AI-Driven Design System for Fabrication of Inhalable Nanocatchers for Virus Capture and Neutralization.

The global pandemic presents a critical threat to humanity, with no effective rapid-response solutions for early-stage virus dissemination. This study aims to create an AI-driven entry-blocker design system (AIEB) to fabricate inhalable virus-like nanocatchers (VLNCs) fused with entry-blocking peptides (EBPs) to counter pandemic viruses and explore therapeutic applications. This work focuses on developing angiotensin-converting enzyme 2 (ACE2)-mimic domain-fused VLNCs (ACE2@VLNCs) using AIEB and analyzing their interaction with the SARS-CoV-2 receptor binding domain (RBD), demonstrating their potential to hinder SARS-CoV-2 infection. Aerosol-based tests show ACE2@VLNCs persist over 70 min in the air and neutralize pseudoviruses within 30 min, indicating their utility in reducing airborne virus transmission. In vivo results reveal ACE2@VLNCs mitigate over 67% of SARS-CoV-2 infections. Biosafety studies confirm their safety, causing no damage to eyes, skin, lungs, or trachea, and not eliciting significant immune responses. These findings offer crucial insights into pandemic virus prevention and treatment, highlighting the potential of the ACE2@VLNCs system as a promising strategy against future pandemics.

Bioengineered Bacteriophage-Like Nanoparticles as RNAi Therapeutics to Enhance Radiotherapy against Glioblastomas.

Since glioblastomas (GBMs) are radioresistant malignancies and most GBM recurrences occur in radiotherapy, increasing the effectiveness of radiotherapy by gene-silencing has recently attracted attention. However, the difficulty in precisely tuning the composition and RNA loading in nanoparticles leads to batch-to-batch variations of the RNA therapeutics, thus significantly restricting their clinical translation. Here, we bioengineer bacteriophage Qß particles with a designed broccoli light-up three-way junction (b-3WJ) RNA scaffold (contains two siRNA/miRNA sequences and one light-up aptamer) packaging for the silencing of genes in radioresistant GBM cells. The in vitro results demonstrate that the cleavage of de novo designed b-3WJ RNA by Dicer enzyme can be easily monitored in real-time using fluorescence microscopy, and the TrQß@b-3WJLet-7gsiEGFR successfully knocks down EGFR and IKKα simultaneously and thereby inactivates NF-κB signaling to inhibit DNA repair. Delivery of TrQß@b-3WJLet-7gsiEGFR through convection-enhanced delivery (CED) infusion followed by 2Gy X-ray irradiation demonstrated that the median survival was prolonged to over 60 days compared with the 2Gy X-ray irradiated group (median survival: 31 days). Altogether, the results of this study could be critical for the design of RNAi-based genetic therapeutics, and CED infusion serves as a powerful delivery system for promoting radiotherapy against GBMs without evidence of systemic toxicity.

Microneedle patches integrated with lateral flow cassettes for blood-free chronic kidney disease point-of-care testing during a pandemic.

Chronic kidney disease (CKD) is the most neglected chronic disease affecting over 750 million persons in the world. Currently, many patients with cancers or other chronic diseases (i.e., CKD) struggle to receive clinical treatment or examination due to hospitals cancelling or delaying in the COVID-19 pandemic, which may increase the risk of death. Cystatin C (Cys C) has been proposed as a potential glomerular filtration rate (GFR) marker for the early detection of acute kidney injury and CKD. However, most traditional methods for Cys C detection are immunoassays using serum as a sample and are tedious to perform and economically burdensome. To diagnose the disease in the early stage and carry out daily management during the current pandemic, we developed an integration of hydrogel microneedle patch (HMNP) and lateral flow cassette (LFC) to rapidly detect Cys C in skin interstitial fluid (ISF) in 25 min for blood-free CKD management anytime and anywhere by the naked eye that can reduce the impact of an individual's quality of life and life expectancy. Conceivably, this strategy presents a wide scope in the application of numerous other diseases if corresponding analytes are available in skin ISF.

A new lateral flow plasmonic biosensor based on gold-viral biomineralized nanozyme for on-site intracellular glutathione detection to evaluate drug-resistance level.

Temozolomide (TMZ)-resistant glioblastoma multiforme (GBM) cells would have abnormal redox status due to bio-thiols, like glutathione (GSH), which constitute the most crucial defense system that protects cells from therapeutic agents. Current strategies for GSH detection often require sophisticated instruments that may not be available in laboratories with fewer resources. Here, we circumvent this problem by introducing a lateral flow plasmonic biosensor (LFPB) based on gold-viral biomineralized nanoclusters (AuVCs) as nanozymes that enables the detection of a few molecules with the naked eye and quantified by an auto-analysis software. The GSH level controls the growth of gold nanoparticles (AuNPs) and generates coloured patterns with distinct tonality, which are then auto-analyzed to calculate the GSH concentrations by smartphone with an auto-analysis software. Under the optimized conditions, grayscale value plotted against GSH concentration exhibited a linear relationship within the range of 25-500 µM with a limit of detection (LoD) of 9.80 µM and highly positive correlation between detected GSH level and TMZ drug-resistance level in GBM cells. This excellent property allowed our approach to be used for on-site determination of GSH levels in a rapid (i.e., within 30 min), simple (i.e., auto-analysis software), and cost-effective process (i.e., instrument-free) for cancer precision therapy.

Mobile healthcare system based on the combination of a lateral flow pad and smartphone for rapid detection of uric acid in whole blood.

Excessive production of uric acid (UA) in blood may lead to gout, hyperuricaemia and kidney disorder; thus, a fast, simple and reliable biosensor is needed to routinely determine the UA concentration in blood without pretreatment. The purpose of this study was to develop a mobile healthcare (mHealth) system using a drop of blood, which comprised a lateral flow pad (LFP), mesoporous Prussian blue nanoparticles (MPBs) as artificial nanozymes and auto-calculation software for on-site determination of UA in blood and data management. A standard curve was found to be linear in the range of 1.5-8.5 mg/dL UA, and convenience, cloud computing and personal information management were simultaneously achieved for the proposed mHealth system. Our mHealth system appropriately met the requirements of application in patients' homes, with the potential of real-time monitoring by their primary care physicians (PCPs).

Instrument-Free Detection of FXYD3 Using Vial-Based Immunosensor for Earlier and Faster Urothelial Carcinoma Diagnosis.

The incidence and 5 year recurrence rate of urothelial carcinomas (UCs), including UC of the bladder (UCB) and upper urinary tract UC (UTUC), have increased annually. There is a great need for a simple and fast point-of-care (POC) test for early diagnosis and amelioration in the survival rate. We present a POC test comprising a new vial-immunosensor, nanoenzyme, and iPhone 7 plus, which detects and quantifies the new biomarker FXYD domain-containing ion transport regulator 3 (FXYD3) in human urine for specific UC screening, tumor-grade classification, and postoperative monitoring by the grayscale value of the photograph taken. The performance of the proposed POC test was then verified using urine from 4 healthy people, 40 UCB patients (10 patients were low-grade and 30 patients were high-grade), and 13 UTUC patients (2 patients were low-grade and 11 patients were high-grade), confirming the accuracy and specificity by comparing the results with those obtained by enzyme-linked immunosorbent assay (ELISA). Moreover, we also designed a correction method that can make the grayscale values calculated by different smartphones close to the values calculated by iPhone 7 plus, resulting in the POC test enabling simple, fast, universal, and portable testing, data storage, and sharing for personal UCs screening and postoperative monitoring.

A serological point-of-care test for Zika virus detection and infection surveillance using an enzyme-free vial immunosensor with a smartphone.

Zika virus (ZIKV) is a mosquito-borne flavivirus (FLAV) that emerged in Brazil in 2015 and has rapidly spread to more than 50 countries worldwide. However, early, accurate, and specific point-of-care (POC) diagnosis of ZIKV is very difficult because most infected patients are asymptomatic or display nonspecific symptoms similar to those of other viral infections, and most of the analysis also requires instruments. Herein, an instrument-free ZIKV POC test using a drop of blood comprising a vial immunosensor, artificial nanozyme platinum/gold core-shell nanoparticles (Pt@Au NPs) as a signal probe, and a smartphone was developed to specifically detect ZIKV without cross-reaction with other FLAVs. A high sensitivity of 1 pg/mL ZIKV, desirable specificity, data storage, and geographic location surveillance were simultaneously achieved for the proposed POC test. Our POC test suitably met the urgent needs of ports of entry, airports, and endemic regions with stressed resources, as well as strict clinical requirements for ZIKV detection.

Direct glucose detection in whole blood by colorimetric assay based on glucose oxidase-conjugated graphene oxide/MnO2 nanozymes.

We herein report a facile approach for the preparation of horseradish peroxidase (HRP)-mimic glucose oxidase-conjugated graphene oxide/MnO2 (GOD-GO/MnO2) as new nanozyme to detect the glucose concentration in whole blood. The nano-sized of MnO2 nanoparticles embedded in bovine serum albumin (BSA)-coated GO by in situ growth were evaluated focusing on the principle of HRP-mimic activity catalyzing the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of hydrogen peroxide. Furthermore, we constructed dual sensing platforms based on the combination of a plasma separation pad and GOD-GO/MnO2 for direct detection of glucose concentration in whole blood by colorimetric assay without blood sample pretreatment. As a proof-of-concept, a limit of detection of 3.1 mg dL-1 for glucose was obtained with a wide linear quantification range from 25 mg dL-1 to 300 mg dL-1 through visual observation and quantitative analysis, suggesting potential clinical applications in blood glucose monitoring for diabetic patients.

Combined Detection of CA19-9 and MUC1 Using a Colorimetric Immunosensor Based on Magnetic Gold Nanorods for Ultrasensitive Risk Assessment of Pancreatic Cancer.

We herein report a facile approach for developing an enzyme-free colorimetric immunosensor based on a magnetic iron oxide (IO)-coated gold nanorod (MGNR) nanocomposite with high electron transfer ability to accelerate the color bleaching reaction of methyl orange (MO) in the presence of NaBH4 for ultrasensitive detection of cancer antigens. In the case of MO, the reaction rate of MGNRs showed approximately 45.6-fold and 1520.8-fold higher than that of Cys-GNRs and NaBH4, respectively. The proposed colorimetric immunosensor was demonstrated to enable simple, cost-effective, sensitive, and specific carbohydrate antigen 19-9 (CA19-9) and mucin 1 (MUC1) detection for risk evaluation of pancreatic cancer (PC) with a small volume of serum sample without the use of any enhancing solutions or enzymes. By increasing the concentration of CA19-9 and MUC1, more MGNRs remained in the plate well to enhance the color bleaching of MO. As a proof-of-concept, the limit of detection (LOD) of 3.5 × 10-5 U/mL for CA19-9 and 5.2 × 10-6 U/mL for MUC1 was obtained with a wide linear quantification range from 8.6 × 10-5 U/mL to 1.4 × 10-2 U/mL for CA19-9 and 1.3 × 10-5 U/mL to 2.1 × 10-3 U/mL for MUC1, suggesting potential clinical applications for the early risk evaluation of PC.

A colorimetric immunosensor based on self-linkable dual-nanozyme for ultrasensitive bladder cancer diagnosis and prognosis monitoring.

We developed self-linkable Prussian blue (PB)-incorporated magnetic graphene oxide (PMGO) as a peroxidase-mimicking nanozyme with high oxidizability to 3,3',5,5'-tetramethylbenzidine (TMB), which generates significant absorption intensity for the colorimetric immunosensing of apolipoprotein A1 (ApoA1) in early bladder cancer (BC) diagnosis and prognosis monitoring. The ultrasensitive immunosensor was constructed using an ApoA1 antibody (AbApoA1)-functionalized chip (biochipApoA1) and self-linkable peroxidase-mimicking, PB-incorporated magnetic graphene oxide (PMGO). After incubating the sample and capturing ApoA1 proteins captured on the biochipApoA1, the PMGO was functionalized with AbApoA1, and then mouse IgG (PMGO-1), rabbit anti-mouse IgG antibody (PMGO-2), and goat anti-rabbit IgG antibody (PMGO-3) were added together. We envisioned that each captured ApoA1 protein would allow the retention of a large amount of PMGO through a self-linking process to amplify the colorimetric signal of TMB in the presence of H2O2. The linear detection range could be obviously widened in the presence of self-linkable PMGO-from 0.05 ng/mL to 100 ng/mL-compared with the group without signal amplification (from 1 ng/mL to 100 ng/mL). Our immunosensor analysis of ApoA1 in the urine of BC patients and healthy individuals was highly correlated with enzyme-linked immunosorbent assay measurements; moreover, the ApoA1 concentrations of patients with high-grade BC were significantly higher than those of patients with low-grade BC. After standard clinical treatment, a significant drop of ApoA1 concentration occurred in urine that was lower than the cut-off concentration, suggesting potential clinical applications of the new self-linkable PMGO-generating colorimetric immunosensor in early BC diagnosis and prognosis monitoring.