Immunodetection of Extracellular Vesicles

Procedures were performed essentially as described [25 (link),32 (link),33 (link)]. Immunodetection was carried out with the following primary antibodies: mouse anti-human CD63 (Abcam ab23792), mouse anti-human CD9, or mouse anti-human CD81 (both from Dr E. Rubinstein, Université Paris-Sud, Institut André Lwoff, Villejuif, France). Secondary incubation was performed with a rabbit anti mouse Fc fragment (Dako Agilent Z0412), then grids were incubated with Protein A-Gold 10 nm (Cell Microscopy Center, Department of Cell Biology, Utrecht University). All samples were observed with a Tecnai Spirit electron microscope (FEI, Eindhoven, The Netherlands), and digital acquisitions were made with a numeric 4k CCD camera (Quemesa, Olympus, Münster, Germany). Images were analysed with iTEM software (EMSIS) and statistical studies were done with GraphPad Prism software (v8).

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Bergamelli M., Martin H., Aubert Y., Mansuy J.M., Marcellin M., Burlet-Schiltz O., Hurbain I., Raposo G., Izopet J., Fournier T., Benchoua A., Bénard M., Groussolles M., Cartron G., Tanguy Le Gac Y., Moinard N., D’Angelo G, & Malnou C.E. (2022). Human Cytomegalovirus Modifies Placental Small Extracellular Vesicle Composition to Enhance Infection of Fetal Neural Cells In Vitro. Viruses, 14(9), 2030.

Publication 2022

ab23792 Z0412 Tecnai Spirit electron microscope Quemesa

Antibodies Cell Digital Fc fragment Gold Human Microscope electron Microscopy Mouse Prism Protein a Rabbit

Corresponding Organization : Centre National de la Recherche Scientifique

Other organizations : Hôpital Purpan, Institut National Polytechnique de Toulouse, Institut de Pharmacologie et de Biologie Structurale, Institut Curie, Université Paris Cité, Association Francaise contre les Myopathies, Institut des Cellules Souches pour le Traitement et l'Étude des Maladies Monogéniques, Hôpital des Enfants, Hôpital Paule de Viguier, Laboratoire Epidémiologie et Analyses en Santé Publique : Risques, Maladies Chroniques et Handicaps, Université de Montpellier

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Variable analysis

independent variables

Procedures performed as described in references [25], [32], and [33]

dependent variables

Immunodetection of CD63, CD9, and CD81 proteins

control variables

Mouse anti-human CD63 (Abcam ab23792), mouse anti-human CD9, and mouse anti-human CD81 (from Dr E. Rubinstein, Université Paris-Sud, Institut André Lwoff, Villejuif, France) used as primary antibodies
Rabbit anti mouse Fc fragment (Dako Agilent Z0412) used as secondary antibody
Protein A-Gold 10 nm (Cell Microscopy Center, Department of Cell Biology, Utrecht University) used for incubation
Tecnai Spirit electron microscope (FEI, Eindhoven, The Netherlands) used for sample observation
Quemesa 4k CCD camera (Olympus, Münster, Germany) used for digital acquisitions
ITEM software (EMSIS) used for image analysis
GraphPad Prism software (v8) used for statistical studies

positive controls

Not specified

negative controls

Not specified

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