamine-modified electrode via the amidation reaction. A lectin–target-antibody sandwich-type conjugate was formed when the biosensor was successively reacted with target protein and horseradish peroxidase labeled anti-CEA antibody probe. In the presence of hydroquinone and hydrogen peroxide, the resulted biosensor produced an obvious catalysis current signal. The catalytic current at −0.25 V was linear with the concentration of CEA in the range of 1 ng/mL to 10 ng/mL with detection limit of 0.03 ng/mL and 0.05 ng/mL using concanavalin A and wheat-germ agglutinin as molecular recognition element, respectively, and in the range of 0.5 ng/mL to 7 ng/mL with detection limit of 0.01 ng/mL using Lens culinaris agglutinin as molecular recognition element. Additionally, the lectin-based biosensor array was successfully applied to evaluate the glycan expression of CEA N -glycan and discriminate CEA between healthy and cancer serum samples. The lectin-based biosensor array endows a feasibility tool for clinical diagnosis in complex biological systems and shows great promise for cancer detection and further pathological study of carcinogenesis. [ABSTRACT FROM AUTHOR]

Two electrogenerated chemiluminescence (ECL) biosensors for selective and sensitive detection of Escherichia coli O157:H7 were first designed by employing the antimicrobial peptide Magainin I as a biological recognition element and the ruthenium complex (Ru1) as an ECL label. In a direct format, the biosensor (I) was fabricated by self-assembling the Ru1-labeled peptide taken as a capture and signal probe onto the gold electrode surface. This system showed that the decreased ECL intensity was logarithmically proportional to the concentration of E. coli O157:H7 in the range of 1.0 × 10 3 –5.0 × 10 5 CFU/mL, with a detection limit of 2.3 × 10 2 CFU/mL. In a sandwich format, the biosensor (II) was fabricated by self-assembling the modified Magainin I taken as capture probe onto the gold electrode surface. After E. coli O157:H7 was captured by the capture probe on the biosensor (II) and then bound with the Ru1-labeled peptide as the signal probe, the increased ECL intensity was logarithmically proportional to the concentration of E. coli from 5.0 × 10 2 CFU/mL to 5.0 × 10 5 CFU/mL, with a detection limit of 1.2 × 10 2 CFU/mL. Importantly, the designed ECL biosensors showed a satisfactory selectivity in discriminating the gram-negative E. coli O157:H7 from gram-positive bacteria, and pathogenic from nonpathogenic E. coli . The strategy of using a Ru1-labeled peptide as a capture/signal probe is a promising approach to designing ECL biosensors for highly sensitive and rapid detection of E. coli O157:H7; this approach could be extended to the design of ECL biosensors for other desired bacteria as well. [ABSTRACT FROM AUTHOR]

A highly selective and sensitive electrogenerated chemiluminescence (ECL) biosensor for the detection of prostate PC-3 cancer cells was designed using a prostate specific antibody as a capture probe and ruthenium complex-labelled wheat germ agglutinin as a signal probe. The ECL biosensor was fabricated by covalently immobilising the capture probe on a graphene oxide-coated glassy carbon electrode. Target PC-3 cells were selectively captured on the surface of the biosensor, and then, the signal probe was bound with the captured PC-3 cells to form a sandwich. In the presence of tripropylamine, the ECL intensity of the sandwich biosensor was logarithmically directly proportion to the concentration of PC-3 cells over a range from 7.0 × 10 2 to 3.0 × 10 4 cells mL −1 , with a detection limit of 2.6 × 10 2 cells mL −1 . The ECL biosensor was also applied to detect prostate specific antigen with a detection limit of 0.1 ng mL −1 . The high selectivity of the biosensor was demonstrated in comparison with that of a lectin-based biosensor. The strategy developed in this study may be a promising approach and could be extended to the design of ECL biosensors for highly sensitive and selective detection of other cancer-related cells or cancer biomarkers using different probes. [ABSTRACT FROM AUTHOR]

Abstract: A label-free electrochemical impedance spectroscopy (EIS) biosensor for the sensitive determination and discrimination of alpha-fetoprotein (AFP) was developed by employing wheat-germ agglutinin (WGA) lectin as molecular recognition element. The EIS biosensor was fabricated by adsorbing carboxyl-functionalized single-wall carbon nanotubes (SWNTs) onto a screen-printed carbon electrode (SPCE) and subsequently covalently coupling WGA onto the surface of the SWNTs-modified electrode. Upon binding of AFP to the biosensor, the electron transfer resistance was increased and the increase in the electron transfer resistance was linearly proportional to the logarithm of the concentration of AFP in the range from 1 to 100ng/L with a detection limit of 0.1ng/L. It was found that the employment of SWNTs as immobilization platform could reduce the background and enhance the EIS response. Moreover, the lectin-based biosensor array fabricated with different lectins was used to evaluate the glycan expression of AFP N-glycan and discriminate AFP between healthy and cancer patients serum samples. This work demonstrates that the employment of carbon nanotubes as immobilization platform and lectin as molecular recognition element in biosensor array is a promising approach for the determination and discrimination of glycoproteins for cancer diagnosis. The strategy proposed in this work could further be used for high-throughput, label-free profiling of the glycan expression of cancer-related glycoproteins and to develop methods for cancer diagnosis in the early stages. [Copyright &y& Elsevier]

Abstract: A highly sensitive and attractive antifouling impedimetric aptasensor for the determination of thrombin in undiluted serum sample was developed. The aptasensor was fabricated by co-assembling thiol-modified anti-thrombin binding aptamer, dithiothreitol and mercaptohexanol on the surface of gold electrode. The performance of aptasensor was characterized by atomic force microscopy, contact angle and electrochemical impedance spectroscopy. In the measurement of thrombin, the change in interfacial electron transfer resistance of aptasensor was monitored using a redox couple of Fe(CN)6 3−/4−. The increase in the electron transfer resistance was linearly proportional to the concentration of thrombin in the range from 1.0 to 20ng/mL and a detection limit of 0.3ng/mL thrombin was achieved. The fabricated aptasensor displayed attractive antifouling properties and allowed direct quantification of extrinsic thrombin down to 0.08ng/mL in undiluted serum sample. This work provides a promising strategy for clinical application with impressive sensitivity and antifouling characteristics. [Copyright &y& Elsevier]

Abstract: 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 NH2-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 &y& Elsevier]

Abstract: An ultra-sensitive impedimetric DNA sensor based on cascade signal amplification in sandwich manner was developed. A hairpin DNA was immobilized on a gold electrode and was acted as a probe DNA to improve the selectivity of the device. Without the target DNA, the hairpin DNA probe was in a “closed” state. However, in the presence of a target DNA, the hybridization between the loop moiety and the target broke the stem duplex. Consequently, the stem moiety at the 3′-end of the probes was moved away from the electrode surface and available for hybridization with the reporter DNA–Au nanoparticles conjugates (reporter DNA–AuNPs). The thiolated reporter DNA was complementary to the stem moiety at the 3′-end of the probe. After the probe DNA/target DNA/reporter DNA–AuNPs sandwich complex was formed on the electrode surface, the AuNPs were enlarged by immersing the electrode in a growth solution containing a negatively charged surfactant. Cascaded signal amplification was achieved because the electrode surface became more negatively charged after each assembly step. The electron-transfer resistance (Ret) of the DNA sensor increased with the increase in target DNA concentration through a wide linear range from 0.1fM to 300fM. Under optimized conditions, the DNA sensor could detect as low as 30aM of the fully matched target DNA. The sensor also showed excellent differentiation ability for single-base mismatch. [Copyright &y& Elsevier]

Abstract: A novel electrogenerated chemiluminescence (ECL) biosensor for highly sensitive detection of Escherichia coli (E. coli) was first developed by employing Concanavalin A (Con A) as a biological recognition element and bis(2,2′-bipyridine)-4′-methyl-4-carboxybipyridine ruthenium (II) (Ru1) complex as the detector. The ECL biosensor was fabricated by adsorbing carboxyl-functionalised single-wall carbon nanotubes (SWNTs) onto a paraffin-impregnated graphite electrode and further covalently coupling the Ru1–Con A probe onto the surface of the SWNT-modified electrode. Upon the binding of E. coli O157:H7 (as a model target), the biosensor showed a decreased ECL intensity in the presence of tri-n-propylamine (TPrA), which was in logarithmically direct proportion to the concentration of E. coli over a range from 5.0×102 to 5.0×105 cells/mL. The detection limit of this sensor was 127cells/mL. Additionally, the ECL biosensor also showed satisfactory selectivity in discriminating gram-negative E. coli from gram-positive bacteria. The strategy developed in this study may be a promising approach and could be extended to the design of ECL biosensors for highly sensitive and rapid detection of other desired bacteria. [Copyright &y& Elsevier]

Abstract: An ultrasensitive DNA detection method based on coulometric measurement of enzymatic silver deposition on gold nanoparticle (AuNP)-modified screen-printed carbon electrode (SPCE) was developed. The DNA sensor was prepared by self-assembly of a hairpin probe DNA, dually labeled with thiol at its 5′ end and biotin at its 3′ end, on AuNPs on SPCEs. The immobilized probe can shielded the biotin from being approached by a bulky alkaline phosphatase-linked streptavidin (Sv-ALP) conjugate. In the presence of the target DNA, hybridization caused conformational change in the probe DNA and forced biotin away from the surface of the AuNPs. Sv-ALP was bound to the biotin and catalyzed the hydrolysis of ascorbic acid 2-phosphate to form ascorbic acid. The latter reduced the silver ions for the deposition of metallic silver on the electrode surface. The deposited silver was then measured by coulometric analysis in the H2SO4 solution. A considerable increase in signal was obtained because of the accumulation and near-exhaustive coulometric oxidation of metallic silver. Under optimal conditions, the dynamic detection range of the target DNA was from 3.0×10−17 to 1.0×10−14 molL−1, and the detection limit was 1.5×10−17 molL−1. Further, the DNA sensor exhibited selectivity against single-base mismatched DNA. [Copyright &y& Elsevier]

Abstract: A novel electrogenerated chemiluminescence (ECL) biosensor for highly sensitive and selective detection of mercury ion was developed on the basis of mercury-specific oligonucleotide (MSO) served as a molecular recognition element and the ruthenium(II) complex (Ru1) as an ECL emitting species. The biosensor was fabricated on a glassy carbon electrode coated with a thin layer of single wall carbon nanotubes, where the ECL probe, NH2-(CH2)6-oligo(ethylene oxide)6-MSO↔Dend-Ru1, was covalently attached. The Dend-Ru1 pendant was prepared by covalent coupling Ru1 with the 4th generation polyamidoamine dendrimer (Dend), in which each dendrimer contained 35 Ru1 units so that a large amplification of ECL signal was obtained. Upon binding of Hg2+ to thymine (T) bases of the MSO, the T–Hg–T structure was formed, and the MSO changed from its linear shape to a “hairpin” configuration. Consequently, the Dend-Ru1 approached the electrode surface resulting in the increase of anodic ECL signal in the presence of the ECL coreactant tri-n-propylamine. The reported biosensor showed a high reproducibility and possessed long-term storage stability (92.3% initial ECL recovery over 30 day''s storage). An extremely low detection limit of 2.4pM and a large dynamic range of 7.0pM to 50nM Hg2+ were obtained. An apparent binding constant of 1.6×109 M−1 between Hg2+ and the MSO was estimated using an ECL based extended Langmuir isotherm approach involving multilayer adsorption. Determination of Hg2+ contents in real water samples was conducted and the data were consistent with the results from cold vapor atomic fluorescence spectroscopy. [Copyright &y& Elsevier]

Abstract: A glucose biosensor based on a nanocomposite made by layer-by-layer electrodeposition of the redox polymer into a multilayer containing glucose oxidase (GOx) and single-walled carbon nanotubes (SWCNT) on a screen-printed carbon electrode (SPCE) surface was developed. The objectives of the electrodeposition of redox polymer are to stabilize further the multilayer using a coordinative cross-linked redox polymer and to wire the GOx. The electrochemistry of the layer-by-layer assembly of the GOx/SWCNT/redox polymer nanocomposite was followed by cyclic voltammetry. The resultant biosensor provided stable and reproducible electrocatalytic responses to glucose, and the electrocatalytic current for glucose oxidation was enhanced with an increase in the number of layers. The biosensor displayed a linear range from 0.5 to 6.0mM, a sensitivity of 16.4μA/(mMcm2), and a response time of about 5s. It shows no response to 0.05mM of ascorbic acid, 0.32mM of uric acid and 0.20mM of acetaminophen using a Nafion membrane covering the nanocomposite-modified electrode surface. [Copyright &y& Elsevier]

Abstract: An amperometric glucose biosensor based on a multilayer made by layer-by-layer assembly of single-walled carbon nanotubes modified with glucose oxidase (GOx-SWCNT conjugates) and redox polymer (PVI-Os) on a screen-printed carbon electrode (SPCE) surface was developed. The SPCE surface was functionalized with a cationic polymer by electrodeposition of the PVI-Os, followed by alternating immersions in anionic GOx-SWCNT conjugate solutions and cationic PVI-O solutions. The purpose is to build a multilayer structure which is further stabilized through the electrodeposition of PVI-Os on the multilayer film. The electrochemistry of the layer-by-layer assembly of the GOx-SWCNT conjugate/PVI-Os bilayer was followed by cyclic voltammetry. The resultant glucose biosensor provided stable and reproducible electrocatalytic responses to glucose, and the electrocatalytic current for glucose oxidation was enhanced with an increase in the number of bilayers. The glucose biosensor displayed a wide linear range from 0.5 to 8.0mM, a high sensitivity of 32μAmM−1 cm−2, and a response time of less than 5s. The glucose biosensor proved to be promising amperometric detectors for the flow injection analysis of glucose. [Copyright &y& Elsevier]

Abstract: 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. [ABSTRACT FROM AUTHOR]

Abstract: 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 Pb2+ 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 Pb2+ in the range from 5.0 to 80pM with a detection limit of 1.4pM 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 &y& Elsevier]

Abstract: 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. [ABSTRACT FROM AUTHOR]

Abstract: A label-free electrochemical DNA biosensor array was developed as a model system for simultaneous detection of multiplexed DNAs using microlitres of sample. A novel multi-electrode array was comprised of six gold working electrodes and a gold auxiliary electrode, which were fabricated by gold sputtering technology, and a printed Ag/AgCl reference electrode was fabricated by screen-printing technology. The DNA biosensor array for simultaneous detection of the human immunodeficiency virus (HIV) oligonucleotide sequences, HIV-1 and HIV-2, was fabricated in sequence by self-assembling each of two kinds of thiolated hairpin-DNA probes onto the surfaces of the corresponding three working electrodes, respectively. The hybridization events were monitored by square wave voltammetry using methylene blue (MB) as a hybridization redox indicator. The oxidation currents of MB accumulated on the array decreased with increasing the concentration of HIVs due to higher affinity of MB for single strand rather than double strands of DNA. Under the optimized conditions, the peak currents were linear over ranges from 20 to 100nM for HIV-1 and HIV-2, with the same detection limits of 0.1nM (S/N=3), respectively. The biosensor array showed a good specificity without the obvious cross-interference. Furthermore, single-base mutation oligonucleotides and random oligonucleotides can be easily discriminated from complementary target DNAs. This work demonstrates that different hairpin-DNA probes can be used to design the label-free electrochemical biosensor array for simultaneous detection of multiplexed DNA sequences for various clinical applications. [Copyright &y& Elsevier]

Abstract: A label-free and sensitive faradic impedance spectroscopy (FIS) aptasensor based on target-induced aptamer displacement was developed for the determination of lysozyme as a model system. The aptasensor was fabricated by self-assembling the partial complementary single strand DNA (pcDNA)–lysozyme binding aptamer (LBA) duplex on the surface of a gold electrode. To measure lysozyme, the change in interfacial electron transfer resistance of the aptasensor using a redox couple of [Fe(CN)6]3−/4− as the probe was monitored. The introduction of target lysozyme induced the displacement of the LBA from the pcDNA–LBA duplex on the electrode into the solution, decreasing the electron transfer resistance of the aptasensor. The decrease in the FIS signal is linear with the concentration of lysozyme in the range from 0.2nM to 4.0nM, with a detection limit of 0.07nM. The fabricated aptasensor shows a high sensitivity, good selectivity and satisfactory regeneration. This work demonstrates that a high sensitivity of the fabricated aptasensor can be obtained using a relatively short pcDNA. This work also demonstrates that the target-induced aptamer displacement strategy is promising in the design of an electrochemical aptasensor for the determination of lysozyme with good selectivity and high sensitivity. [Copyright &y& Elsevier]

Abstract: A novel electrogenerated chemiluminescence (ECL) aptamer-based biosensing method for the determination of thrombin was developed on basis of a structure-switching ECL-dequenching mechanism. A thiolated ss-DNA capture probe, composing of a ss-DNA sequence to adopt two distinct structures-a DNA double strand with a complementary DNA sequence tagged with a ECL signal producer tris(2,2′-bipyridyl)ruthenium derivative and a DNA duplex with a thrombin-binding aptamer tagged with a ECL-quencher ferrocene (FcDNA), was self-assembled onto surface of a gold electrode. In the presence of thrombin, the aptamer sequence prefers to form the aptamer-thrombin complex and the switch of the binding partners occurs in conjunction with the generation of a strong ECL signal owing to the dissociation of FcDNA. The integrated ECL intensity versus the concentration of thrombin was linear in the range from 2.0×10−10 M to 2.0×10−7 M. The detection limit was 6×10−11 M. The relative standard derivation for 2.0×10−9 M was 5.7% (n =7). This work demonstrates that the sensitivity and specificity of ECL aptamer-based method for proteins can be greatly improved using quenching ECL signal producer by a chemical quencher such as ferrocene. [Copyright &y& Elsevier]

Abstract: A simple and highly sensitive electrochemical impedance spectroscopy (EIS) biosensor based on a thrombin-binding aptamer as molecular recognition element was developed for the determination of thrombin. The signal enhancement was achieved by using gold nanoparticles (GNPs), which was electrodeposited onto a glassy carbon electrode (GCE), as a platform for the immobilization of the thiolated aptamer. In the measurement of thrombin, the change in interfacial electron transfer resistance of the biosensor using a redox couple of [Fe(CN)6]3−/4− as the probe was monitored. The increase of the electron transfer resistance of the biosensor is linear with the concentration of thrombin in the range from 0.12nM to 30nM. The association and dissociation rate constants of the immobilized aptamer–thrombin complex were 6.7×103 M−1 s−1 and 1.0×10−4 s−1, respectively. The association and dissociation constants of three different immobilized aptamers binding with thrombin were measured and the difference of the dissociation constants obtained was discussed. This work demonstrates that GNPs electrodeposited on GCE used as a platform for the immobilization of the thiolated aptamer can improve the sensitivity of an EIS biosensor for the determination of protein. This work also demonstrates that EIS method is an efficient method for the determination of association and dissociation constants on GNPs modified GCE. [Copyright &y& Elsevier]

Abstract: A novel electrogenerated chemiluminescence aptamer-based (ECL-AB) biosensor for the determination of a small molecule drug is designed employing cocaine-binding aptamer as molecular recognition element for cocaine as a model analyte and ruthenium complex served as an ECL label. A 5′-terminal cocaine-binding aptamer with the ECL label at 3′-terminal of the aptamer was utilized as an ECL probe. The ECL-AB biosensors were fabricated by immobilizing the ECL probe onto a gold electrode surface via thiol-Au interactions. An enhanced ECL signal is generated upon recognition of the target cocaine, attributed to a change in the conformation of the ECL probe from random coil-like configuration on the probe-modified film to three-way junction structure, in close proximity to the sensor interface. The integrated ECL intensity versus the concentration of cocaine was linear in the range from 5.0×10−9 to 3.0×10−7 M. The detection limit was 1.0×10−9 M. This work demonstrates that the combination of a highly binding aptamer to analyte with a highly sensitive ECL technique to design ECL-AB biosensor is a great promising approach for the determination of small molecule drugs. [Copyright &y& Elsevier]