Electrochemical sensor for glutathione reductase based on screen-printed electrodes coated with 3,5-dinitrobenzoic acid-modified carbon nanotubes.

The preparation of a hybrid nanomaterial is reported by covalently attaching 3,5-dinitrobenzoic acid groups to the surface of oxidized multi-walled carbon nanotubes using 1,6-diaminohexane as cross-linking agent. This nanomaterial, modified with the redox mediator, was used as transduction element to construct an amperometric sensor for the efficient indirect determination of glutathione reductase at a low working potential of - 0.05 V, through the oxidation of unconsumed nicotinamide adenine dinucleotide phosphate (NADPH) in the enzymatic reaction. The sensor exhibited an excellent linear response in the range 1.6 to 174 µU/µL, with high reproducibility and selectivity. The developed device was successfully validated in real samples, accurately determining the active enzyme in diluted human serum, making it a promising alternative for the determination of glutathione reductase and other related NADPH-dependent enzymes with relevance in clinical analysis.

Sandwich-Type Electrochemical Aptasensor with Supramolecular Architecture for Prostate-Specific Antigen.

A novel sandwich-type electrochemical aptasensor based on supramolecularly immobilized affinity bioreceptor was prepared via host-guest interactions. This method utilizes an adamantane-modified, target-responsive hairpin DNA aptamer as a capture molecular receptor, along with a perthiolated ß-cyclodextrin (CD) covalently attached to a gold-modified electrode surface as the transduction element. The proposed sensing strategy employed an enzyme-modified aptamer as the signalling element to develop a sandwich-type aptasensor for detecting prostate-specific antigen (PSA). To achieve this, screen-printed carbon electrodes (SPCEs) with electrodeposited reduced graphene oxide (RGO) and gold nanoferns (AuNFs) were modified with the CD derivative to subsequently anchor the adamantane-modified anti-PSA aptamer via supramolecular associations. The sensing mechanism involves the affinity recognition of PSA molecules on the aptamer-enriched electrode surface, followed by the binding of an anti-PSA aptamer-horseradish peroxidase complex as a labelling element. This sandwich-type arrangement produces an analytical signal upon the addition of H2O2 and hydroquinone as enzyme substrates. The aptasensor successfully detected the biomarker within a concentration range of 0.5 ng/mL to 50 ng/mL, exhibiting high selectivity and a detection limit of 0.11 ng/mL in PBS.

Electrochemical aptasensor based on anisotropically modified (Janus-type) gold nanoparticles for determination of C-reactive protein.

Novel Janus nanoparticles based on Au colloids anisotropically modified with polyamidoamine dendrons were prepared though a masking/toposelective modification approach. These nanomaterials were further functionalized with horseradish peroxidase on the dendritic face and provided on the opposite metal surface with a ssDNA aptamer for C-reactive protein (CRP). The resulting nanoparticles were employed as biorecognition/signaling elements to construct an amperometric aptasensor with sandwich-type architecture for the specific detection of this cardiac biomarker. To do this, screen-printed carbon electrodes modified with electrodeposited Au nanoparticles and functionalized with anti-CRP aptamers were used as transduction interface. The aptasensor was employed for the amperometric detection of CRP (working potential: - 200 mV vs pseudo-Ag/AgCl) in the broad range from 10 pg·mL-1 to 1.0 ng·mL-1 with a detection limit of 3.1 pg·mL-1. This electroanalytical device also showed good specificity, reproducibility (RSD = 9.8%, n = 10), and stability and was useful to quantify CRP in reconstituted human serum samples, with a RSD of 13.3%.

Electrochemical biosensors based on nucleic acid aptamers.

During recent decades, nucleic acid aptamers have emerged as powerful biological recognition elements for electrochemical affinity biosensors. These bioreceptors emulate or improve on antibody-based biosensors because of their excellent characteristics as bioreceptors, including limitless selection capacity for a large variety of analytes, easy and cost-effective production, high stability and reproducibility, simple chemical modification, stable and oriented immobilization on electrode surfaces, enhanced target affinity and selectivity, and possibility to design them in target-sensitive 3D folded structures. This review provides an overview of the state of the art of electrochemical aptasensor technology, focusing on novel aptamer-based electroanalytical assay configurations and providing examples to illustrate the different possibilities. Future prospects for this technology are also discussed. Graphical abstract.

Self-propelled enzyme-controlled IR-mesoporous silica Janus nanomotor for smart delivery.

Here, we report the preparation of a novel Janus nanoparticle with opposite Ir and mesoporous silica nanoparticles through a partial surface masking with toposelective modification method. This nanomaterial was employed to construct an enzyme-powered nanomachine with self-propulsion properties for on-command delivery. The cargo-loaded nanoparticle was provided with a pH-sensitive gate and unit control at the mesoporous face by first attaching boronic acid residues and further immobilization of glucose oxidase through reversible boronic acid esters with the carbohydrate residues of the glycoenzyme. Addition of glucose leads to the enzymatic production of H2O2 and gluconic acid, being the first compound catalytically decomposed at the Ir nanoparticle face producing O2 and causing the nanomachine propulsion. Gluconic acid leads to a pH reduction at the nanomachine microenvironment causing the disruption of the gating mechanism with the subsequent cargo release. This work demonstrates that enzyme-mediated self-propulsion improved release efficiency being this nanomotor successfully employed for the smart release of Doxorubicin in HeLa cancer cells.

An intercommunicated nanosystem for dual delivery.

Here, we describe the design of a novel particle-to-particle intercommunicated nanosystem for dual delivery, triggered by physical and chemical inputs. The nanosystem was composed of an Au-mesoporous silica Janus nanoparticle loaded with paracetamol, mechanized with light-sensitive supramolecular gates at the mesoporous face and functionalized on the metal surface with the enzyme acetylcholinesterase. The second component was a mesoporous silica nanoparticle loaded with rhodamine B and gated with thiol-sensitive ensembles. Upon irradiation of this nanosystem with a near-UV light laser, an analgesic drug was released from the Janus nanomachine due to disassembling of the photosensitive gating mechanism. Further addition of N-acetylthiocholine leads to the enzymatic production of thiocholine at the Janus nanomachine, thus acting as a "chemical messenger" causing the disruption of the gating mechanism at the second mesoporous silica nanoparticle with the subsequent dye release.

Supramolecular Enzymatic Labeling for Aptamer Switch-Based Electrochemical Biosensor.

Here we report a novel labeling strategy for electrochemical aptasensors based on enzymatic marking via supramolecular host-guest interactions. This approach relies on the use of an adamantane-modified target-responsive hairpin DNA aptamer as an affinity bioreceptor, and a neoglycoconjugate of ß-cyclodextin (CD) covalently attached to a redox enzyme as a labeling element. As a proof of concept, an amperometric aptasensor for a carcinoembryonic antigen was assembled on screen-printed carbon electrodes modified with electrodeposited fern-like gold nanoparticles/graphene oxide and, by using a horseradish peroxidase-CD neoglycoenzyme as a biocatalytic redox label. This aptasensor was able to detect the biomarker in the concentration range from 10 pg/mL to 1 ng/mL with a high selectivity and a low detection limit of 3.1 pg/mL in human serum samples.

An enzyme-controlled mesoporous nanomachine for triple-responsive delivery.

The construction of a novel enzyme-controlled nanomachine with multiple release mechanisms for on-command delivery is described. This nanodevice was assembled by modifying mesoporous silica nanoparticles with 2-(benzo[d]thiazol-2-yl)phenyl 4-aminobenzoate moieties, and further capped with ß-cyclodextrin-modified glucose oxidase neoglycoenzyme. The device released the encapsulated payload in the presence of H2O2 and acidic media. The use of glucose as an input chemical signal also triggered cargo release through the enzymatic production of gluconic acid and hydrogen peroxide, and the subsequent disruption of the gating mechanism at the mesoporous surface. The nanodevice was successfully employed for the enzyme-controlled release of doxorubicin in HeLa cancer cells.

Amperometric aptasensor with sandwich-type architecture for troponin I based on carboxyethylsilanetriol-modified graphene oxide coated electrodes.

A novel amperometric aptasensor for the specific detection of cardiac troponin I (cTnI) was constructed by using screen-printed carbon electrodes coated with a carboxyethylsilanetriol-modified graphene oxide derivative as transduction element. This novel carboxylic acid-enriched nanomaterial allows easy and high load immobilization of the capture aptamer molecules on the electrode surface. The biosensing interface was assembled by covalent attachment of an amino-functionalized DNA aptamer on the carboxylic acid-enriched electrode surface. The sensing approach relies on the specific recognition of cTnI by the aptamer and further assembly of a sandwich-type architecture with a novel aptamer-peroxidase conjugate as signaling element. The aptasensor was employed to detect the cardiac biomarker in the broad range from 1.0 pg/mL to 1.0 µg/mL with a detection limit of 0.6 pg/mL. This electroanalytical device also showed high specificity, reproducibility and stability, and was useful to quantify cTnI in reconstituted human serum samples.

A glutathione disulfide-sensitive Janus nanomachine controlled by an enzymatic AND logic gate for smart delivery.

This work describes the assembly of a novel enzyme-controlled nanomachine operated through an AND Boolean logic gate for on-command delivery. The nanodevice was constructed on Au-mesoporous silica Janus nanoparticles capped with a thiol-sensitive gate-like molecular ensemble on the mesoporous face and functionalized with glutathione reductase on the gold face. This autonomous nanomachine employed NADPH and glutathione disulfide as input chemical signals, leading to the enzymatic production of reduced glutathione that causes the disruption of the gating mechanism on the mesoporous face and the consequent payload release as an output signal. The nanodevice was successfully used for the autonomous release of doxorubicin in HeLa cancer cells and RAW 264.7 macrophage cells.

Dendrimers as Soft Nanomaterials for Electrochemical Immunosensors.

Electrochemical immunosensors are antibody-based affinity biosensors with a high impact on clinical, environmental, food, and pharmaceutical analysis. In general, the analytical performance of these devices is critically determined by the materials and reagents used for their construction, signal production and amplification. Dendrimers are monodisperse and highly branched polymers with three-dimensional structures widely employed as "soft" nanomaterials in electrochemical immunosensor technology. This review provides an overview on the state-of-the-art in dendrimer-based electrochemical immunosensors, focusing on those using polyamidoamine and poly (propylene imine) dendrimers. Special emphasis is given to the most original methods recently reported for the construction of immunosensor architectures incorporating dendrimers, as well as to novel sensing approaches based on dendrimer-assisted signal enhancement strategies.

Amperometric aptasensor for carcinoembryonic antigen based on the use of bifunctionalized Janus nanoparticles as biorecognition-signaling element.

We report herein the design of a novel biosensing strategy for the detection of carcinoembryonic antigen (CEA), based on the use of Janus-type nanoparticles having Au and silica opposite faces as integrated electrochemical biorecognition-signaling system. The Janus nanoparticles were properly functionalized with horseradish peroxidase on the silica surface to act as signaling element, and a biotin thiol-modified anti-CEA DNA hairpin aptamer the Au face to assemble the biorecognition element. The sensing approach relies on the first specific recognition of CEA by the bifunctionalized Janus nanoparticles, causing unfolding of the DNA hairpin structure and unmasking the biotin residues at the aptamer chain. This CEA-Janus nanoparticle complex was then captured by avidin-modified Fe3O4@SiO2 NanoCaptors®, allowing further magnetic deposition on carbon screen printed electrodes for the amperometric detection of the cancer biomarker. The Janus nanoparticles-based aptasensor was able to detect CEA in the range from 1 to 5000 ng mL-1 (5.5 pM-28 nM) with a detection limit of 210 pg mL-1 (1.2 pM). The aptasensor also showed high reproducibility and storage stability, and was successfully validated in human serum.