Quantitative Detection of Multiple Cardiovascular Biomarkers by an Antibody Microarray-Based Metal-Enhanced Fluorescence Assay.

Accurate detection of multiple cardiovascular biomarkers is crucial for the timely screening of acute coronary syndrome (ACS) and differential diagnosis from acute aortic syndrome (AAS). Herein, an antibody microarray-based metal-enhanced fluorescence assay (AMMEFA) has been developed to quantitatively detect 7 cardiovascular biomarkers through the formation of a sandwich immunoassay on the poly(glycidyl methacrylate-co-2-hydroxyethyl methacrylate)-decorated GNR-modified slide (GNR@P(GMA-HEMA) slide). The AMMEFA exhibits high specificity and sensitivity, the linear ranges span 5 orders of magnitude, and the limits of detection (LODs) of cardiac troponin I (cTnI), heart-type fatty acid binding protein (H-FABP), C-reactive protein (CRP), copeptin, myoglobin, D-Dimer, and N-terminal pro-brain natriuretic peptide (NT-proBNP) reach 0.07, 0.2, 65.7, 0.6, 0.2, 8.3, and 0.3 pg mL-1, respectively. To demonstrate its practicability, the AMMEFA has been applied to quantitatively analyze 7 cardiovascular biomarkers in 140 clinical plasma samples. In addition, the expression levels of cardiovascular biomarkers were analyzed by the least absolute shrinkage and selector operator (LASSO) regression, and the area under receiver operator characteristic curves (AUCs) of healthy donors (HDs), ACS patients, and AAS patients are 0.99, 0.98, and 0.97, respectively.

A Multichannel Fluorescence Isothermal Amplification Device with Integrated Internet of Medical Things for Rapid Sensing of Pathogens through Deep Learning.

The landscape of diagnostic assessments has experienced a paradigm shift driven by the advent of isothermal amplification techniques on point-of-care testing (POCT). The development of compact, portable isothermal amplification devices further emphasizes their transformative influence on diagnostic approaches. However, in prioritizing portability, these devices may exhibit limitations in functionality, rendering them less effective in addressing urgent public health emergencies during sudden pathogen outbreaks. In this paper, an efficient isothermal fluorescence amplification device has been fabricated for the rapid detection of pathogens during public health crises. The device features multichannel capability for simultaneous detection of various targets, integrates with the Internet of Medical Things (IoMT) for remote control and data uploading, and includes a deep learning-based batch processing system for rapid (9.4 ms) and accurate discrimination of pathogen type with excellent accuracy. The device has been successfully employed to simultaneously detect Staphylococcus aureus (SA) and methicillin-resistant Staphylococcus aureus (MRSA) with limits of detection (LODs) of 18 CFU/mL (SA) and 20 CFU/mL (MRSA) within 35 min by multiplex RPA assay and CRISPR/Cas12a-mediated nucleic acid detection assay.

Lateral flow immunoassay with peptide-functionalized gold nanoparticles for rapid detection of protein tyrosine phosphatase 1B.

In this work, a lateral flow immunoassay (LFIA) with peptide functionalized gold nanoparticles (termed as biotin-ppeptide-AuNPs) has been developed for rapid, semi-quantitative detection of PTP1B activity without using any sophisticated equipment. In this method, the anti-phosphotyrosine (anti-pY) monoclonal antibody and streptavidin were used as test line and control line, respectively. The biotin-ppeptide-AuNPs contain 10% biotinylated peptide ligand carry a motif SDGHEpYIYVDP with pY (phosphotyrosine) and 90% pentapeptide (CALNN) ligand, which are used as PTP1B substrates and LFIA labelling probes. The experimental results demonstrate that the as-proposed LFIA with biotin-ppeptide-AuNPs exhibits a wide linear range (from 50 ng/mL to 10 µg/mL), a relatively low limit of detection (LOD, 44 ng/mL), and good specificity. In addition, the LFIA with biotin-ppeptide-AuNPs has been successfully used to evaluate activity levels of PTP1B in four cell lysates and the detection results exhibit a consistent trend with that of commercial kit.

Discovery of Phenolic Matrix Metalloproteinase Inhibitors by Peptide Microarray for Osteosarcoma Treatment.

Because of the abnormal upregulation of matrix metalloproteinase (MMP) activities in tumors, MMP inhibitors (MMPIs) are validated anticancer drug candidates. We identified several MMPIs including mangiferin as an MMP-9 inhibitor with a half maximal inhibitory concentration (IC50) value of 250 nM, isosilybin as an MMP-13 inhibitor with an IC50 value of 250 nM, and isoliquiritigenin as a broad-spectrum MMPI (with IC50 values of 16 nM for MMP-1, 10 nM for MMP-2, 81 nM for MMP-3, 8 nM for MMP-7, 10 nM for MMP-9, and 14 nM for MMP-13) through studying the interactions of 6 MMPs secreted by U-2OS cells with 51 phenolic natural products on the peptide microarray platform. In addition, the inhibitory mechanisms of as-discovered MMPIs were evaluated by a molecular docking simulation. The antitumor efficiencies of MMPIs were demonstrated by both a cell scratch test and growth suppression of mouse-born OS tumors. The results of the cell scratch test suggested that isoliquiritigenin significantly inhibited the migration of U-2OS cells. In addition, administration of isoliquiritigenin significantly reduced the tumor size (by about 80%) and prolonged the survival time (by more than 70 days). This study suggests that the discovery of MMPIs from phenolic natural products is a meaningful way to screen anticancer agents.

The Peptide Functionalized Inorganic Nanoparticles for Cancer-Related Bioanalytical and Biomedical Applications.

In order to improve their bioapplications, inorganic nanoparticles (NPs) are usually functionalized with specific biomolecules. Peptides with short amino acid sequences have attracted great attention in the NP functionalization since they are easy to be synthesized on a large scale by the automatic synthesizer and can integrate various functionalities including specific biorecognition and therapeutic function into one sequence. Conjugation of peptides with NPs can generate novel theranostic/drug delivery nanosystems with active tumor targeting ability and efficient nanosensing platforms for sensitive detection of various analytes, such as heavy metallic ions and biomarkers. Massive studies demonstrate that applications of the peptide-NP bioconjugates can help to achieve the precise diagnosis and therapy of diseases. In particular, the peptide-NP bioconjugates show tremendous potential for development of effective anti-tumor nanomedicines. This review provides an overview of the effects of properties of peptide functionalized NPs on precise diagnostics and therapy of cancers through summarizing the recent publications on the applications of peptide-NP bioconjugates for biomarkers (antigens and enzymes) and carcinogens (e.g., heavy metallic ions) detection, drug delivery, and imaging-guided therapy. The current challenges and future prospects of the subject are also discussed.

Development of a peptide microarray-based metal-enhanced fluorescence assay for ultrasensitive detection of multiple matrix metalloproteinase activities by using a gold nanorod-polymer substrate.

Matrix metalloproteinases (MMPs) are attractive biomarkers for cancer diagnosis and treatment, while it is still a challenge to precise analysis of MMP activities owing to their very low abundance in the biological samples, especially at the early stages of tumors. Herein, a peptide microarray-based metal-enhanced fluorescence assay (PMMEFA) is proposed to simultaneously detect MMP-1, -2, -3, -7, -9, and -13 activities. The assay involves immobilization of Förster resonance energy transfer dye pair decorated peptides (FRET-peptides) on a poly(glycidyl methacrylate-co-2-hydroxyethyl methacrylate) coated gold nanorod modified glass slide (GNR@P(GMA-HEMA)). To fabricate the GNR@P(GMA-HEMA) slide, GNRs are self-assembled onto an aminated glass slide, and a polymer brush (P(GMA-HEMA)) is grown through a surface-initiated atom transfer radical polymerization reaction (SI-ATRP). Upon the addition of MMPs, the FRET pairs are broken due to the specific cleavage of FRET-peptides by enzymes, resulting in the recovery of fluorescence signals and further enhancement by the MEF of GNRs. The fluorescence recovery degree provides a direct indicator for MMP activity. The PMMEFA exhibits excellent sensitivity, which enables to detect MMP-1, -2, -3, -7, -9, and -13 activities, with low limits of detection (LODs) of 1.7 fg mL-1, 0.3 fg mL-1, 2.0 fg mL-1, 1.8 fg mL-1, 2.2 fg mL-1 and 14.0 fg mL-1, respectively. To substantiate the practicability of PMMEFA, MMP activities were measured in a range of matrices, encompassing cell culture medium, serum, and tumor tissue homogenate, and MMP activities can be detected only in 0.15 µL serum and 0.025 mg tumor tissue.

Scalable synthesis of Au@CeO2 nanozyme for development of colorimetric lateral flow immunochromatographic assay to sensitively detect heart-type fatty acid binding protein.

Nanozymes with core@shell nanostructure are considered promising biolabeling materials for their multifunctional properties. In this work, a simple one-pot strategy has been proposed for scalable synthesis of gold@cerium dioxide core@shell nanoparticles (Au@CeO2 NPs) with strong localized surface plasmon resonance (LSPR) absorption and high peroxidase-like catalytic activity by redox reactions of Ce3+ ions and AuCl4- ions in diluted ammonia solution under room temperature. A colorimetric lateral flow immunochromatographic assay (LFIA) has been successfully fabricated for sensitive detection of heart-type fatty acid binding protein (H-FABP, an early cardiac biomarker) by using the Au@CeO2 NPs as reporters. The as-developed LFIA with Au@CeO2 NP reporter (termed as Au@CeO2-LFIA) exhibits a dynamic range of nearly two orders of magnitude, and a limit of detection (LOD) as low as 0.35 ng mL-1 H-FABP with nanozyme-triggered 3,3',5,5'-tetramethylbenzidine (TMB) colorimetric amplification. Furthermore, the practicality of Au@CeO2-LFIA has been demonstrated by profiling the concentrations of H-FABP in 156 blood samples of acute myocardial infarction (AMI) patients, and satisfactory results are obtained.

Lectin microarray based glycan profiling of exosomes for dynamic monitoring of colorectal cancer progression.

BACKGROUND: Exosomes, as emerging biomarkers in liquid biopsies in recent years, offer profound insights into cancer diagnostics due to their unique molecular signatures. The glycosylation profiles of exosomes have emerged as potential biomarkers, offering a novel and less invasive method for cancer diagnosis and monitoring. Colorectal cancer (CRC) represents a substantial global health challenge and burden. Thus there is a great need for the aberrant glycosylation patterns on the surface of CRC cell-derived exosomes, proposing them as potential biomarkers for tumor characterization. RESULTS: The interactions of 27 lectins with exosomes from three CRC cell lines (SW480, SW620, HCT116) and one normal colon epithelial cell line (NCM460) have been analyzed by the lectin microarray. The result indicates that Ulex Europaeus Agglutinin I (UEA-I) exhibits high affinity and specificity towards exosomes derived from SW480 cells. The expression of glycosylation related genes within cells has been analyzed by high-throughput quantitative polymerase chain reaction (HT-qPCR). The experimental result of HT-qPCR is consistent with that of lectin microarray. Moreover, the limit of detection (LOD) of UEA-I microarray is calculated to be as low as 2.7 × 105 extracellular vehicles (EVs) mL-1 (three times standard deviation (3σ) of blank sample). The UEA-I microarray has been successfully utilized to dynamically monitor the progression of tumors in mice-bearing SW480 CRC subtype, applicable in tumor sizes ranging from 2 mm to 20 mm in diameter. SIGNIFICANCE: The results reveal that glycan expression pattern of exosome is linked to specific CRC subtypes, and regulated by glycosyltransferase and glycosidase genes of mother cells. Our findings illuminate the potential of glycosylation molecules on the surface of exosomes as reliable biomarkers for diagnosis of tumor at early stage and monitoring of cancer progression.

Profiling of Multiple Matrix Metalloproteinases Activities in the Progression of Osteosarcoma by Peptide Microarray-Based Fluorescence Assay on Polymer Brush-Coated Zinc Oxide Nanorod Substrate.

Peptide microarray provides the ability to miniaturize, parallelize, and automate high-throughput screening substrate specificities of enzymes, profiling of multiple enzyme activities, discovery of disease biomarkers, and development of drugs. Matrix metalloproteinases (MMPs) are demonstrated as important biomarkers of tumor invasion and metastasis. Herein, a peptide microarray-based fluorescence assay is proposed to profile multiple MMPs (MMP-1, MMP-2, MMP-3, MMP-7, MMP-9, and MMP-13) activities in the culture medium of four human osteosarcoma (OS) cells and in the progression of OS by using the mouse-bearing xenograft OSs including U-2OS and Saos-2 human. This method has excellent selectivity and sensitivity, which enables to detect the activities of cellular secreted MMP-1, MMP-2, MMP-3, MMP-7, MMP-9, and MMP-13 with limit of detection downs to 10 pM, 30 pM, 113 pM, 13 pM, 93 pM, and 12 pM, respectively. Furthermore, it is demonstrated that the activity pattern of MMPs is serum closely relevant to the disease progression and type of tumor.

Analysis and Modeling of Aged SAC-Bi Solder Joints Subjected to Varying Stress Cycling Conditions.

Solder joints are subjected to varied stress cycle circumstances in the electronic packaging service life but are also influenced by aging. There has been limited investigation into the influence of aging and varying cycles on SnAgCu-Bi (SAC-Bi) solder joint fatigue. Cyclic fatigue tests were performed on solder joints of several alloys, including SnAgCu (SAC305), SnAgCu-Bi (SAC-Q), and SnCu-Bi (SAC-R). Individual solder joints were cycled under varying stress levels, alternating between mild and harsh stress levels. At least seven samples were prepared for each alloy by alternating between 25 mild stress (MS) cycles and three harsh stress (HS) cycles until the solder joint broke off. The impact of aging on Bi-doped solder joints fatigue under varied amplitude stress was examined and predicted for 10 and 1000 h under 125 °C. Because of the "Step-up" phenomenon of inelastic work, a new fatigue model was developed based on the common damage accumulation (CDA) model. The experimental results revealed that aging reduced the fatigue life of the tested solder alloys, particularly that of SAC305. According to the CDA model, all solder alloys failed earlier than expected after aging. The proposed model uses the amplification factor to assess inelastic work amplification after switching between the MS and HS cycles under varying stress amplitude conditions. The amplification factor for the SAC-Bi solder alloys increased linearly with fracture initiation and substantially followed crack propagation until the final failure. Compared with existing damage accumulation models, the proposed fatigue model provides a more accurate estimation of damage accumulation. For each case, the cut-off positions were examined. The SAC-Q amplification factor increased linearly to 83% of its overall life, which was much higher than that of SAC305 and SAC-R. This study identified three distinct failure modes: ductile, brittle, and near intermetallic compound (IMC) failure. It was also observed that SAC-Q with an organic solderability preservatives (OSP) surface finish was more susceptible to brittle failure owing to the excessive brittleness of the alloy material.

Biosensors and bioassays for determination of matrix metalloproteinases: state of the art and recent advances.

Matrix metalloproteinases (MMPs) are closely associated with various physiological and pathological processes, and have been regarded as potential biomarkers for severe diseases including cancer. Accurate determination of MMPs would advance our understanding of their roles in disease progression, and is of great significance for disease diagnosis, treatment and prognosis. In this review, we present a comprehensive overview of the developed bioassays/biosensors for detection of MMPs, and highlight the recent advancement in nanomaterial-based immunoassays for MMP abundance measurements and nanomaterial-based biosensors for MMP activity determination. Enzyme-linked immunosorbent assay (ELISA)-based immunoassays provide information about total levels of MMPs with high specificity and sensitivity, while target-based biosensors measure the amounts of active MMPs, and allow imaging of MMP activities in vivo. For multiplex and high-throughput analysis of MMPs, microfluidics and microarray-based assays are described. Additionally, we put forward the existing challenges and future prospects from our perspective.

Synthesis of heteronanostructures for multimodality molecular imaging-guided photothermal therapy.

Combining photothermal therapy (PTT) and multimodality molecular imaging into one nanotheranostic can improve the diagnostics and therapeutics outcome of malignant tumors significantly. Herein, a heteronanostructure (named Au-Fe3O4@PDA-PEG-DTPA-Gd) has been synthesized for multimodality molecular imaging-guided PTT by conjugation of gadolinium-diethylenetriaminepentaacetic acid (Gd-DTPA) on a polydopamine (PDA) coated Au-Fe3O4 heterodimeric via a polyethylene glycol (PEG) crosslinker. Au-Fe3O4@PDA-PEG-DTPA-Gd exhibits reasonable biocompatibility, high photothermal conversion efficiency (η = 41.3%), integration of excellent T1-/T2-weighted magnetic resonance imaging (MRI, the molar longitudinal relaxivity rate (r1) = 6.14 mM-1 s-1/transverse relaxation rate (r2) = 71.88 mM-1 s-1) and X-ray computed tomography imaging (CT, approximately 1.94 times higher than that of the commercial CT contrast agent ioversol) contrast functionality. The theranostic capability of Au-Fe3O4@PDA-PEG-DTPA-Gd has been verified on the triple negative breast tumor cell (MDA-MB-231 cell) and MDA-MB-231 tumor-bearing mouse model. From MR/CT imaging, Au-Fe3O4@PDA-PEG-DTPA-Gd shows remarkable PTT efficacy, which completely inhibits MDA-MB-231 tumor growth in vivo with a single treatment.

Array-based in situ fluorescence assay for profiling multiplex matrix metalloproteinases activities in tissue section.

Herein, an array-based in situ fluorescence assay is proposed for high-throughput analysis and localization of multiplex matrix metalloproteinases (MMPs) activities in cell monolayers and tissue sections. Five specific MMPs (MMP-2, -3, -7, -9, and -14) peptide substrates containing FAM/Dabcyl fluorescent resonance energy transfer (FRET) pair are directly spotted on the surface of cell monolayers or tissue sections, and hydrolyzed by localized MMPs, resulting in fluorescence recovery of FAM. MMPs activities are determined by the fluorescence intensity of stained cells/tissues due to the cellular internalization of peptide fragments with FAM moiety. We demonstrate that the array-based in situ fluorescence assay is suitable for identifying the MMPs expression patterns of cells, as well as determining the secreted MMPs activities in cell monolayer with high sensitivity (as low as hundreds of cells per square centimeter). The feasibility of the assay is further confirmed by evaluating inhibition potencies of six compounds toward five MMPs. Profiling of five MMPs activities in the localized parts of 32 thyroid tissues is performed without separation or extraction procedures, demonstrating the good practicality of the method.

Two-Dimensional Layered Nanomaterial-Based Electrochemical Biosensors for Detecting Microbial Toxins.

Toxin detection is an important issue in numerous fields, such as agriculture/food safety, environmental monitoring, and homeland security. During the past two decades, nanotechnology has been extensively used to develop various biosensors for achieving fast, sensitive, selective and on-site analysis of toxins. In particular, the two dimensional layered (2D) nanomaterials (such as graphene and transition metal dichalcogenides (TMDs)) and their nanocomposites have been employed as label and/or biosensing transducers to construct electrochemical biosensors for cost-effective detection of toxins with high sensitivity and specificity. This is because the 2D nanomaterials have good electrical conductivity and a large surface area with plenty of active groups for conjugating 2D nanomaterials with the antibodies and/or aptamers of the targeted toxins. Herein, we summarize recent developments in the application of 2D nanomaterial-based electrochemical biosensors for detecting toxins with a particular focus on microbial toxins including bacterial toxins, fungal toxins and algal toxins. The integration of 2D nanomaterials with some existing antibody/aptamer technologies into electrochemical biosensors has led to an unprecedented impact on improving the assaying performance of microbial toxins, and has shown great promise in public health and environmental protection.