Ultrasensitive Electrochemical Biosensor Based on Efficient PDA-APDMAO Antifouling Interface and Dual-Signal Ratio Strategy for Trace Detection of Alpha-Fetoprotein in Human Serum.

In electrochemical analysis, developing biosensors that can resist the nonspecific adsorption of interfering biomolecules in human serum remains a huge challenge, which depends on the design of efficient antifouling materials. Herein, 3-aminopropyldimethylamine oxide (APDMAO) biomimetic zwitterions were prepared as antifouling interfaces. Among them, the unique positive and negative charges (N+-O-) of APDMAO promoted its hydrogen bonding with water molecules, forming a firm hydration barrier that endowed it with strong and stable antifouling performance. Meanwhile, its inherent amino groups could copolymerize with the biomimetic adhesive dopamine to form a thin layer of quinone intermediates, providing conditions for the subsequent binding of aptamers and signal probes. Importantly, the biomimetic APDMAO with functional groups and one-step oxidation characteristics solved the challenges of zwitterionic synthesis and modification, as well as improved biocompatibility of the sensing interface, thereby expanding the application potential of zwitterions as antifouling materials in sensing analysis. Thiol-containing alpha-fetoprotein (AFP) aptamers modified with methylene blue (MB) were coupled under controllable potential, greatly reducing the incubation time, which promoted the productization application of biosensors. In addition, the ratio sensing strategy using MB as internal standard factors and concanavalin-silver nanoparticles (ConA-Ag NPs) as signal probes was introduced to reduce background and instrument interferences, thus improving detection accuracy. On this basis, the proposed antifouling electrochemical biosensor achieved sensitive and accurate AFP detection over a wide dynamic range (10 fg/mL-10 ng/mL), with a low detection limit of 3.41 fg/mL (3σ/m). This work provides positive insights into the development of zwitterionic antifouling materials and clinical detection of liver cancer markers in human serum.

Highly Efficient Signal On/Off Electrochemiluminescence Gel Aptasensor Based on a Controlled Release Strategy for the Sensitive Detection of Prostate Specific Antigen.

The controlled release strategy can make the constructed sensor have the function of self-on/off, which has an obvious effect on improving the sensitivity in immunoassays. Metal organic gels (MOGs) are the most noteworthy. They are materials with ultrahigh surface area, highly dispersed atomical metal sites, and well-defined porosity and can be used as an efficient luminophore to cause the developed sensor to have good hydrophilicity and adjustability, thus further improving the detection sensitivity. In this work, a novel on/off electrochemiluminescence (ECL) gel aptasensor was constructed using the Cys-[Ru(dcbpy)3]2+ gel as a luminophore, ZnS quantum dots (QDs) as quenchers, and aminated mesoporous silica nanocontainers (SiO2-NH2) as carriers of controlled release for prostate specific antigen (PSA) detection. Specifically, the ssDNA and PSA aptamer made up clamp-like molecules to block holes of the SiO2-NH2 after encapsulating the quencher ZnS QDs. Because of the specific binding between the PSA antigen and aptamer, the clamp-like molecules of ssDNA and the PSA aptamer were disassembled. Finally, the release of ZnS QDs was triggered, thereby realizing a self-off mode of the ECL signals under a co-reactant-free environment by ECL resonance energy transfer (ECL-RET) between the Cys-[Ru(dcbpy)3]2+ and ZnS QDs. In addition, the quenching mechanism was confirmed by molecular orbitals from the theoretical calculation level. The detection limit of the gel aptasensor for PSA was as low as 1.01 fg/mL, showing excellent sensitivity and accuracy. These strategies provided a feasible idea for PSA and even other tumor marker immunoassays.

Ultrasensitive Electrochemiluminescence Biosensor with Silver Nanoclusters as a Novel Signal Probe and α-Fe2O3-Pt as an Efficient Co-reaction Accelerator for Procalcitonin Immunoassay.

Herein, a high-efficiency biosensor based on ternary electrochemiluminescence (ECL) system was constructed for procalcitonin (PCT) detection. Specifically, silver nanoclusters (Ag NCs) with stable luminescence properties were prepared with small-molecule lipoic acid (LA) as the ligand, and its ECL emission in persulfate (S2O82-) was first reported. Meanwhile, the prepared Ag NCs possessed ligand-to-metal charge-transfer characteristics, thus transferring energy from LA to Ag+ for luminescence. Based on the small particle size, good biocompatibility, and molecular binding ability, Ag NCs-LA was used as an ideal luminescent probe. In addition, α-Fe2O3-Pt was introduced to facilitate the activation of S2O82-, thereby generating more sulfate radicals to react with the free radicals of Ag NCs to enhance ECL emission. The synergistic effect of the variable valence state of transition metals and high catalytic activity of noble metals endows α-Fe2O3-Pt with excellent catalytic ability for S2O82-. Importantly, the sensing mechanism was systematically demonstrated by UV-vis, fluorescence, and ECL analysis, as well as density functional theory calculations. At last, NKFRGKYKC was designed for specific immobilization of antibodies, thus releasing the antigen binding sites to improve the antigen recognition efficiency. Based on this, the developed biosensor showed high sensitivity for PCT detection, with a wide linear range (10 fg/mL-100 ng/mL) and a low detection limit (3.56 fg/mL), which could be extended to clinical detection of multiple biomarkers.

Europium-based metal-organic framework with acid-base buffer structure as electrochemiluminescence luminophore for hyperstatic trenbolone trace monitoring under wide pH range.

The wide and even whole pH range electrochemiluminescence (ECL) is attractive for steroid estrogens detection under harsh conditions (such as strong acid and alkali). Herein, we presented an efficient europium-based metal-organic framework (Eu-MOF) as ECL luminophore, which has been synthesized via the specific 2, 4-bis(3, 5-dicarboxyphenylamino)-6-oltriazine (H4BDPO) ligand with acid-base buffering effect. The functional groups with weak acid and base endowed the H4BDPO with eight ionogenic group states, thereout different total charges of H4BDPO were derived, thus high and steady ECL signals of Eu-MOF were acquired under different environments with pH = 1.0-14.0. Most notably, combined with the means of UV-vis, fluorescence spectra, cyclic voltammetry (CV) and density functional theory (DFT) calculations, the Eu-MOF has been explored different luminescence mechanisms with variational total charges. The constructed ECL biosensor based on the Eu-MOF realized sensitive detection of trenbolone under wide pH range (In order to maintain the biological activity of antigen and antibody, the studied pH value is 5-8.5), in which the limits of detection were 3.95 fg/mL (pH = 5.0), 2.36 fg/mL (pH = 7.4) and 5.48 fg/mL (pH = 8.5) respectively. This work provides a considerable method to realize efficient trace detection of steroid estrogens under the wide or even whole pH conditions.

High-bioactivity microfluidic immunosensing platform for electrochemiluminescence determination of CYFRA 21-1 with the introduction of Fe3O4@Cu@Cu2O.

Relying on the electrochemiluminescence (ECL) and microfluidic technology, an immunosensor chip with high bioactivity was designed for sensitive determination of cytokeratin 19 fragment 21-1 (CYFRA 21-1). The mesoporous nanomaterial Fe3O4@Cu@Cu2O as the co-reaction accelerator was used to catalyze the S2O82- to produce more SO4•- to achieve the amplification of the ECL signal. In fact, the generating of SO4•- could not only be done with the aid of the reversible cycles of Fe2+ and Fe3+ and Cu+ and Cu2+, but could be achieved also through the catalase-like function of Fe3O4. What is more, it has also been proved that Fe3O4@Cu@Cu2O exhibited better catalytic performance than single Fe3O4, Cu2O, and Cu@Cu2O, which supported its application in this system. In addition, a portable microfluidic immunosensor chip for CYFRA 21-1-sensitive determination was assembled, which showed high selectivity, sensitivity, and strong universality in clinical cancer screening and diagnosis. It should be noted that HWRGWVC (HWR) was introduced as the antibody fixator to improve the incubation and binding efficiency of the antibody, which increased the ECL intensity and improved the sensitivity of the immunosensor. This strategy provided a new idea for cancer identification and diagnosis in clinical medicine.

Catalytic Decomposition of the Hole-Derived H2O2 by AgBiS2@Ag Nanozyme to Enhance the Photocurrent of Z-Scheme BiVO4/ZnIn2S4 Photoelectrode in Microfluidic Immunosensing Platform.

A novel microfluidic photoelectrochemical (PEC) analytical device based on AgBiS2@Ag nanozyme-mediated signal amplification was developed for ultrasensitive detection of cytokeratin 19 fragment 21-1 (CYFRA 21-1). First, a brand new Z-scheme BiVO4/ZnIn2S4 (BZIS) photoactive material was utilized as a sensing matrix to supply a stable photocurrent. Under anodic bias, the photoexcited holes in BiVO4 could oxidize water to produce hydrogen peroxide (H2O2), which markedly enhanced the separation efficiency of the electron-hole pairs. Besides, the Z-scheme heterojunction formed between BiVO4 and ZnIn2S4 further accelerated the transport of the electron. Second, for improving the sensitivity of the PEC sensor, a new strategy of catalytic dissociation of the hole-derived H2O2 by AgBiS2@Ag nanozyme was proposed to amplify the PEC signal. AgBiS2@Ag composites, possessing an excellent peroxidase-mimicking feature, could efficiently catalyze the H2O2 to produce hydroxyl radicals (•OH) and lead to the significant enhancement of the photocurrent. Third, automatic sample injection and detection were successfully realized by integrating the photoelectrode into microfluidic chips. Based on this advanced sensing strategy, the designed microfluidic PEC sensor displayed a wide linear range (0.1 pg/mL - 100 ng/mL) and a low detection limit of 35 fg/mL (S/N = 3), which could be efficiently applied to the ultrasensitive determination of CYFRA 21-1 in a human serum sample.

Efficient ABEI-Dissolved O2-Ce(III, IV)-MOF Ternary Electrochemiluminescent System Combined with Self-Assembled Microfluidic Chips for Bioanalysis.

A signal-amplified electrochemiluminescent (ECL) sensor chip was developed for sensitive analysis of procalcitonin (PCT). Herein, we first prepared a self-enhanced luminophore, which enhanced ECL responses through intramolecular reactions. Second, Au-Pd bimetallic nanocrystals and mixed-valence Ce-based metal-organic frameworks (MOFs) were introduced as co-reaction promoters to facilitate the reduction of dissolved O2. Based on the synergistic catalysis of Au and Pd, the spontaneous cyclic reaction of Ce(III)/Ce(IV), and the high electrochemical active surface area of Ce(III, IV) MOF, a large number of superoxide anion radicals (O2•-) and hydroxyl radicals (OH•) were produced. Therefore, the luminescence efficiency of N-(aminobutyl)-N-(ethylisoluminol)-dissolved O2 (ABEI-O2) systems were greatly improved, providing a new prospect for the application of dissolved O2 in ECL analysis. In addition, the affinity peptide ligands were used for the directional connection of antibodies to provide protection for the bioactivity of the proposed sensor. Finally, the microfluidic technology was applied to ECL analysis to integrate the three-electrode detection system into the self-assembled microfluidic chip, which realized the automation and portability of the detection process. The developed sensor showed high sensitivity for PCT detection with a detection limit of 3.46 fg/mL, which possessed positive significance for the clinical diagnosis of sepsis.

High-Efficiency CdSe Quantum Dots/Fe3O4@MoS2/S2O82- Electrochemiluminescence System Based on a Microfluidic Analysis Platform for the Sensitive Detection of Neuron-Specific Enolase.

In this work, based on electrochemiluminescence (ECL) technology and self-assembled portable disease detection chips, a bioactivity-maintained sensing platform was developed for the quantitative detection of neuron-specific enolase. First, we prepared Fe3O4@MoS2 nanocomposites as an efficient catalyst to accelerate the reduction of persulfate (S2O82-). Specifically, abundant sulfate radicals (SO4•-) were generated because of cyclic conversion between Fe2+ and Fe3+. Meanwhile, MoS2 nanoflowers with a high specific surface area could not only load more Fe3O4 but also solve its agglomeration problem, which greatly improved the catalytic efficiency. Moreover, a biosensor chip was constructed by standard lithography processes for disease detection, which had good sensitivity and portability. According to the above strategies, the developed portable sensing platform played the part of promoting the practical application of bioanalysis in early tumor screening and clinical diagnosis. In addition, we developed a short peptide ligand (NARKFYKG, NAR) to avoid the occupation of antigen binding sites by specifically connecting to Fc fragments in antibodies. Thus, the binding efficiency of antibodies and the activity of biosensors were improved due to the introduction of NAR.

A dual signal-amplified electrochemiluminescence immunosensor based on core-shell CeO2-Au@Pt nanosphere for procalcitonin detection.

A dual signal-amplified sandwich electrochemiluminescence (ECL) immunosensor was fabricated for trace detection of procalcitonin (PCT). CeO2-Au@Pt composed of sea urchin-like Au@Pt nanoparticles coated on CeO2 hollow nanospheres was immobilized on electrode surface to electrochemically catalyze H2O2 to produce a large number of superoxide anion (O2•-). The immunosensor was prepared by linking the capture antibody on immobilized CeO2-Au@Pt with heptapeptide (HWRGWVC), which could maintain the activity of the antibody. The prepared Au star@BSA was used to bind abundant luminol for labeling the secondary antibody (Ab2). Upon the sandwich-typed immunoreactions, the O2•- could react with the introduced luminol on the immunosensor surface to produce strong ECL intensity. With an outstanding linear detection range and a low detection limit of 17 fg/mL, the ECL immunosensor permitted ultrasensitive detection of PCT at a low H2O2 concentration and demonstrated its high application potential in the clinical assay.

Peptide-Based Electrochemiluminescence Biosensors Using Silver Nanoclusters as Signal Probes and Pd-Cu2O Hybrid Nanoconcaves as Coreactant Promoters for Immunoassays.

Metal nanoclusters (NCs) possess high light stability and biocompatibility because of their unique quantum size effect, which has gradually become a new type of electrochemiluminescence (ECL) nanomaterial for immunoassays. However, the luminescence efficiency of metal NCs is too low to meet the needs of trace analysis, which limits its application. Herein, Ag NCs served as signal probes and Pd-Cu2O hybrid nanoconcaves served as coreaction promoters, developing a highly efficient peptide-based biosensor for neuron-specific enolase (NSE) detection. Utilizing the reversible cycle of Cu+/Cu2+ and the reduction characteristics of Pd NPs, Pd-Cu2O greatly accelerates the reduction of S2O82-. Meanwhile, Pd-Cu2O has good hydrogen evolution activity, which promotes the generation of oxygen by improving the redox efficiency of the overall reaction, thus increasing the yield of active intermediates (OH•) to promote the reduction of S2O82-. Specially, this is an effective attempt to use the hydrogen evolution reaction (HER) to accelerate the ECL emission of the S2O82- system. In addition, a short peptide ligand (NARKFYKGC, NFC) was developed to implement the targeted immobilization of antibodies, which can specifically bind to the Fc fragment of antibodies, thereby avoiding the occupation of the antigen binding site (Fab fragment). The introduction of NFC not only improves the binding efficiency of antibodies but also protects its bioactivity, thus significantly improving the sensitivity of the biosensor. Based on these strategies, the proposed biosensor provides a new perspective for the applications of metal NCs in ECL systems.

Peptide-Based Biosensor with a Luminescent Copper-Based Metal-Organic Framework as an Electrochemiluminescence Emitter for Trypsin Assay.

A copper-based metal-organic framework (JUC-1000) has emerged as a promising electrochemiluminescence (ECL) emitter in the domains of bioanalysis and immunoassay. Herein, a highly efficient signal "on-off" peptide-based biosensor was constructed for trypsin (TPN) assay. JUC-1000 synthesized using an organic ligand of H4BDPO was functionalized as the ECL emitter, whose cathodic ECL behavior in aqueous media was first investigated using potassium persulfate (K2S2O8) as the coreactant. To further amplify the ECL signal, highly catalytic Ag@CeO2 nanoparticles were fabricated as both a substrate and an coreaction accelerator, which can efficiently catalyze the reduction of S2O82- to generate more sulfate anion radicals (SO4•-) for ECL enhancement, thereby generating strong and stable ECL signals in a "signal on" state. The functionalized JUC-1000 emitter was connected to the Ag@CeO2 sensing layer though a heptapeptide (HWRGWVC, HGC), and TPN as the target can specifically cleave the carboxyl side of arginine residues in HGC, leading to the release of emitters in a "signal off" state. Based on the efficient signal-switching, the biosensor exhibited linear ECL responses to the added TPN concentration, realizing sensitive detection of TPN in 10 fg/mL to 100 ng/mL with a limit of detection of 3.46 fg/mL. This work proposed an attractive orientation for the fundamental research of applying transition metal-organic frameworks as ECL emitters in bioanalysis and immunoassay.

Ultrasensitive near-infrared electrochemiluminescence biosensor derived from Eu-MOF with antenna effect and high efficiency catalysis of specific CoS2 hollow triple shelled nanoboxes for procalcitonin.

In this paper, we report a novel multiple amplification strategy for ultrasensitive near-infrared electrochemiluminescence (ECL) immunoassay in K2S2O8 solution. The realization of this strategy is based on the antenna effect of Eu-MOF (EuBTC) and a high efficiency catalysis of CoS2 hollow triple shelled nanoboxes (TSNBs). The H3BTC ligand in the antenna effect first undergoes π-π* absorption and a singlet-singlet electronic transition. Its energy passes through the intersystem to the triplet state, next transfers from the lowest excited triplet state to the vibrational energy level of the rare earth ion, finally realizing sensitizing center ion luminescence. Moreover, ionic reaction and structural advantages endow CoS2 TSNBs a dual signal enhancement effect. This sandwich-type ECL biosensor has a near-infrared luminescence in 800-900 nm, thus avoiding damage to the sample in the meantime. In practical diagnosis, the normal critical value of procalcitonin (PCT) (<0.5 ng/mL) is much higher than the detection limit (3.65 fg/mL) and is in the detection range (10 fg/mL-100 ng/mL), which means that the ECL biosensor has a high sensitivity in the detection of PCT and meet the requirement for diagnosis of disease completely. Therefore, the strategy provides a feasible method for efficient and stable analysis of systemic inflammatory response such as fearful bacterial infection, hepatitis B, and peritonitis.

Rare Self-Luminous Mixed-Valence Eu-MOF with a Self-Enhanced Characteristic as a Near-Infrared Fluorescent ECL Probe for Nondestructive Immunodetection.

Steady and efficient sensitized emission of Eu2+ to Eu3+ can be achieved through a rare mixed-valence Eu-MOF (L4EuIII2EuII). Compared with the sensitization of other substances, the similar ion radius and configuration of the extranuclear electron between Eu2+ and Eu3+ make sensitization easier and more efficient. The sensitization of Eu2+ to Eu3+ is of great assistance for the self-enhanced luminescence of L4EuIII2EuII, the longer luminous time, and the more stable electrochemiluminescence (ECL) signal. Simultaneously, L4EuIII2EuII possesses near-infrared (NIR) fluorescence of around 900 nm and a mighty self-luminous characteristic, which render it useful as a NIR fluorescent probe and as a luminophore to establish a NIR ECL biosensor. This NIR biosensor can greatly reduce the damage to the detected samples and even achieve a nondestructive test and improve the detection sensitivity by virtue of strong susceptibility and environmental suitability of NIR. In addition, the CeO2@Co3O4 triple-shelled microspheres further enhanced the ECL intensity due to two redox pairs of Ce3+/Ce4+ and Co2+/Co3+. The NIR ECL biosensor based on these strategies owns an ultrasensitive detection ability of CYFRA 21-1 with a low limit of detection of 1.70 fg/mL and also provides a novel idea for the construction of a highly effective nondestructive immunodetection biosensor.

Triple Amplification of 3,4,9,10-Perylenetetracarboxylic Acid by Co2+-Based Metal-Organic Frameworks and Silver-Cysteine and Its Potential Application for Ultrasensitive Assay of Procalcitonin.

In this work, a triple-amplified biosensor with a bioactivity-maintained peculiarity was constructed for quantitative procalcitonin (PCT) detection. As everyone knows, a strong electrochemiluminescence (ECL) signal is the premise to ensure high sensitivity for trace target detection. Hence, a valid tactic was developed to achieve signal amplification of luminophor by using Co2+-based metal-organic frameworks (ZIF-67) and silver-cysteine (AgCys). The ZIF-67 particles, which have more atomically dispersed Co2+, could play the role of a co-reaction accelerator to catalyze S2O82- to generate abundant Co3+ and sulfate radical anions (SO4•-). Afterward, a mass of Co3+ was reduced to more hydroxyl radicals (OH•) by H2O, thus ulteriorly reducing S2O82- to generate more SO4•-. Remarkably, S2O82- was reduced to SO4•- continuously with the recycling of Co2+ and Co3+, which realized an effective signal amplification. Meanwhile, the AgCys complex with superior catalysis and biocompatibility was prepared to further improve the ECL signal and maintain the bioactivity of the biomolecule. Furthermore, HWRGWVC, a heptapeptide that was used for combining the Fc fragments of an antibody by Au-S bonding to achieve the fixed point fixation, could not only maintain bioactivity of an antibody but also improved its incubation efficiency, thus further enhancing biosensor sensitivity. Under optimum conditions, the proposed biosensor realized highly sensitive assay for PCT with a wide dynamic range from 10 fg/mL to 100 ng/mL and a detection limit as low as 3.67 fg/mL. With superior stability, selectivity, and repeatability, the prepared biosensor revealed immense potential application of ultrasensitive assay for PCT in human serum.