title: Labeling of immune cells for in vivo imaging using magnetofluorescent nanoparticles.
abstract: Observation of immune and stem cells in their native microenvironments requires the development of imaging agents to allow their in vivo tracking. We describe here the synthesis of magnetofluorescent nanoparticles for cell labeling in vitro and for multimodality imaging of administered cells in vivo. MION-47, a prototype monocrystalline iron oxide nanoparticle, was first converted to an intermediate bearing a fluorochrome and amine groups, then reacted with either HIV-Tat peptide or protamine to yield a nanoparticle with membrane-translocating properties. We describe how to assess optimal cell labeling with tests of cell phenotype and function. Synthesis of magnetofluorescent nanoparticles and cell-labeling optimization can be realized in 48 h, whereas nanoparticle uptakes and retention studies may generally take up to 120 h. Labeled cells can be detected by magnetic resonance imaging, fluorescence reflectance imaging, fluorescence-mediated tomography, confocal microscopy and flow cytometry, and can be purified based on their fluorescent or magnetic properties. The present protocol focuses on T-cell labeling but can be used for labeling a variety of circulating cells.
authors.affiliation: Center for Molecular Imaging Research, Massachusetts General Hospital, Charlestown, Massachusetts 02129, USA. mpittet@hms.harvard.edu
authors.forename: Mikael J
authors.initials: MJ
authors.lastname: Pittet
authors.forename: Filip K
authors.initials: FK
authors.lastname: Swirski
authors.forename: Frederick
authors.initials: F
authors.lastname: Reynolds
authors.forename: Lee
authors.initials: L
authors.lastname: Josephson
authors.forename: Ralph
authors.initials: R
authors.lastname: Weissleder
mesh_terms: D000818:Animals; D005290:Ferric Compounds / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry*; D008279:Magnetic Resonance Imaging; D008280:Magnetics; D051379:Mice; D053758:Nanoparticles / Q000737:chemistry*; D013194:Staining and Labeling / Q000379:methods*; D013601:T-Lymphocytes / Q000737:chemistry* / Q000166:cytology
chemical_list: D005290:Ferric Compounds; D005456:Fluorescent Dyes; C000499:ferric oxide