VIDEO DROP ナノ粒子イメージングアナライザー 文献


The von Willebrand factor stamps plasmatic extracellular vesicles from glioblastoma patients
Quentin Sabbagh, Gwennan André‑Grégoire, Carolina Alves‑Nicolau, Aurélien Dupont, Nicolas Bidère, Emmanuel Jouglar, Laëtitia Guével, Jean‑Sébastien Frénel & Julie Gavard.
Scientific Reports volume 11, Article number: 22792 (2021)


Glioblastoma is a devastating tumor of the central nervous system characterized by a poor survival and an extremely dark prognosis, making its diagnosis, treatment and monitoring highly challenging. Numerous studies have highlighted extracellular vesicles (EVs) as key players of tumor growth, invasiveness and resistance, as they carry and disseminate oncogenic material in the local tumor microenvironment and at distance. However, whether their quality and quantity reflect individual health status and changes in homeostasis is still not fully elucidated. Here, we separated EVs from plasma collected at different time points alongside with the clinical management of GBM patients. Our findings confirm that plasmatic EVs could be separated and characterized with standardized protocols, thereby ensuring the reliability of measuring vesiclemia, i.e. extracellular vesicle concentration in plasma. This unveils that vesiclemia is a dynamic parameter, which could be reflecting tumor burden and/or response to treatments. Further label-free liquid chromatography tandem mass spectrometry unmasks the von Willebrand Factor (VWF) as a selective protein hallmark for GBM-patient EVs. Our data thus support the notion that EVs from GBM patients showed differential protein cargos that can be further surveyed in circulating EVs, together with vesiclemia.

The Necroptosis Effector MLKL drives Small Extracellular Vesicle Release and Tumour Growth in Glioblastoma
Gwennan André-Grégoire, Tiphaine Douanne, An Thys, Clément Maghe, Kathryn Jacobs, Cyndie Ballu, Kilian Trillet, Ignacio Busnelli, Vincent Hyenne, Jacky G Goetz, Nicolas Bidère, Julie Gavard
bioRxiv January 14, 2021.
doi: https://doi.org/10.1101/2021.01.12.426398


Extracellular vesicles (EVs) are lipid-based nano-sized particles that convey biological material from donor to recipient cells. They play key roles in tumour progression, notably in glioblastoma in which the subpopulation of Glioblastoma Stem-like Cells (GSCs) might represent a meaningful source of tumour-derived EVs. However, the mechanisms involved in the production and release of EVs by GSCs are still poorly understood. Here, we report the identification of MLKL, a crucial effector of cell death by necroptosis, as a regulator of the constitutive secretion of small EVs from GSCs. The targeting of MLKL by genetic, protein depletion or chemical approaches alters endosomal trafficking and EV release and reduces GSC expansion in vitro. This function ascribed to MLKL appears independent of its role during necroptosis. In vivo, pharmacological inhibition of MLKL triggers a reduction of both the tumour burden in xenografted mice and of the level of plasmatic EVs. This work reinforces the idea of a non-deadly role for MLKL in endosomal trafficking and suggests that interfering with EV biogenesis is a promising therapeutic option to sensitize glioblastoma cells to death.


Experimental Evaluation of an Interferometric Light Microscopy Particle Counter for Titering and Characterization of Virus Preparations
Vesa Turkki,Elisa Alppila,Seppo Ylä-Herttuala,and Hanna P.Lesch
Viruses.2021 May 19.doi:10.3390/v13050939

レンチ、アデノ、バキュロウイルスのサンプルを異なる濃度で分析し、測定値を従来の滴定および特性評価方法(TRPS,NTA,nFCM,VirusCounter3100)と比較した。一部の精製されていないサンプルや小さなウイルスは、特性評価が不可能な場合や標準曲線やバックグラウンド減算法を使用しなければならない場合があり、分析の期間が長くなるが、VIDEODROPの干渉光顕微鏡(ILM)法は、未精製でも従来の光学顕微鏡の回折限界内サイズの粒子を検出することができる。 したがって、VIDEODROPは高濃度の精製ウイルス製剤を測定する際に特に有用である。


Virus particle concentration is a critical piece of information for virology, viral vaccines and gene therapy research. We tested a novel nanoparticle counting device, “Videodrop”, for its efficacy in titering and characterization of virus particles. The Videodrop nanoparticle counter is based on interferometric light microscopy (ILM). The method allows the detection of particles under the diffraction limit capabilities of conventional light microscopy. We analyzed lenti-, adeno-, and baculovirus samples in different concentrations and compared the readings against traditional titering and characterization methods. The tested Videodrop particle counter is especially useful when measuring high-concentration purified virus preparations. Certain non-purified sample types or small viruses may be impossible to characterize or may require the use of standard curve or background subtraction methods, which increases the duration of the analysis. Together, our testing shows that Videodrop is a reasonable option for virus particle counting in situations where a moderate number of samples need to be analyzed quickly.

Keywords: automated nanoparticle counter; titering; viruses; virus vectors; interferometric light microscopy; particle count

Analysis of viromes and microbiomes from pig fecal samples reveals that phages and prophages are not vectors of antibiotic resistance genes
Maud Billaud, Quentin Lamy-Besnier, Julien Lossouarn, Elisabeth Moncaut, Moira B. Dion, Sylvain Moineau, Fatoumata Traoré, Emmanuelle Le Chatelier, Catherine Denis, Jordi Estelle, Caroline Achard, Olivier Zemb, Marie-Agnès Petit
bioRxiv May 20, 2021. 
doi: https://doi.org/10.1101/2021.05.20.444921


Understanding the transmission of antibiotic resistance genes (ARGs) is critical for human health. For this, it is necessary to identify which type of mobile genetic elements is able to spread them from animal reservoirs into human pathogens. Previous research suggests that in pig feces, ARGs may be encoded by bacteriophages. However, convincing proof for phage-encoded ARGs in pig viromes is still lacking, because of bacterial DNA contaminating issues. We collected 14 pig fecal samples and performed deep sequencing on both highly purified viral fractions and total microbiota, in order to investigate phage and prophage-encoded ARGs. We show that ARGs are absent from the genomes of active, virion-forming phages (below 0.02% of viral contigs from viromes), but present in three prophages, representing 0.02% of the viral contigs identified in the microbial dataset. However, the corresponding phages were not detected in the viromes, and their genetic maps suggest they might be defective. Furthermore, our dataset allows for the first time a comprehensive view of the interplay between prophages and viral particles.

Viral metagenomic analysis of the cheese surface:A comparative study of rapid procedures for extracting viral particles.
Eric Dugat-Bony, Julien Lossouarn, Marianne De Paepe, Anne-Sophie Sarthou, Yasmina Fedala, Marie-Agnès Petit, Stéphane Chaillou Science Direct January 11, 2019. 


The structure and functioning of microbial communities from fermented foods, including cheese, have been extensively studied during the past decade. However, there is still a lack of information about both the occurrence and the role of viruses in modulating the function of this type of spatially structured and solid ecosystems. Viral metagenomics was recently applied to a wide variety of environmental samples and standardized procedures for recovering viral particles from different type of materials has emerged. In this study, we adapted a procedure originally developed to extract viruses from fecal samples, in order to enable efficient virome analysis of cheese surface. We tested and validated the positive impact of both addition of a filtration step prior to virus concentration and substitution of purification by density gradient ultracentrifugation by a simple chloroform treatment to eliminate membrane vesicles. Viral DNA extracted from the several procedures, as well as a vesicle sample, were sequenced using Illumina paired-end MiSeq technology and the subsequent clusters assembled from the virome were analyzed to assess those belonging to putative phages, plasmid-derived DNA, or even from bacterial chromosomal DNA. The best procedure was then chosen, and used to describe the first cheese surface virome, using Epoisses cheese as example. This study provides the basis of future investigations regarding the ecological importance of viruses in cheese microbial ecosystems.

Utilization of interferometric light microscopy for the rapid analysis of virus abundance in a river.
Céline Roose-Amsaleg, Yasmina Fedala, Catherine Vénien-Bryan, Josette Garnier, Albert-Claude Boccara, Martine Boccara
National Library of Medicine Mar 2, 2017.DOI: 10.1016/j.resmic.2017.02.004


There is a constant need for direct counting of biotic nanoparticles such as viruses to unravel river functioning. We used, for the first time in freshwater, a new method based on interferometry differentiating viruses from other particles such as membrane vesicles. In the French Marne River, viruses represented between 42 and 72% of the particles. A spring monitoring in 2014 revealed their increase (2.1 × 107 to 2.1 × 108 mL-1) linked to an increase in algal biomass and diversity of bacterial plankton. Predicted virus size distributions were in agreement with transmission electron microscopy analysis suggesting a dominance of large viruses (≥60 nm).

Keywords: Interferometric light microscopy; Membrane vesicle; River; Virus.

Full-field interferometry for counting and differentiating aquatic biotic nanoparticles:from laboratory to Tara Oceans.
Martine Boccara, Yasmina Fedala, Catherine Venien Bryan, Marc Bailly-Bechet, Chris Bowler, Albert Claude Boccara
National Library of Medicine Aug 29, 2016. DOI: 10.1364/BOE.7.003736


There is a huge abundance of viruses and membrane vesicles in seawater. We describe a new full-field, incoherently illuminated, shot-noise limited, common-path interferometric detection method that we couple with the analysis of Brownian motion to detect, quantify, and differentiate biotic nanoparticles. We validated the method with calibrated nanoparticles and homogeneous DNA or RNA viruses. The smallest virus size that we characterized with a suitable signal-to-noise ratio was around 30 nm in diameter. Analysis of Brownian motions revealed anisotropic trajectories for myoviruses.We further applied the method for vesicles detection and for analysis of coastal and oligotrophic samples from Tara Oceans circumnavigation.

Keywords: (010.0280) Remote sensing and sensors; (110.3175) Interferometric imaging; (120.5820) Scattering measurements; (170.4580) Optical diagnostics for medicine.


Highly Stable Polymer Coating on Silver Nanoparticles for Efficient Plasmonic Enhancement of Fluorescence
Ryo Kato, Mitsuhiro Uesugi, Yoshie Komatsu, Fusatoshi Okamoto, Takuo Tanaka, Fumihisa Kitawaki, and Taka-aki Yano.
ACS Omega 2022, 7, 5, 4286–4292 Publication Date:January 7, 2022


Surface coating of plasmonic nanoparticles is of huge importance to suppress fluorescence quenching in plasmon-enhanced fluorescence sensing. Herein, a one-pot method for synthesizing polymer-coated silver nanoparticles was developed using a functional polymer conjugated with disulfide-containing anchoring groups. The disulfides played a crucial role in covalently bonding polymers to the surface of the silver nanoparticles. The covalent bond enabled the polymer layer to form a long-term stable coating on the silver nanoparticles. The polymer layer coated was adequately thin to efficiently achieve plasmonic enhancement of fluorescence and also thick enough to effectively suppress quenching of fluorescence, achieving a huge net enhancement of fluorescence. The polymer-coated plasmonic nanoparticles are a promising platform for demonstrating highly sensitive biosensing for medical diagnostics.