Single-Atomic Site Catalyst with Heme Enzymes-Like Active Sites for Electrochemical Sensing of Hydrogen Peroxide.

Heme enzymes, with the pentacoordinate heme iron active sites, possess high catalytic activity and selectivity in biosensing applications. However, they are still subject to limited catalytic stability in the complex environment and high cost for broad applications in electrochemical sensing. It is meaningful to develop a novel substitute that has a similar structure to some heme enzymes and mimics their enzyme activities. One emerging strategy is to design the Fe-N-C based single-atomic site catalysts (SASCs). The obtained atomically dispersed Fe-Nx active sites can mimic the active sites of heme enzymes effectively. In this work, a SASC (Fe-SASC/NW) is synthesized by doping single iron atoms in polypyrrole (PPy) derived carbon nanowire via a zinc-atom-assisted method. The proposed Fe-SASC/NW shows high heme enzyme-like catalytic performance for hydrogen peroxide (H2 O2 ) with a specific activity of 42.8 U mg-1 . An electrochemical sensor based on Fe-SASC/NW is developed for the detection of H2 O2 . This sensor exhibits a wide detection concentration range from 5.0 × 10-10 m to 0.5 m and an excellent limit of detection (LOD) of 46.35 × 10-9 m. Such excellent catalytic activity and electrochemical sensing sensitivity are attributed to the isolated Fe-Nx active sites and their structural similarity with natural metalloproteases.

Direct Cytosolic MicroRNA Detection Using Single-Layer Perfluorinated Tungsten Diselenide Nanoplatform.

Intracellular miRNA detection is vitally important for diagnosing severe diseases like cancer and for resolving the ensemble of gene products that orchestrate the living state of cells. Recent advances in the design, synthesis, and application of biocompatible nanomaterials as platforms for probing miRNAs have enabled widespread efforts to mobilize new compounds in biomedical research. Two-dimensional graphene-like nanomaterials exhibit desirable physical properties such as convenient quantum size and dynamic interface functionality. Because miRNAs regulate gene expression in the cytoplasm, it is imperative that nanomaterials targeting them are properly delivered. Unloading of nanomaterials into the cytosol using the cellular endocytic transport pathway is often hindered by an inability to cross the endosomal membrane. To address this challenge, we designed a strategy to deliver functionalized WSe2 nanosheets (FWNs) to the cytosol using perfluorinated surface functionalization. Perfluorinated compounds are both hydrophobic and lipophobic, exhibiting excellent phase-separation tendency in both polar and nonpolar environments. FWNs are ∼120 nm in diameter, feature low toxicity, and exhibit excellent stability in serum. The fluorinated nanostructure of FWNs enabled efficient cytosolic delivery from the endomembrane system. The fidelity of this approach was confirmed through intracellular delivery of two DNA probes (ssDNA-21 and ssDNA-210), which resulted in specific labeling of cytosolic miRNA and demonstrated the utility of this system for direct cytosolic biosensing.

Efficient Cytosolic Delivery Using Crystalline Nanoflowers Assembled from Fluorinated Peptoids.

The design and synthesis of biocompatible nanomaterials as cargoes for the intracellular delivery of therapeutic proteins or genes have attracted intense attention because of their potential for use in therapeutics. Despite the advances in this area, very few nanomaterials can be efficiently delivered to the cytosol. To address these challenges, crystalline nanoflower-like particles are designed and synthesized from fluorinated sequence-defined peptoids; the crystallinity and fluorination of these particles enable highly efficient cytosolic delivery with minimal cytotoxicity. A cytosol delivery rate of 80% has been achieved for the fluorinated peptoid nanoflowers. Furthermore, these nanocrystals can carry therapeutic genes, such as mRNA and effectively deliver the payload into the cytosol, demonstrating the universal delivery capability of the nanocrystals. The results indicate that self-assembly of crystalline nanomaterials from fluorinated peptoids paves a new way toward development of nanocargoes with efficient cytosolic gene delivery capability.

Model-Based Angular Scan Error Correction of an Electrothermally-Actuated MEMS Mirror.

In this paper, the actuation behavior of a two-axis electrothermal MEMS (Microelectromechanical Systems) mirror typically used in miniature optical scanning probes and optical switches is investigated. The MEMS mirror consists of four thermal bimorph actuators symmetrically located at the four sides of a central mirror plate. Experiments show that an actuation characteristics difference of as much as 4.0% exists among the four actuators due to process variations, which leads to an average angular scan error of 0.03°. A mathematical model between the actuator input voltage and the mirror-plate position has been developed to predict the actuation behavior of the mirror. It is a four-input, four-output model that takes into account the thermal-mechanical coupling and the differences among the four actuators; the vertical positions of the ends of the four actuators are also monitored. Based on this model, an open-loop control method is established to achieve accurate angular scanning. This model-based open loop control has been experimentally verified and is useful for the accurate control of the mirror. With this control method, the precise actuation of the mirror solely depends on the model prediction and does not need the real-time mirror position monitoring and feedback, greatly simplifying the MEMS control system.

Single-Atomic Site Catalyst Enhanced Lateral Flow Immunoassay for Point-of-Care Detection of Herbicide.

Point-of-care (POC) detection of herbicides is of great importance due to their impact on the environment and potential risks to human health. Here, we design a single-atomic site catalyst (SASC) with excellent peroxidase-like (POD-like) catalytic activity, which enhances the detection performance of corresponding lateral flow immunoassay (LFIA). The iron single-atomic site catalyst (Fe-SASC) is synthesized from hemin-doped ZIF-8, creating active sites that mimic the Fe active center coordination environment of natural enzyme and their functions. Due to its atomically dispersed iron active sites that result in maximum utilization of active metal atoms, the Fe-SASC exhibits superior POD-like activity, which has great potential to replace its natural counterparts. Also, the catalytic mechanism of Fe-SASC is systematically investigated. Utilizing its outstanding catalytic activity, the Fe-SASC is used as label to construct LFIA (Fe-SASC-LFIA) for herbicide detection. The 2,4-dichlorophenoxyacetic acid (2,4-D) is selected as a target here, since it is a commonly used herbicide as well as a biomarker for herbicide exposure evaluation. A linear detection range of 1-250 ng/mL with a low limit of detection (LOD) of 0.82 ng/mL has been achieved. Meanwhile, excellent specificity and selectivity towards 2,4-D have been obtained. The outstanding detection performance of the Fe-SASC-LFIA has also been demonstrated in the detection of human urine samples, indicating the practicability of this POC detection platform for analyzing the 2,4-D exposure level of a person. We believe this proposed Fe-SASC-LFIA has potential as a portable, rapid, and high-sensitive POC detection strategy for pesticide exposure evaluation.

Molecularly imprinted polypyrrole nanotubes based electrochemical sensor for glyphosate detection.

An electrochemical sensor based on molecularly imprinted polypyrrole nanotubes (MIPNs) has been developed for the detection of glyphosate (Gly) with high sensitivity and specificity. Herein, the MIPNs are prepared by imprinting Gly sites on the surface of polypyrrole (PPy) nanotubes. The synthesized MIPNs have high electrical conductivity and exhibit rapid adsorption rate, enhanced affinity and specificity to Gly. An electrochemical sensor for Gly detection is fabricated by assembling MIPNs-modified screen-printed electrodes with a 3D-printed electrode holder, which is highly portable and suitable for real-time detection. The results demonstrate that the MIPNs-based electrochemical sensor for Gly exhibits a wide detection range of 2.5-350 ng/mL with a limit of detection (LOD) of 1.94 ng/mL. Besides, the Gly sensor possessed good stability, reproducibility, and excellent selectivity against other interferents. The practicability of the sensor is verified by detecting Gly in orange juice and rice beverages, indicating that the sensor is suitable for monitoring pesticides in actual food and environmental samples.

CdTe@SiO2 signal reporters-based fluorescent immunosensor for quantitative detection of prostate specific antigen.

In this paper, an immunosensor using CdTe@SiO2 core-shell nanoparticles as labels was constructed for highly sensitive detection of prostate-specific antigen (PSA). In this approach, CdTe@SiO2 core-shell nanoparticles were synthesized using the sol-gel method. The additional Cd ions and sulfur source in SiO2 shell can greatly enhance the fluorescence intensity of CdTe nanocrystals. The reason is the formation of CdS-like cluster in SiO2 shell, which reduced the quantum size effect. The obtained CdTe@SiO2 nanoparticles also exhibited excellent biocompatibility, which was ideal for applying in biomarker detection. Furthermore, PSA capture antibodies functionalized magnetic Fe3O4 nanoparticles (Fe3O4-Ab1) were utilized in the proposed immunosensor to capture and enrich the PSA. The captured PSA was then immuno-recognized by CdTe@SiO2 labeled with PSA detection antibodies (CdTe@SiO2-Ab2) by forming the sandwich complex Fe3O4-Ab1/PSA/Ab2-CdTe@SiO2. The construction of this immunosensor was confirmed by fluorescence spectroscopy. The proposed immunosensor showed a good linear relationship between the fluorescent intensity and the target PSA concentration ranging from 0.01 to 5 ng/mL, and a detection limit as low as 0.003 ng/mL was achieved. The sensor also exhibited good specificity to PSA. This highly sensitive and specific immunosensor has great potential to be used in other biological detection.

Pt-Ni(OH)2 nanosheets amplified two-way lateral flow immunoassays with smartphone readout for quantification of pesticides.

Excessive use of herbicide and insecticide causes bioaccumulation in the environment and increases potential toxicity for people and animals. Portable systems for rapid assays of herbicide and insecticide residues have attracted prominent interests. Here, we developed a two-dimensional (2D) Pt-Ni(OH)2 nanosheets (NSs) amplified two-way lateral flow immunoassay (LFI) with a smartphone-based readout for simultaneous detection of acetochlor and fenpropathrin. The 2D Pt-Ni(OH)2 NSs were synthesized and used as the enhanced signal label in the immunoassay due to their high peroxidase-like activity and low migration speed. The two-way LFI was designed to eliminate potential cross-reaction between two targets. Portable detection system was developed based on a smartphone-based readout, which scans the LFI and provides the accurate testing result. The universal use of smartphones makes the developed platform suitable for cheap and on-site applications. Using the integrated platform, detection of acetochlor and fenpropathrin simultaneously was successfully achieved with the detection limits of 0.63 ng/mL and 0.24 ng/mL, respectively. To confirm the performance of the on-site application, we detected 10 non-spiked samples and 3 spiked samples. The obtained detection results were consistent with the data from gas chromatography analysis. The estimated recoveries ranged from 97.12% to 111.46%, indicating the practical reliability of our developed assay. The developed smartphone-based platform exhibits enhanced sensitivity, which provides a promising technique for on-site, multiplex, highly sensitive detection of pesticides.

Visualization of endogenous hydrogen sulfide in living cells based on Au nanorods@silica enhanced fluorescence.

Hydrogen sulfide as a gas indicator molecule plays an important role in various human physiological processes. However, due to the high volatility and diffusivity of H2S in biological systems, it is very difficult to implement a precise assay for H2S detection. Compared with the destructive instrumental methods, assays based on fluorescence probes provide noninvasive and real-time detections of H2S in living cells. In this work, we presented a fluorescent nanoprobe based on dye-functionalized Au nanorods (NRs)@silica for sensitive and selective detection of H2S in vitro and living cells. With the metal enhanced fluorescence effect, the fluorescence turn-on and turn-off were controlled by the formation and disassembly of coordination compound between dyes and copper ions. Silica matrix was used to coat the Au NRs to prevent them from the biological cytotoxicity. The effects of the different distances between Au NRs and fluorophores on fluorescent enhancement were explored and approximately 5-fold fluorescence enhancement was obtained with a distance of 22 nm. A detection of limit of 17 nM was achieved. In addition, visualization of exogenous and endogenous H2S in living cells was validated.

Unprecedented peroxidase-mimicking activity of single-atom nanozyme with atomically dispersed Fe-Nx moieties hosted by MOF derived porous carbon.

Due to robustness, easy large-scale preparation and low cost, nanomaterials with enzyme-like characteristics (defined as 'nanozymes') are attracting increasing interest for various applications. However, most of currently developed nanozymes show much lower activity in comparison with natural enzymes, and the deficiency greatly hinders their use in sensing and biomedicine. Single-atom catalysts (SACs) offer the unique feature of maximum atomic utilization, providing a potential pathway to improve the catalytic activity of nanozymes. Herein, we propose a Fe-N-C single-atom nanozyme (SAN) that exhibits unprecedented peroxidase-mimicking activity. The SAN consists of atomically dispersed Fe─Nx moieties hosted by metal-organic frameworks (MOF) derived porous carbon. Thanks to the 100% single-atom active Fe dispersion and the large surface area of the porous support, the Fe-N-C SAN provided a specific activity of 57.76 U mg-1, which was almost at the same level as natural horseradish peroxidase (HRP). Attractively, the SAN presented much better storage stability and robustness against harsh environments. As a proof-of-concept application, highly sensitive biosensing of butyrylcholinesterase (BChE) activity using the Fe-N-C SAN as a substitute for natural HRP was further verified.

Highly photoluminescent carbon dots derived from linseed and their applications in cellular imaging and sensing.

Carbon dots (CDs) synthesized from natural organic precursors, such as glucose, citric acid, glycerol, and chitosan, have attracted great interest since natural organic precursors provide abundant carbon sources, a variety of heteroatoms for doping (such as N, S, and P) and good biocompatibility. However, previous approaches utilized organic solvents during synthesis procedures, which limited their widespread development in biomedical applications. Herein, the facile synthesis of a new type of bright CDs through an eco-friendly method that employs linseed as a natural precursor has been reported. The as-obtained CDs possessed high quantum yield of 14.2%, excellent solubility and photostability as well as excitation-dependent photoluminescence (PL). In addition, the as-prepared CDs exhibited great potential in cell imaging owing to negligible cytotoxicity as well as excellent biocompatibility and great resistance to photobleaching. Subsequently, the as-prepared CDs were also applied in the fabrication of a biosensor for sensitive detection of butyrylcholinesterase (BChE) based on the fluorescence quenching mechanism, which could be used as an indicator for detecting pesticides and nerve gases. By monitoring the change in the fluorescence intensity of the CDs, the activity of BChE was sensitively analyzed. The limit of detection (LOD) of BChE was 0.035 mU mL-1. The as-prepared CDs have potential applications in both biosensors and bio-imaging.

Synthetic Polymer Nanoparticles Functionalized with Different Ligands for Receptor-mediated Transcytosis across Blood-Brain Barrier.

Polymeric nanoparticles have been investigated as biocompatible and promising nano-carriers to deliver drugs across the blood-brain barrier (BBB). However, most of the polymeric nanoparticles cannot be observed without attaching them with fluorescent dyes. Generally complex synthesis process is required to attach fluorescent dye tracing molecules with drug carrier nanoparticles. In this paper, we synthesized a novel fluorescent polymer based on poly [Triphenylamine-4-vinyl-(P-methoxy-benzene)] (TEB). This polymer was prepared from TEB polymer through coprecipitation. Furthermore, three types of ligands, transferrin (TfR), lactoferrin (LfR) and lipoprotein (LRP), were covalently attached on the nanoparticle surface to improve the BBB transport efficiency. All of prepared TEB-based nanoparticles were biocompatible, exhibited excellent fluorescence properties and could be observed in vivo. The transcellular transportation of these TEB-based nanoparticles across the BBB was evaluated by observing the fluorescent intensity. The translocation study was performed in an in vitro BBB model that were constructed based on mouse cerebral endothelial cells (bEnd.3). The results showed that ligand-coated TEB nanoparticles can be transported across BBB with high efficiencies (up to 29.02%). This is the first time that the fluorescent TEB nanoparticles were applied as nano-carriers for transport across the BBB. Such fluorescent polymeric nanoparticles have the potential applications in brain imaging or drug delivery.

Magnetostrictive Microcantilever as an Advanced Transducer for Biosensors.

The magnetostrictive microcantilever (MSMC) as a high-performance transducer was introduced for the development of biosensors. The principle and characterization of MSMC are presented. The MSMC is wireless and can be easily actuated and sensed using magnetic field/signal. More importantly, the MSMC exhibits a high Q value and works well in liquid. The resonance behavior of MSMC is characterized in air at different pressures and in different liquids, respectively. It is found that the Q value of the MSMC in water reaches about 40. Although the density and viscosity of the surrounding media affect the resonance frequency and the Q value of MSMC, the density has a stronger influence on the resonance frequency and the viscosity has a stronger influence on the Q value, which result in that, for MSMC in air at pressure of less than 100 Pa, the resonance frequency of MSMC is almost independent of the pressure, while the Q value increases with decreasing pressure. MSMC array was developed and characterized. It is experimentally demonstrated that the characterization of an MSMC array is as simple as the characterization of a single MSMC. A filamentous phage against Salmonella typhimurium was utilized as bio-recognition unit to develop an MSMC based biosensor. The detection of S. typhimurium in water demonstrated that the MSMC works well in liquid.

A surface-scanning coil detector for real-time, in-situ detection of bacteria on fresh food surfaces.

Proof-in-principle of a new surface-scanning coil detector has been demonstrated. This new coil detector excites and measures the resonant frequency of free-standing magnetoelastic (ME) biosensors that may now be placed outside the coil boundaries. With this coil design, the biosensors are no longer required to be placed inside the coil before frequency measurement. Hence, this new coil enables bacterial pathogens to be detected on fresh food surfaces in real-time and in-situ. The new coil measurement technique was demonstrated using an E2 phage-coated ME biosensor to detect Salmonella typhimurium on tomato surfaces. Real-time, in-situ detection was achieved with a limit of detection (LOD) statistically determined to be lower than 1.5×10(3) CFU/mm(2) with a confidence level of difference higher than 95% (p<0.05).

Rapid and sensitive detection of Salmonella Typhimurium on eggshells by using wireless biosensors.

This article presents rapid, sensitive, direct detection of Salmonella Typhimurium on eggshells by using wireless magnetoelastic (ME) biosensors. The biosensor consists of a freestanding, strip-shaped ME resonator as the signal transducer and the E2 phage as the biomolecular recognition element that selectively binds with Salmonella Typhimurium. This ME biosensor is a type of mass-sensitive biosensor that can be wirelessly actuated into mechanical resonance by an externally applied timevarying magnetic field. When the biosensor binds with Salmonella Typhimurium, the mass of the sensor increases, resulting in a decrease in the sensor's resonant frequency. Multiple E2 phage-coated biosensors (measurement sensors) were placed on eggshells spiked with Salmonella Typhimurium of various concentrations (1.6 to 1.6 × 10(7) CFU/cm(2)). Control sensors without phage were also used to compensate for environmental effects and nonspecific binding. After 20 min in a humidity-controlled chamber (95%) to allow binding of the bacteria to the sensors to occur, the resonant frequency of the sensors was wirelessly measured and compared with their initial resonant frequency. The resonant frequency change of the measurement sensors was found to be statistically different from that of the control sensors down to 1.6 × 10(2) CFU/cm(2), the detection limit for this work. In addition, scanning electron microscopy imaging verified that the measured resonant frequency changes were directly related to the number of bound cells on the sensor surface. The total assay time of the presented methodology was approximately 30 min, facilitating rapid detection of Salmonella Typhimurium without any preceding sampling procedures.

Effects of surface functionalization on the surface phage coverage and the subsequent performance of phage-immobilized magnetoelastic biosensors.

One of the important applications for which phage-immobilized magnetoelastic (ME) biosensors are being developed is the wireless, on-site detection of pathogenic bacteria for food safety and bio-security. Until now, such biosensors have been constructed by immobilizing a landscape phage probe on gold-coated ME resonators via physical adsorption. Although the physical adsorption method is simple, the immobilization stability and surface coverage of phage probes on differently functionalized sensor surfaces need to be evaluated as a potential way to enhance the detection capabilities of the biosensors. As a model study, a filamentous fd-tet phage that specifically binds streptavidin was adsorbed on either bare or surface-functionalized gold-coated ME resonators. The surface functionalization was performed through the formation of three self-assembled monolayers with a different terminator, based on the sulfur-gold chemistry: AC (activated carboxy-terminated), ALD (aldehyde-terminated), and MT (methyl-terminated). The results, obtained by atomic force microscopy, showed that surface functionalization has a large effect on the surface phage coverage (46.8%, 49.4%, 4.2%, and 5.2% for bare, AC-, ALD-, and MT-functionalized resonators, respectively). In addition, a direct correlation of the observed surface phage coverage with the quantity of subsequently captured streptavidin-coated microbeads was found by scanning electron microscopy and by resonance frequency measurements of the biosensors. The differences in surface phage coverage on the differently functionalized surfaces may then be used to pattern the phage probe layer onto desired parts of the sensor surface to enhance the detection capabilities of ME biosensors.

Direct detection of Salmonella typhimurium on fresh produce using phage-based magnetoelastic biosensors.

Current bacterial detection methods require the collection of samples followed by preparation and analysis in the laboratory, both time and labour consuming steps. More importantly, because of cost, only a limited number of samples can be taken and analyzed. This paper presents the results of an investigation to directly detect Salmonella typhimurium on fresh tomato surfaces using phage-based magnetoelastic (ME) biosensors. The biosensor is composed of a ME resonator platform coated with filamentous E2 phage, engineered to bind with S. typhimurium. The ME biosensors are wireless sensors, whose resonance oscillation and resonance frequency are actuated and detected through magnetic fields. The sensors used in this study were 0.028 mm×0.2 mm×1 mm in size. In this study, the tomato surface was spiked with S. typhimurium suspensions with concentrations ranging from 5×10(1) to 5×10(8)CFU/ml and then allowed to dry in air. The detection was conducted by directly placing ME measurement biosensors and control sensors on the spiked surface for 30 min in a humid environment. The control sensors were identical to the measurement biosensors, but without phage. Both measurement and control sensors were blocked with BSA to reduce non-specific binding. The resonance frequencies of both measurement and control sensors were measured prior to and after the placement of the sensors on the tomato. Shifts in the resonance frequency of the measurement biosensors were observed, while the control sensors showed negligible change. Scanning electron microscopy (SEM) was used to verify the specific binding of S. typhimurium to the biosensor. Results of multiple biosensor detection and corresponding analyzes showed statistically different responses between the measurement and control sensors for tomatoes spiked with S. typhimurium suspensions with concentrations of 5×10(2)CFU/ml and greater. This study demonstrates the direct detection of food-borne bacteria on fresh produce.