Phospholipid Bilayer Integrated with Multifunctional Peptide for Ultralow-Fouling Electrochemical Detection of HER2 in Human Serum.

Electrochemical biosensing devices face challenges of severe nonspecific adsorption in complex biological matrices for the detection of biomarkers, and thus, there is a significant need for sensitive and antifouling biosensors. Herein, a sensitive electrochemical biosensor with antifouling and antiprotease hydrolysis ability was constructed for the detection of human epidermal growth factor receptor 2 (HER2) by integrating multifunctional branched peptides with distearoylphosphatidylethanolamine-poly(ethylene glycol) (DSPE-PEG) self-assembled bilayer. The peptide was designed to possess antifouling, antiprotease hydrolysis, and HER2 recognizing capabilities. Molecular dynamics simulations demonstrated that the DSPE was able to effectively self-assemble into a bilayer, and the water contact angle and electrochemical experiments verified that the combination of peptide with the DSPE-PEG bilayer was conducive to enhancing the hydrophilicity and antifouling performance of the modified surface. The constructed HER2 biosensor exhibited excellent antifouling and antiprotease hydrolysis capabilities, and it possessed a linear range of 1.0 pg mL-1 to 1.0 µg mL-1, and a limit of detection of 0.24 pg mL-1. In addition, the biosensor was able to detect HER2 in real human serum samples without significant biofouling, and the assaying results were highly consistent with those measured by the enzyme-linked immunosorbent assay (ELISA), indicating the promising potential of the antifouling biosensor for clinical diagnosis.

DNA liquid crystals with AIE effect toward humidity-indicating biomaterials.

In this study, by fabricating DNA doped with tetraphenylethene-containing ammonium surfactant, the resulting solvent-free DNA ionic complex could undergo a humidity-induced phase change that could be well tracked by the fluorescence signal of the surfactant. Taking advantage of the humidity-induced change in fluorescence, the reported ionic DNA complex could accurately indicate the humidity in real time.

Antifouling strategies for electrochemical sensing in complex biological media.

Surface fouling poses a significant challenge that restricts the analytical performance of electrochemical sensors in both in vitro and in vivo applications. Biofouling resistance is paramount to guarantee the reliable operation of electrochemical sensors in complex biofluids (e.g., blood, serum, and urine). Seeking efficient strategies for surface fouling and establishing highly sensitive sensing platforms for applications in complex media have received increasing attention in the past. In this review, we provide a comprehensive overview of recent research efforts focused on antifouling electrochemical sensors. Initially, we present a detailed illustration of the concept about biofouling along with an exploration of four key antifouling mechanisms. Subsequently, we delve into the commonly employed antifouling strategies in the fabrication of electrochemical sensors. These encompass physical surface topography (micro/nanostructure coatings and filtration membranes) and chemical surface modifications (PEG and its derivatives, zwitterionic polymers, peptides, proteins, and various other antifouling materials). The progress in antifouling electrochemical sensors is proposed concerning the antifouling mechanisms as well as sensing capability assessments (e.g., sensitivity, stability, and practical application ability). Finally, we summarize the evolving trends in the field and highlight some key remaining limitations.

Phosphorylation of Oligopeptides: Design of Ultra-Hydrophilic Zwitterionic Peptides for Anti-Fouling Detection of Nucleic Acids in Saliva.

The construction of low-fouling biosensors for assaying biomarkers in complex biological samples remains a challenge, and the key limitation is the lack of effective anti-fouling materials. Inspired by the biomimetic process of protein phosphorylation, we herein designed a new phosphorylated peptide modified with the dihydrogen phosphate (-PO4H2) group, which significantly increased the hydrophilicity and anti-fouling capability of the peptide when compared with natural and normal peptides. Molecular simulation (MS) illustrated that, compared with the -COOH and -NH2 groups, the -PO4H2 group formed the most numbers of hydrogen bonds and stronger hydrogen bonds with water molecules. As a result, the PO4H2-oligopeptide was proved by MS to be able to attract the greatest number of water molecules, so as to form a compact layer of H2O to resist further adsorption of nonspecific biomolecules. The modification of electrodes with the designed PO4H2-oligopeptides, in addition to the adoption of neutral peptide nucleic acids (PNAs) as the sensing probes, ensured the fabrication of anti-fouling electrochemical biosensors capable of detecting nucleic acids in complex saliva. The constructed anti-fouling biosensor was able to detect the nucleic acid of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in undiluted saliva, with a wide linear response range (0.01 pM-0.01 µM) and a low limit of detection (LOD) of 3.4 fM (S/N = 3). The phosphorylation of oligopeptides offers an effective strategy to designing ultra-hydrophilic peptides suitable for the construction of promising anti-biofouling biosensors and bioelectronics.

The construction of elastin-like polypeptides and their applications in drug delivery system and tissue repair.

Elastin-like polypeptides (ELPs) are thermally responsive biopolymers derived from natural elastin. These peptides have a low critical solution temperature phase behavior and can be used to prepare stimuli-responsive biomaterials. Through genetic engineering, biomaterials prepared from ELPs can have unique and customizable properties. By adjusting the amino acid sequence and length of ELPs, nanostructures, such as micelles and nanofibers, can be formed. Correspondingly, ELPs have been used for improving the stability and prolonging drug-release time. Furthermore, ELPs have widespread use in tissue repair due to their biocompatibility and biodegradability. Here, this review summarizes the basic property composition of ELPs and the methods for modulating their phase transition properties, discusses the application of drug delivery system and tissue repair and clarifies the current challenges and future directions of ELPs in applications.

Antifouling Electrochemical Biosensor Based on the Designed Functional Peptide and the Electrodeposited Conducting Polymer for CTC Analysis in Human Blood.

Circulating tumor cells (CTCs) are considered reliable cancer biomarkers for the liquid biopsy of many types of tumors. The direct detection of CTCs in human blood with normal biosensors, however, remains challenging because of severe biofouling in blood that contains various proteins and a large number of cells. Herein, we report the construction of an antifouling electrochemical biosensor capable of assaying CTCs directly in blood, based on a designed multifunctional peptide and the electrodeposited conducting polymer poly(3,4-ethylenedioxythiophene) (PEDOT). The designed peptide possesses antifouling capability in complex biological media and specific recognition ability to capture breast cancer cells MCF-7. Meanwhile, electrodeposited PEDOT can promote electron transfer at the sensing interface, improve the signal-to-noise ratio for the detection, and thus enhance the sensitivity of the biosensor. The integration of the multifunctional peptide and conducting polymer PEDOT ensures that the developed biosensor is able to perform directly in blood samples without purification or separation. The antifouling electrochemical biosensor for the detection of MCF-7 cells exhibits a wide linear range over 4 orders, with a limit of detection (LOD) of 17 cells mL-1. More interestingly, even when performing in 25% human blood, the biosensor still retains a linear response with an LOD of 22 cells mL-1, without suffering significantly from biofouling in real blood. This work provides a promising strategy for the direct analysis of CTCs in human blood without a complicated pretreatment, and it may find practical application in the liquid biopsy of cancers.

DNA Computation-Modulated Self-Assembly of Stimuli-Responsive Plasmonic Nanogap Antennas for Correlated Multiplexed Molecular Imaging.

Nanogap antennas with strong electromagnetic fields of the "hot spot" in the gap region of two adjacent particles that can significantly improve the optical properties of fluorophores hold great potential for ultrasensitive bioanalysis. Herein, a DNA computation-mediated self-assembly of Au NBP dimer-based plasmonic nanogap antennas was designed for imaging of intracellular correlated dual disease biomarkers. It is worth noting that with the benefit from the electromagnetic fields of the "hot spot" in the gap region and strand displacement amplification, the fluorescence intensity can be enhanced ∼14.7-fold by Au NBP dimer-based plasmonic nanogap antennas. In addition, the AND-gate sensing mechanism was confirmed through monitoring the response of three designed nAP-PH1, m-PH1, and PH1 probes, the fluorescence recovery in different cell lines (Hela and L02), and inhibitor-treated cells, respectively. Furthermore, thanks to the "dual keys" activation design, such an "AND-gate" sensing manner can be used for ultrasensitive correlated multiplexed molecular imaging, demonstrating its feasible prospect in correlated multiplexed molecular imaging.

Antifouling Strategies for Selective In Vitro and In Vivo Sensing.

The ability to fabricate sensory systems capable of highly selective operation in complex fluid will undoubtedly underpin key future developments in healthcare. However, the abundance of (bio)molecules in these samples can significantly impede performance at the transducing interface where nonspecific adsorption (fouling) can both block specific signal (reducing sensitivity) and greatly reduce assay specificity. Herein, we aim to provide a comprehensive review discussing concepts and recent advances in the construction of antifouling sensors that are, through the use of chemical, physical, or biological engineering, capable of operating in complex sample matrix (e.g., serum). We specifically highlight a range of molecular approaches to the construction of solid sensory interfaces (planar and nanoparticulate) and their characterization and performance in diverse in vitro and in vivo analyte (e.g., proteins, nucleic acids, cells, neuronal transmitters) detection applications via derived selective optical or electrochemical strategies. We specifically highlight those sensors that are capable of detection in complex media or those based on novel architectures/approaches. Finally, we provide perspectives on future developments in this rapidly evolving field.

Electrochemical Biosensor with Enhanced Antifouling Capability Based on Amyloid-like Bovine Serum Albumin and a Conducting Polymer for Ultrasensitive Detection of Proteins in Human Serum.

Biofouling has been a substantial burden on biomarker analysis in complex biological media, leading to poor sensitivity and selectivity or even malfunction of the sensing devices. In this work, an electrochemical biosensor with excellent antifouling ability and high stability was fabricated based on amyloid-like bovine serum albumin (AL-BSA) crosslinked with the conducting polymer polyaniline (PANI). Compared with the crosslinked conventional bovine serum albumin (BSA), the crosslinked AL-BSA exhibited enhanced antifouling capability, and it was able to form an effective antifouling film within a significantly short reaction time. With further immobilization of immunoglobulin M (IgM) antibodies onto the prepared AL-BSA surface via the formation of amide bonds, an electrochemical biosensor capable of assaying IgM in human serum samples with superior selectivity and sensitivity was constructed. The biosensor exhibited excellent antifouling performance even in 100% human serum, a low limit of detection down to 2.32 pg mL-1, and acceptable accuracy for real sample analysis compared with the standard enzyme-linked immunosorbent assay for IgM detection. This strategy of using AL-BSA to construct antifouling sensing interfaces provided a reliable diagnostic method for the detection of a series of protein biomarkers in complex biological media.

Free-standing electrochemical biosensor for carcinoembryonic antigen detection based on highly stable and flexible conducting polypyrrole nanocomposite.

A flexible free-standing electrochemical biosensor to detect carcinoembryonic antigen (CEA) is described based on a conducting polypyrrole (PPy) nanocomposite film electrode. The conducting PPy composite was constructed by the sandwiched structure formed by PPy doped with pentaerythritol ethoxylate (PEE) and 2-naphthalene sulfonate (2-NS-PPy) separately via electropolymerization. Gold nanoparticles (AuNPs) were fixed on the PPy composite film by electrodeposition and then connected to CEA aptamer through self-assembly to construct a free-standing electrochemical biosensor breaking away from additional soft substrates and current collector. This PPy composite film-based electrochemical biosensor exhibits satisfying sensing performance for CEA detection, with a linear range from 10-10 g/mL to 10-6 g/mL and a detection limit of 0.033 ng/mL, good specificity and long-term sensing stability (96.8% of the original signal after 15 days). The biosensor also presents acceptable reproducibility with 1.7% relative standard deviation. Moreover, this electrochemical biosensor owns the deformation stability that could bear various deformations (twisting, folding, and knotting) without affecting device's sensing performance. It can even maintain 99.4% of the original signal under 25% strain deformation. Due to the superior sensing performance, high stability (mechanical deformation and long-term storage), and flexibility, this free-standing electrochemical biosensor proves huge potential in application of flexible and wearable electronics.

Target-triggered configuration change of DNA tetrahedron for SERS assay of microRNA 122.

A surface-enhanced Raman scattering (SERS) method is proposed for the assay of microRNA 122 based on configuration change of DNA tetrahedron. Firstly, a DNA tetrahedron was self-assembled with one vertex labeled with toluidine blue (TB). Then, it was immobilized on the porous Ni/SiO2@PEI@Au as a SERS platform, which was characterized by scanning electron microscopy (SEM) and X-ray diffraction (XRD). At this time, the DNA tetrahedron was contracted; so, the TB is close to AuNPs and the Raman signal is high. When target microRNA 122 existed, with the nicking enzyme amplification strategy, a great deal of DNA signal chains (S5) was obtained, which can extend the contracted DNA tetrahedron and change it into a three-dimensional DNA tetrahedron. In this case, the TB was far from AuNPs, resulting in a lower Raman signal. Due to the configuration change of DNA tetrahedron, the Raman signal at 1624 cm-1 (with the excitation wavelength of 633 nm) has a linear relationship with the logarithm concentration of microRNA 122. This SERS assay has high sensitivity for microRNA 122 with a determination range from 0.01 aM to 10 fM and a detection limit of 0.009 aM. The recoveries from spiked samples were in the range 95 to 109%. This SERS strategy is designed based on the target-triggered configuration change of DNA tetrahedron, which can give new insight for DNA structures in bioanalysis. Graphical abstract A sensitive surface-enhanced Raman scattering (SERS) biosensor was developed to detect microRNA 122 using the configuration change of DNA tetrahedron to indirectly control the position of TB and hot spot.

A nanocomposite consisting of MnO2 nanoflowers and the conducting polymer PEDOT for highly sensitive amperometric detection of paracetamol.

An electrochemical sensor for paracetamol is described that consists of a glassy carbon electrode (GCE) that was modified with the conducting polymer poly(3,4-ethylenedioxythiophene) (PEDOT) doped with MnO2 nanoflowers. The hydrothermally synthesized MnO2 nanoflowers possess a large surface area and can be doped into PEDOT through electrochemical deposition to form a conducting polymer nanocomposite. The nanoflowers are shown to be uniformly distributed within the nanocomposite as revealed by elemental mapping analysis. The nanocomposite displays excellent catalytic activity toward the electrochemical oxidation of paracetamol. The modified GCE, best operated at a working potential of around 0.37 V (vs. SCE) has a linear response in 0.06 to 435 µM paracetamol concentration range and a very low limit of detection (31 nM at a signal-to-noise ratio of 3). The sensor exhibits excellent reproducibility and stability, and satisfying accuracy for paracetamol detection in pharmaceutical samples. Graphical abstract A highly sensitive electrochemical sensor capable of detecting paracetamol with a limit of detection down to 31 nM was developed based on MnO2 nanoflowers doped conducting polymer PEDOT.

Polydopamine Nanosphere/Gold Nanocluster (Au NC)-Based Nanoplatform for Dual Color Simultaneous Detection of Multiple Tumor-Related MicroRNAs with DNase-I-Assisted Target Recycling Amplification.

A novel fluorescence resonance energy transfer (FRET)-based platform using polydopamine nanospheres (PDANSs) as energy acceptors and dual colored Au NCs as energy donors for simultaneous detection of multiple tumor-related microRNAs with DNase-I-assisted target recycling amplification was developed for the first time. On the basis of monitoring the change of the recovered fluorescence intensity at 445 and 575 nm upon the addition of targets miRNA-21 and let-7a, these two microRNAs (miRNAs) can be simultaneously quantitatively detected, with detection limits of 4.2 and 3.6 pM (3σ) for miRNA-21 and let-7a, which was almost 20 times lower than that without DNase I. Additionally, semiquantitative determination of miRNA-21 and let-7a can also be realized through photovisualization. Most importantly, serums from normal and breast cancer patients can be visually and directly discriminated without any sample pretreatment by confocal microscope experiments, demonstrating promising potential for auxiliary clinical diagnosis.

Antifouling and ultrasensitive biosensing interface based on self-assembled peptide and aptamer on macroporous gold for electrochemical detection of immunoglobulin E in serum.

Accurate detection of protein biomarkers in complex media remains a challenge due to severe nonspecific adsorption and biofouling, and sensing interfaces that combine the high sensitivity and antifouling ability are highly desirable. Herein, an antifouling sensing interface capable of sensitively assaying immunoglobulin E (IgE) in biological samples was constructed. The sensing interface was fabricated through the self-assembly of a zwitterionic peptide and the IgE aptamer onto a macroporous Au substrate, which was electrochemically fabricated with the aid of multilayer polystyrene nanospheres self-assembled on glassy carbon electrode. Due to the huge surface area arising from porous morphology and high specificity of aptamer, the developed electrochemical biosensor exhibits ultrahigh sensitivity and selectivity towards IgE, with the linear range of 0.1-10 pg mL-1, and a very low limit of detection down to 42 fg mL-1. Interestingly, owing to the presence of the zwitterionic peptide, the biosensing interface can satisfyingly reduce the nonspecific adsorption and fouling effect. Consequently, the biosensor was successfully applied to detect IgE in complex biological samples, indicating great promise of this peptide-based sensing interface for antifouling assays. Graphical abstract ᅟ.

Redox and label-free array detection of protein markers in human serum.

A substantial outstanding challenge in diagnostics and disease monitoring is an ability to rapidly and conveniently assay for protein biomarkers within complex biological media. Label-free electroanalytical methods present, arguably, the most promising and scalable means of achieving this but, as with all label-free assays, can struggle with response selectivity issues that arise from nonspecific surface interactions. Impedimetric methods are ultrasensitive and have been applied to the quantification of a wide range of proteins but have not previously been utilized in a multiplexed format capable of operation in complex analytical fluid. Herein, we present the use of thermally cross-linked poly(ethylene glycol) (PEG) polymer sensory array interfaces in the ultrasensitive quantification of two protein markers, insulin and C-reactive protein (CRP). This was achieved with detection limits of 171 ± 19 fM and 150 ± 10 pM, respectively. Significantly, the arrays not only enable the simultaneous, fast, nonamplified, and label-free detection of both markers without reagent addition but do so with little cross talk, even in human serum. A blind analysis of 17 real patient samples generated results in excellent agreement with those obtained through a clinically approved chemiluminescence assay.

A low-fouling electrochemical biosensor based on BSA hydrogel doped with carbon black for the detection of cortisol in human serum.

Electrochemical biosensors with high sensitivity can detect low concentrations of biomarkers, but their practical detection applications in complex biological environments such as human serum and sweat are severely limited by the biofouling. Herein, a conductive hydrogel based on bovine serum albumin (BSA) and conductive carbon black (CCB) was prepared for the construction of an antifouling biosensor. The BSA hydrogel (BSAG) was doped with CCB, and the prepared composite hydrogel exhibited good conductivity originated from the CCB and antifouling capability owing to the BSA hydrogel. An antifouling biosensor for the sensitive detection of cortisol was fabricated by drop-coating the conductive hydrogel onto a poly(3,4-ethylenedioxythiophene) (PEDOT) modified electrode and further immobilizing the cortisol aptamer. The constructed biosensor showed a linear range of 100 pg mL-1 - 10 µg mL-1 and a limit of detection of 26.0 pg mL-1 for the detection of cortisol, and it was capable of assaying cortisol accurately in complex human serum. This strategy of preparing antifouling and conductive hydrogels provides an effective way to develop robust electrochemical biosensors for biomarker detection in complex biological media.

An antifouling electrochemical biosensor based on oxidized bacterial cellulose and quaternized chitosan for reliable detection of involucrin in wound exudate.

The monitoring of biomarkers in wound exudate is of great importance for wound care and treatment, and electrochemical biosensors with high sensitivity are potentially useful for this purpose. However, conventional electrochemical biosensors always suffer from severe biofouling when performed in the complex wound exudate. Herein, an antifouling electrochemical biosensor for the detection of involucrin in wound exudate was developed based on a wound dressing, oxidized bacterial cellulose (OxBC) and quaternized chitosan (QCS) composite hydrogel. The OxBC/QCS hydrogel was prepared using an in-situ chemical oxidation and physical blending method, and the proportion of OxBC and QCS was optimized to achieve electrical neutrality and enhanced hydrophilicity, therefore endowing the hydrogel with exceptional antifouling and antimicrobial properties. The involucrin antibody SY5 was covalently bound to the OxBC/QCS hydrogel to construct the biosensor, and it demonstrated a low limit of detection down to 0.45 pg mL-1 and a linear detection range from 1.0 pg mL-1 to 1.0 µg mL-1, and it was capable of detecting targets in wound exudate. Crucially, the unique antifouling and antimicrobial capability of the OxBC/QCS hydrogel not only extends its effective lifespan but also guarantees the sensing performance of the biosensor. The successful application of this wound dressing, OxBC/QCS hydrogel for involucrin detection in wound exudate demonstrates its promising potential in wound healing monitoring.

A Super-Antifouling Electrochemical Biosensor for Protein Detection in Complex Biofluids Based on PEGylated Multifunctional Peptide.

Overcoming the influence of interfering substances in the environment and achieving superior sensing performance are significant challenges in biomarker detection within complex matrices. Herein, an integrated electrochemical sensing platform for sensitive detection of biomarkers in complex biofluids was developed based on a newly designed PEGylated multifunctional peptide (PEG-MPEP). The designed PEG-MPEP contains a poly(serine) sequence (-ssssss-) as the antifouling part and recognition peptide sequence (-avwgrwh) specific for the target human immunoglobulin G (IgG). To improve the peptide stability to protease hydrolysis, d-amino acids were adopted to synthesize the whole peptide. Additionally, the PEGylation can further enhance the stability of the peptide, and the PEG itself was also antifouling, ensuring superstrong antifouling capability of the PEG-MPEP. The designed PEG-MPEP-based biosensor possessed a high sensitivity for the detection of IgG in the range of 1.0 pg mL-1 to 1.0 µg mL-1, with a low limit of detection (0.41 pg mL-1), and it was capable of assaying targets accurately in real serum samples. Compared with conventional peptide-modified biosensors, the PEG-MPEP-modified biosensor exhibited superior antifouling and antihydrolysis properties in complex biofluid, showcasing promising potential for practical assay applications.

Enhanced split-type photoelectrochemical aptasensor incorporating a robust antifouling coating derived from four-armed polyethylene glycol.

Antifouling biosensors capable of preventing protein nonspecific adhesion in real human bodily fluids are highly sought-after for precise disease diagnosis and treatment. In this context, an enhanced split-type photoelectrochemical (PEC) aptasensor was developed incorporating a four-armed polyethylene glycol (4A-PEG) to construct a robust antifouling coating, enabling accurate and sensitive bioanalysis. The split-type PEC system involved the photoelectrode and the biocathode, effectively separating signal converter with biorecogniton events. Specifically, the TiO2 electrode underwent sequential modification with ZnIn2S4 (ZIS) and polydopamine (PDA) to form the PDA/ZIS/TiO2 photoelectrode. The cathode substrate was synthesized as a hybrid of N-doped graphene loaded with Pt nanoparticles (NG-Pt), and subsequently modified with 4A-PEG to establish a robust antifouling coating. Following the anchoring of probe DNA (pDNA) on the 4A-PEG-grafted antifouling coating, the biocathode for model target of cancer antigen 125 (CA125) was obtained. Leveraging pronounced photocurrent output of the photoelectrode and commendable antifouling characteristics of the biocathode, the split-type PEC aptasensor showcased exceptional detection performances with high sensitivity, good selectivity, antifouling ability, and potential feasibility.

Aqueous Grown Quantum Dots with Robust Near-Infrared Fluorescence for Integrated Traumatic Brain Injury Diagnosis and Surgical Monitoring.

Surgical intervention is the most common first-line treatment for severe traumatic brain injuries (TBIs) associated with high intracranial pressure, while the complexity of these surgical procedures often results in complications. Surgeons often struggle to comprehensively evaluate the TBI status, making it difficult to select the optimal intervention strategy. Here, we introduce a fluorescence imaging-based technology that uses high-quality silver indium selenide-based quantum dots (QDs) for integrated TBI diagnosis and surgical guidance. These engineered, poly(ethylene glycol)-capped QDs emit in the near-infrared region, are resistant to phagocytosis, and importantly, are ultrastable after the epitaxial growth of an aluminum-doped zinc sulfide shell in the aqueous phase that renders the QDs resistant to long-term light irradiation and complex physiological environments. We found that intravenous injection of QDs enabled both the precise diagnosis of TBI in a mouse model and, more importantly, the comprehensive evaluation of the TBI status before, during, and after an operation to distinguish intracranial from superficial hemorrhages, provide real-time monitoring of the secondary hemorrhage, and guide the decision making on the evacuation of intracranial hematomas. This QD-based diagnostic and monitoring system could ultimately complement existing clinical tools for treating TBI, which may help surgeons improve patient outcomes and avoid unnecessary procedures.