ArticleScientific reports2022
Confocal microscopy analysis reveals that only a small proportion of extracellular vesicles are successfully labelled with commonly utilised staining methods.
Article in Scientific reports, 2022. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 22 papers, 1 of them a synthesis that pooled it.
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Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
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Who cites it
22 citing papers in PubMed, 1 synthesis or guideline pooled it.
- Efficacy and safety of small extracellular vesicle interventions in wound healing and skin regeneration: A systematic review and meta-analysis of animal studies.Theranostics · 2022Pooled it
- Natural Product-Derived Vesicles in Nanotherapeutics.Journal of extracellular vesicles · 2026Review
- No Dye Fits All: Illuminating the Challenges of Fluorescent Extracellular Vesicle Labeling.Journal of extracellular vesicles · 2026Article
- Comparative Analysis of Methodological Aspects of the Study of Extracellular Vesicles and Extracellular Mitochondria: From Isolation to Internalization.Current issues in molecular biology · 2026Review
- From Uptake to Therapeutic Function in Engineered Exosome Delivery Systems.Research (Washington, D.C.) · 2026Review
- Benzimidazole-Derived B2 as a Fluorescent Probe for Bacterial Outer Membrane Vesicle (OMV) Labeling: Integrating DFT, Molecular Dynamics, Flow Cytometry, and Confocal Microscopy.International journal of molecular sciences · 2025Article
- A simplified and efficient method for isolating small extracellular vesicles for comparative and comprehensive translational research.Scientific reports · 2025Article
- Non-Specific Particle Formation During Extracellular Vesicle Labelling With the Lipophilic Membrane Dye PKH26.Journal of extracellular vesicles · 2025Article
- Harnessing extracellular vesicle heterogeneity for diagnostic and therapeutic applications.Nature nanotechnology · 2025Review
- Recent Advancements in Imaging Techniques for Individual Extracellular Vesicles.Molecules (Basel, Switzerland) · 2024Review
- Direct imaging with multidimensional labelling and high-content analysis allows quantitative categorization and characterizations of individual small extracellular vesicles and nanoparticles (sEVPs).Journal of extracellular vesicles · 2024Article
- Tracking Small Extracellular Vesicles Using a Minimally Invasive PicoGreen Labeling Strategy.ACS applied bio materials · 2024Article
- Shining a light on fluorescent EV dyes: Evaluating efficacy, specificity and suitability by nano-flow cytometry.Journal of extracellular biology · 2024Article
- An aptamer-guided fluorescence polarisation platform for extracellular vesicle liquid biopsy.Journal of extracellular vesicles · 2024Article
- Recent advances in chemical biology tools for protein and RNA profiling of extracellular vesicles.RSC chemical biology · 2024Review
- The extracellular vesicle proteomes of Sorghum bicolor and Arabidopsis thaliana are partially conserved.Plant physiology · 2024Article
- Factors to consider before choosing EV labeling method for fluorescence-based techniques.Frontiers in bioengineering and biotechnology · 2024Review
- Molecular mapping of neuronal architecture using STORM microscopy and new fluorescent probes for SMLM imaging.Neurophotonics · 2024Article
- Extracellular Vesicles as "Very Important Particles" (VIPs) in Aging.International journal of molecular sciences · 2023Review
- Characterization of the extracellular vesicles, ultrastructural morphology, and intercellular interactions of multiple clinical isolates of the brain-eating amoeba,Frontiers in microbiology · 2023Article
Corrections and comments
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Authors and funding
8 authors.
Funding
Abstract
Assessing genuine extracellular vesicle (EV) uptake is crucial for understanding the functional roles of EVs. This study measured the bona fide labelling of EVs utilising two commonly used fluorescent dyes, PKH26 and C5-maleimide-Alexa633. MCF7 EVs tagged with mEmerald-CD81 were isolated from conditioned media by size exclusion chromatography (SEC) and characterised using Nanoparticle Tracking Analysis (NTA), Transmission Electron Microscopy (TEM), MACsPlex immunocapture assay and immunoblots. These fluorescently tagged EVs were subsequently stained with C5-maleimide-Alexa633 or PKH26, according to published protocols. Colocalisation of dual-labelled EVs was assessed by confocal microscopy and quantified using the Rank-Weighted Colocalisation (RWC) algorithm. We observed strikingly poor colocalisation between mEmerald-CD81-tagged EVs and C5-Maleimide-Alexa633 (5.4% ± 1.8) or PKH26 (4.6% ± 1.6), that remained low even when serum was removed from preparations. Our data confirms previous work showing that some dyes form contaminating aggregates. Furthermore, uptake studies showed that maleimide and mEmerald-CD81-tagged EVs can be often located into non-overlapping subcellular locations. By using common methods to isolate and stain EVs we observed that most EVs remained unstained and most dye signal does not appear to be EV associated. Our work shows that there is an urgent need for optimisation and standardisation in how EV researchers use these tools to assess genuine EV signals.
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