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The isolation and enrichment of specific extracellular vesicle (EV) subpopulations are essential in the context of precision medicine. However, the current methods predominantly rely on a single-positive marker and are susceptible to interference from soluble proteins or impurities. This limitation represents a significant obstacle to the widespread application of EVs in biological research. Herein, a novel approach that utilizes proximity ligation assay (PLA) and DNA-RNA hybridization are proposed to facilitate the binding of two proteins on the EV membrane in advance enabling the isolation and enrichment of intact EVs with double-positive membrane proteins followed by using functionalized magnetic beads for capture and enzymatic cleavage for isolated EVs release. The isolated subpopulations of EVs can be further utilized for cellular uptake studies, high-throughput small RNA sequencing, and breast cancer diagnosis. Hence, developing and implementing a specialized system for isolating and enriching a specific subpopulation of EVs can enhance basic and clinical research in this field., (© 2023 Wiley‐VCH GmbH.)

Red-to-near-infrared (NIR) fluorescent probes, with advantages such as high spatiotemporal resolution and in situ sensing abilities, are highly attractive for diagnosis of gastrointestinal diseases and targeted drug development. However, conventional red-to-NIR fluorophores with electron closed-shell structures require tedious synthetic procedures for preparation, and it is difficult to further decorate them with sensing groups. In this study, a series of easily prepared pyrroles with simple structures that can quickly be transformed into red-to-NIR emissive radical cations in acidic buffer solution and in vivo stomachs is developed. The in-situ-generated red-to-NIR emissive pyrrole radical cations in the stomach have excellent biocompatibility and stability and can be used not only for intravital gastrointestinal imaging with high spatiotemporal resolution, but also for dynamic monitoring of the gastric emptying process and assessment of anti-gastric-acid therapy. The acidity-induced generation of pyrrole radical cations is believed to provide a facile strategy for developing red-to-NIR fluorophores and studying gastrointestinal diseases., (© 2023 Wiley-VCH GmbH.)

Subarachnoid hemorrhage (SAH) is a severe subtype of stroke caused by the rupturing of blood vessels in the brain. The ability to accurately assess the degree of bleeding in an SAH model is crucial for understanding the brain-damage mechanisms and developing therapeutic strategies. However, current methods are unable to monitor microbleeding owing to their limited sensitivities. Herein, a new bleeding assessment system using a bioprobe TTVP with aggregation-induced emission (AIE) characteristics is demonstrated. TTVP is a water-soluble, small-molecule probe that specifically interacts with blood. Taking advantage of its AIE characteristics, cell membranes affinity, and albumin-targeting ability, TTVP fluoresces in bleeding areas and detects the presence of blood with a high signal-to-noise (S/N) ratio. The degree of SAH bleeding in an endovascular perforation model is clearly evaluated based on the intensity of the fluorescence observed in the brain, which enables the ultrasensitive detection of mirco-bleeding in the SAH model in a manner that outperforms the current imaging strategies. This method serves as a promising tool for the sensitive analysis of the degree of bleeding in SAHs and other hemorrhagic diseases., (© 2023 The Authors. Advanced Science published by Wiley-VCH GmbH.)

New, biocompatible materials with favorable antibacterial activity are highly desirable. In this work, we develop a unique conjugated polymer featuring aggregation-induced emission (AIE) for reliable bacterial eradication. Thanks to the AIE and donor-π-acceptor structure, this polymer shows a high reactive oxygen species (ROS)-generation ability compared to a low-mass model compound and the common photosensitizer Chlorin E6. Moreover, the selective binding of pathogenic microorganisms over mammalian cells was found, demonstrating its biocompatibility. The effective growth inhibition of bacteria upon polymer treatment under light irradiation was validated in vitro and in vivo. Notably, the recovery from infection after treatment with our polymer is faster than that with cefalotin. Thus, this polymer holds great promise in fighting against bacteria-related infections in practical applications., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Noninvasive prenatal detection of monogenic diseases based on cell-free DNA is hampered by challenges in obtaining a sufficient fraction and adequate quality of fetal DNA. Analyzing rare trophoblastic cells from Papanicolaou smears carrying the entire fetal genome provides an alternative method for noninvasive detection of monogenic diseases. However, intracellular labeling for identification of target cells can affect the quality of DNA in varying degrees. Here, a new approach is developed for nondestructive identification of rare fetal cells from abundant maternal cells based on endoplasmic reticulum staining and linear discriminant analysis (ER-LDA). Compared with traditional methods, ER-LDA has little effect on cell quality, allowing trophoblastic cells to be analyzed on the single-cell level. Using ER-LDA, high-purity of trophoblastic cells can be identified and isolated at single cell resolution from 60 pregnancies between 4 and 38 weeks of gestation. Pathogenic variants, including - SEA / deletion mutation and point mutations, in 11 fetuses at risk for α- or β-thalassemia can be accurately detected by this test. The detection platform can also be extended to analyze the mutational profiles of other monogenic diseases. This simple, low-cost, and noninvasive test can provide valuable fetal cells for fetal genotyping and holds promise for prenatal detection of monogenic diseases., Competing Interests: The authors declare no conflict of interest., (© 2020 The Authors. Published by WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim.)

Liquid biopsies of cancer via single-cell molecular profiling of circulating tumor cells (CTCs) are hampered by the lack of ideal CTC markers. In this study, it is reported that TPN, a bioprobe with aggregation-induced emission (AIE) activity is capable of distinguishing various tumor cells from blood leukocytes based on the difference in cell mitochondria. TPN is a cell-permeant live-cell stain that has little effect on cell viability and integrity, enabling single-cell DNA/RNA analysis with improved efficiency compared with traditional antibody-based methods. Using TPN labeling, CTCs and CTC cluster are detected in the blood from patients with lung or liver cancer. The capability of TPN to identify rare tumor cells in the malignant pleural effusion samples is also demonstrated. Furthermore, RNA sequencing of single lung CTC identified by TPN is successfully performed. The findings presented here provide an antibody-free, low-cost, and nondisruptive approach for detection and genomic characterization of viable tumor cells based on a mitochondria-targeting AIE luminogen. It might serve as a new tool for monitoring of genomics dynamic of tumor and unraveling the mechanisms of tumor metastasis., Competing Interests: The authors declare no conflict of interest., (© 2020 The Authors. Published by WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim.)

Two-photon photodynamic therapy (TP-PDT) is emerging as a powerful strategy for stereotactic targeting of diseased areas, but ideal photosensitizers (PSs) are currently lacking. This work reports a smart PS with aggregation-induced emission (AIE) feature, namely DPASP, for TP-PDT with excellent performances. DPASP exhibits high affinity to mitochondria, superior photostability, large two-photon absorption cross section as well as efficient reactive oxygen species generation, enabling it to achieve photosensitization both in vitro and in vivo under two-photon excitation. Moreover, its capability of stereotactic ablation of targeted cells with high-precision is also successfully demonstrated. All these merits make DPASP a promising TP-PDT candidate for accurate ablation of abnormal tissues with minimal damages to surrounding areas in the treatment of various diseases., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Extracellular vesicles (EVs) mediate intercellular communication via transferring proteins and other biological molecules and have been recently investigated as biomarkers of disease. Sensitive and specific biomarkers are required for lung cancer diagnosis and prognosis. The present study screens for abnormal EV proteins in non-small cell lung cancer (NSCLC) using a quantitative proteomics strategy involving LC-MS/MS to identify ideal biomarkers for NSCLC diagnosis. EVs are enriched from the sera of early and advanced NSCLC patients and healthy controls and from cell culture supernatants of lung adenocarcinoma and bronchial epithelial cell lines. In the sera and supernatants, 279 and 632 differentially expressed proteins, respectively, are associated with signaling pathways including extracellular membrane-receptor interaction, focal adhesion, and regulation of the actin cytoskeleton. Thirty-two EV proteins are identified at the intersection of differentially expressed proteins between the NSCLC groups and cell lines. Based on bioinformatics analysis, in silico immunohistochemical, and PRM verification, fibronectin is selected for following in vitro studies and validation with an independent cohort. Fibronectin on EVs is estimated to perform well in the diagnosis of NSCLC patients based on AUC, showing great potential for clinical use and demonstrating the efficacy of this method for EV-associated biomarker screening., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)