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Exosomes are extracellular vesicles with unique size distribution derived from the parent cells. They are involved in intercellular communication and transport, and are also biomarkers for early diagnosis and prognosis of disease. However, the isolation and characterization of exosomes face the challenges of large sample requirements and low enrichment efficiency of traditional methods. Herein, a simple method is proposed for the preparation of an artificial antibody that has a synergistic effect by featuring nanocavities obtained by dull template imprinting and molecular recognition conferred by electrostatic interaction. With this artificial antibody, highly efficient capture and proteome analysis of exosomes from urine and cell culture media are achieved: for urine, the abundance of Top100 exosomal proteins enriched by this artificial antibody increased from 1.85% to 9.66%. For the cell culture medium, the abundance of the Top100 proteins enriched by this artificial antibody was 28.4%. Moreover, this artificial antibody has a comparable effect to the commercial precipitation-based method in capturing exosomes and has advantages in removing contaminants such as prothymosin alpha (PTMA), demonstrating the superior selectivity of the artificial antibody. Overall, this artificial antibody holds promise to capture exosomes from biofluids and is compatible with subsequent proteome analysis. [ABSTRACT FROM AUTHOR]

Nanozymes have great potential applications in tumor treatment due to their good stability, high biocompatibility, easy preparation and versatility. However, it remains a challenge to design highly active nanozymes with tumor cell targeting. Herein, three nanoceria structures (nanoceria-rod; nanoceria polyhedra, abbreviated as nanoceria-poly.; and nanoceria-cube) with different surface oxygen vacancy concentrations are designed. Among them, nanoceria-rod shows the highest enzyme activity and tumor cell toxicity because of its highest concentration of oxygen vacancies on the surface. Further study shows that nanoceria-rod can selectively enter tumor cells because nanoceria-rod with a suitable isoelectric point (IEP) remains positively charged in the acidic microenvironment of the tumor but negatively charged in the physiological microenvironment of normal cells. Nanoceria-rod distributes in lysosomes and phagosomes to produce reactive oxygen species (ROS) in tumor cells. Finally, the mitochondrial membrane potential (MMP) was reduced, which caused cell apoptosis. This study provides an interesting new tumor-targeting therapy method, which could also be extended to other drug nanocarriers and diagnostic imaging nanomaterials for tumors. [ABSTRACT FROM AUTHOR]

The pseudo isobaric peptide termini labeling (IPTL) method is a remarkable strategy in quantitative proteomics, and has been efficiently applied in biological studies due to its high quantitative accuracy. However, irreproducible precursor ion selection caused by data dependent acquisition and the chromatographic shift caused by isotope effects limit the wide application of this method. Herein, we expand the use of pseudo IPTL to SWATH MS application and develop a novel quantitative strategy, termed SWATH-pseudo-IPTL, by which the relative quantification could be achieved by comparing the “complete” extracted ion chromatogram (XIC) intensity of MS/MS scan instead of a single intensity measurement in DDA-pseudo-IPTL which only reflected the peptide abundances at that given time. The quantitative analysis of various proportions of mixed HeLa samples revealed the strong accuracy and precision of our SWATH-pseudo-IPTL method, both of which were better than that of the DDA-pseudo-IPTL strategy. SWATH-pseudo-IPTL was also applied to the quantitative profiling of the proteome from human hepatocellular carcinoma cell lines with high and low metastatic potential, and most of the differentially expressed proteins were related to tumorigenesis and tumor metastasis, demonstrating the feasibility of this methodology for biological applications. [ABSTRACT FROM AUTHOR]

Exosomes are secreted nanovesicles shed by almost all kinds of cells. Recently, increased interest has been focused on these extracellular vesicles as natural carriers transporting biological contents for intercellular communication. However, current isolation techniques, such as ultracentrifugation, are not convenient and often require specialized equipment. Herein, we describe a polyethylene glycol (PEG)-based approach, which could permit facile, low-cost and effective isolation of exosomes from cell culture supernatant. High-resolution electron microscopes clearly visualized the size and morphology of isolated exosome aggregates, implying the mechanism of PEG-based precipitation. Combined with tandem mass spectrometry analysis, 6299 protein groups encoded by 5120 genes were successfully characterized from HeLa cell culture supernatant, including numerous exosome proteins which could overlap 97% of the Top 100 exosome marker proteins recorded in the ExoCarta database, as well as a series of low-abundance cytokines and biomarkers. Furthermore, we found a higher ratio of neo-cleavage sites in proteins identified from exosomes compared with cellular proteins, revealing the potential roles of exosomes in accumulation and transportation of protein degradation intermediates. [ABSTRACT FROM AUTHOR]