Pd@Pt nanoparticle-linked immunosorbent assay for quantification of Collagen type II.

The evaluation of specific protein content in engineered tissues provides a gateway for developing regenerative medicine treatments. Since collagen type II, the major component of articular cartilage, is critical for the blossoming field of articular cartilage tissue engineering, the interest in this protein is growing rapidly. Accordingly, the need for quantification of collagen type II is increasing as well. In this study, we provide recent results for a new quantifying nanoparticle sandwich immunoassay technique for collagen type II. Since mesoporous palladium@platinum (Pd@Pt) nanoparticles have peroxidase-like catalytic activities, these nanoparticles were utilized in an enzyme-linked immunosorbent assay (ELISA)-like format to circumvent the need for traditional enzymes. These nanoparticles were easily conjugated with anti-collagen type II antibodies by the natural affinity interaction and used to develop a direct sandwich ELISA-like format for nanoparticle-linked immunosorbent assays. Using this method, we obtained a limit of detection of 1 ng mL-1, a limit of quantification of 9 ng mL-1. and a broad linear range between 1 ng mL-1 and 50 µg mL-1 for collagen type II with an average relative standard deviation of 5.5%, useable over a pH range of 7 - 9 at least. The assay was successfully applied to quantify collagen type II in cartilage tissues and compared with the results of commercial ELISAs and gene expression by reverse transcription-quantitative polymerase chain reaction. This method provides a thermally stable and cost-efficient alternative to traditional ELISAs. It also extends the application of nanoparticle-linked immunosorbent assays, thereby providing the potential to quantify other proteins and apply the technology in the medical, environmental, and biotechnology industry fields.

Simultaneous detection of two herbicides in fruits and vegetables with nanoparticle-linked immunosorbent and lateral flow immunoassays.

Herbicides atrazine and acetochlor are used in crop production. Because of environmental and health hazards with respective maximum contamination levels of 3 and 20 ng/mL, quantifying these herbicides is important when considering presence in foods and vegetables. We utilized two Pd@Pt nanoparticle-amplified immunoassays, a colorimetric Pd@Pt nanoparticle-linked immunosorbent assay (NLISA) and differential pulse voltammetry (DPV) dependent on catalytic activity of Pd@Pt in a dual-lateral flow immunoassay (dual-LFIA-DPV). We achieved overall recoveries of 88.5-114 % in juice, fruit, and vegetable samples for both immunoassays. The NLISA yielded limits of detection (LODs) of 0.59 and 0.31 µg/kg and the dual-LFIA-DPV 0.27 and 0.51 µg/kg for the two respective species. Results for both immunoassays were validated by high-performance liquid chromatography (HPLC), for all food and drink samples though LODs are compromised when configuring the HPLC for both species with the same chromatogram. We expect Pd@Pt-based immunoassays to prove useful in various fields.

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.

Au@PtPd enhanced immunoassay with 3D printed smartphone device for quantification of diaminochlorotriazine (DACT), the major atrazine biomarker.

Increased use of pesticides in agriculture requires new advanced techniques to monitor both environmental levels and human exposure of pesticides to avoid potential adverse health outcomes in sensitive populations. Atrazine is widely used to control broadleaf weeds, and here we developed a new sensor capable of detecting diaminochlorotriazine (DACT), the major metabolite and biomarker of atrazine exposure. We established an Au@PtPd nanoparticles labeled lateral flow immunoassay (LFIA) for immunochromatographic based rapid detection of urinary DACT. The detection was based on competitive immunoassay between the analyte and the BSA-conjugated antigen. As evaluated, the coupled mesoporous core-shell Au@PtPd nanoparticles, with superior peroxidase-like activity, as the signal indicator offers a rapid direct chromatographic readout inversely correlated with the concentration of analytes, providing a detection limit of 0.7 ng/mL for DACT. Moreover, the detection limits were boosted to as low as 11 pg/mL with the detectable range from 10 pg/ml to 10 ng/mL, through a one-step catalytic chromogenic reaction. A rapid readout device was developed by 3D printing to provide a stable real-time quantification of the color intensity capable of assessing both chromatographic and absorbance results. This Au@PtPd nanoparticle-based immunosensing platform, as well as the 3D printed readout device, provide a promising tool for on-site and ultrasensitive detection of pesticide biomarkers.

Smartphone-Based Dual-Channel Immunochromatographic Test Strip with Polymer Quantum Dot Labels for Simultaneous Detection of Cypermethrin and 3-Phenoxybenzoic Acid.

Currently, gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-MS (LC-MS) are the primary methods used to detect pesticides and their metabolites for biomonitoring of exposure. Although GC-MS and LC-MS can provide accurate and sensitive measurements, these techniques are not suitable for point-of-care or in-field biomonitoring applications. The objective of this work is to develop a smartphone-based dual-channel immunochromatographic test strip (ICTS) for on-site biomonitoring of exposure to cypermethrin by simultaneous detection of cypermethrin and its metabolite, 3-phenoxybenzoic acid (3-PBA). Polymer carbon dots (PCDs) with ultrahigh fluorescent brightness were synthesized and used as a signal amplifier in ICTS assay. Cypermethrin (a representative pyrethroid pesticide) and its major metabolite 3-PBA were simultaneously detected to provide more comprehensive analysis of cypermethrin exposure. After competitive immunoreactions between the target sample and the coating antigens preloaded on the test line, the tracer antibody (PCD-conjugated antibody) was quantitatively captured on the test lines. The captured PCDs were inversely proportional to the amount of the target compound in the sample. The red fluorescence on the test line was then recorded using a smartphone-based device capable of conducting image analysis and recording. Under optimal conditions, the sensor showed excellent linear responses for detecting cypermethrin and 3-PBA ranging from 1 to 100 ng/mL and from 0.1 to 100 ng/mL, respectively, and the limits of detection were calculated to be ∼0.35 ng/mL for cypermethrin and ∼0.04 ng/mL for 3-PBA. The results demonstrate that the ICTS device is promising for accurate point-of-care biomonitoring of pesticide exposure.

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.

Nanomaterial-enhanced 3D-printed sensor platform for simultaneous detection of atrazine and acetochlor.

The widespread use of herbicides in agriculture and gardening causes environmental and safety issues such as water pollution. Thus, efficient and convenient analysis of the levels of herbicide residues is of significant importance. Here, we employed 3D-printing to design a multiplex immunosensor for simultaneous detection of two widely used herbicides, atrazine and acetochlor. Multiplexing was achieved through customization of a lateral flow immunoassay, and then integrated with an electrochemical analyzer for ultrasensitive detection. Quantification of herbicide residues was realized through the detection of a novel nanomaterial label, the mesoporous core-shell palladium@platium nanoparticle (Pd@Pt NP), for its outstanding peroxidase-like property. During the electrochemical analysis, the catalytic activity of Pd@Pt NPs on the redox reaction between thionin acetate and hydrogen peroxide provided an electrochemically driven signal that accurately indicated the level of herbicide residues. Using this Nanomaterial-enhanced multiplex electrochemical immunosensing (NEMEIS) system, simultaneous detection of atrazine and acetochlor was realized with a limit of detection of 0.24 ppb and 3.2 ppb, respectively. To further evaluate the feasibility, the optimized NEMEIS was employed for detection in atrazine and acetochlor residue-containing spiked samples, and an overall recovery with 90.8% - 117% range was obtained. The NEMEIS constructed with the aid of 3D-printing provides a rapid, precise, economical, and portable detection device for herbicides, and its success suggests potential broad applications in chemical analysis, biosensors and point-of-care monitoring.

Metal-organic framework based nanozymes: promising materials for biochemical analysis.

In recent years, there has been rapid growth of enzyme-mimicking catalytic nanomaterials (nanozymes). Compared with biological enzymes, nanozymes exhibit several superiorities, including robust activity, easy production, and low cost, which endow them with promising applications in biochemical analysis. As an emerging member of nanozymes, metal-organic framework (MOF) nanozymes are attracting growing attention because of their composition and structural characteristics. Rationally designing MOFs with enzyme-like catalytic ability is opening up a new avenue for biochemical detection. In this Feature Article, we summarize the latest developments of MOF nanozymes and their applications in biochemical sensing. First, the types of nanozymes derived from MOFs are categorized, and effective strategies to improve the weak activity inherent in MOF nanozymes are introduced. Then, the multi-functionalization of MOFs with enzyme mimic activity and other attractive properties is emphasized. After that, the typical applications of MOF nanozymes in the detection of various analytes are rigorously reviewed. Finally, the current challenges and some development directions in this field are discussed. It is believed that the versatile nature of MOFs will bring a bright future for MOF nanozymes in biochemical analysis.

Mesoporous Pd@Pt nanoparticle-linked immunosorbent assay for detection of atrazine.

Atrazine is a widely used herbicide in the United States; however, the Environmental Protection Agency (EPA) has issued warnings about atrazine because of its reported potential harmful effects on animals and humans. Therefore, developing efficient ways to detect this herbicide's residue are critically important. The competitive ELISA is a useful method for detecting chemicals for which antibodies exist due to its high sensitivity, specificity, and efficiency. However, the assay typically requires a separate application of a secondary antibody linked to an enzyme that catalyzes conversion of a non-colored organic to a detectable colored product. In this study, we used the recently developed peroxidase-like mesoporous core-shell palladium@platinum (Pd@Pt) nanoparticle which can easily be bound directly to primary antibody, thereby eliminating the need for a secondary antibody conjugate. We report a first instance in which this technique is applied for use in a competitive assay for small molecules, in this case the herbicide atrazine. Due to their high-surface area and mesoporous structure, Pd@Pt nanoparticles enable fast mass transfer for reaction with excellent catalytic activity. This leads to high sensitivity in our immunoassay with a limit of detection of 0.5 ng mL-1 defined by selecting an IC10 concentration, i.e., the analyte concentration at which 10% of the available Pd@Pt nanoparticle-labeled antibody is inhibited from binding to a plate coated with a bovine serum albumin-atrazine conjugate. We applied our method to well-water and pond water samples spiked with atrazine. Our tests at 5, 10, and 20 ng mL-1 yielded recoveries of 99 - 115%, offering strong supporting evidence that atrazine and other low molecular weight herbicides and pesticides can be detected using this immunoassay approach. Detection with this method is expected to lead to its use in a wide spectrum of applications in agriculture, medical, and biotechnology arenas.

2D Graphene Oxide/Fe-MOF Nanozyme Nest with Superior Peroxidase-Like Activity and Its Application for Detection of Woodsmoke Exposure Biomarker.

Emerging nanomaterials such as nanozymes have recently been applied for the immunoassay-based detection of biomarkers. However, the inferior catalytic activity and low water solubility of nanozymes remain as the major limitations compared to natural enzymes. To overcome these limitations, we successfully synthesized a superior nanozyme with a structure of enriched 2D catalytic interface, namely Nanozyme Nest, which was composed of Fe-based metal-organic frameworks (Fe-MOF) and graphene oxide (GO). Then, we applied it in an ultrasensitive enzyme-linked immunosorbent assay (ELISA) for the detection of benzo[a]pyrene-7,8-diol 9,10-epoxide-DNA adduct (BPDE-DNA), which is a metabolite of benzo[a]pyrene (BP) and used as a typical biomarker of woodsmoke exposure in human blood. The Nanozyme Nest features amplified peroxidase-like catalytic ability from graphene and Fe-MOF due to their large surface area and abundant active sites. By using the proposed Nanozyme Nest-based ultrasensitive ELISA, the BPDE-DNA could be detected at a level as low as 0.268 ng/mL, and the obtained sensitivity was much higher than most of the widely used methods. Our work provides a novel strategy to design ultrasensitive immunosensors with advantages of amplified catalytic activity and improved water solubility compared to classic nanozymes. This illustrates the promising applications of the Nanozyme Nest-based immunosensors in point-of-care settings to conveniently detect exposures and diagnose diseases.

Micro additive manufacturing of glucose biosensors: A feasibility study.

Flexible electrochemical sensors for measurement and quantification of biomarkers are attracting a great deal of attention in non-invasive medical applications, due to their high mechanical compatibility and conformability with the human body. Realization of the full potential of such novel systems relies heavily on their effective manufacturing. Particularly, there is a need for manufacturing techniques that can realize complex designs, consisting of multiple functional materials which are required for sensor functionality. Among emerging additive manufacturing techniques, Direct-Ink-Writing (DIW), where polymer nanocomposite inks are dispensed through nozzles and deposited with high spatial control, carries a great potential to address this need. Here, we introduce a 3D printed flexible electrochemical biosensor for glucose detection. We show that our biosensor works linearly in glucose solution with a concentration range between 100 and 1000 µM. The sensitivity of glucose biosensor is estimated to be 17.5 nA µM-1, and the calculated value of the detection limit (S/N = 3) is 6.9 µM. The demonstrated electrochemical performance and surface properties of the printed sensors show the promising advantages of using this technique over the conventional screen printing method. These advantages include higher sensitivity and specificity and, reduced material consumption.

Fabrication of a SnO2-based acetone gas sensor enhanced by molecular imprinting.

This work presents a new route to design a highly sensitive SnO2-based sensor for acetone gas enhanced by the molecular imprinting technique. Unassisted and acetone-assisted thermal synthesis methods are used to synthesis SnO2 nanomaterials. The prepared SnO2 nanomaterials have been characterized by X-ray powder diffraction, scanning electron microscopy and N2 adsorption-desorption. Four types of SnO2 films were obtained by mixing pure deionized water and liquid acetone with the two types of as-prepared powders, respectively. The acetone gas sensing properties of sensors coated by these films were evaluated. Testing results reveal that the sensor coated by the film fabricated by mixing liquid acetone with the SnO2 nanomaterial synthesized by the acetone-assisted thermal method exhibits the best acetone gas sensing performance. The sensor is optimized for the smooth adsorption and desorption of acetone gas thanks to the participation of acetone both in the procedure of synthesis of the SnO2 nanomaterial and the device fabrication, which results in a distinct response-recovery behavior.