title: Control and manipulation of pathogens with an optical trap for live cell imaging of intercellular interactions.
abstract: The application of live cell imaging allows direct visualization of the dynamic interactions between cells of the immune system. Some preliminary observations challenge long-held beliefs about immune responses to microorganisms; however, the lack of spatial and temporal control between the phagocytic cell and microbe has rendered focused observations into the initial interactions of host response to pathogens difficult. This paper outlines a method that advances live cell imaging by integrating a spinning disk confocal microscope with an optical trap, also known as an optical tweezer, in order to provide exquisite spatial and temporal control of pathogenic organisms and place them in proximity to host cells, as determined by the operator. Polymeric beads and live, pathogenic organisms (Candida albicans and Aspergillus fumigatus) were optically trapped using non-destructive forces and moved adjacent to living cells, which subsequently phagocytosed the trapped particle. High resolution, transmitted light and fluorescence-based movies established the ability to observe early events of phagocytosis in living cells. To demonstrate the broad applicability of this method to immunological studies, anti-CD3 polymeric beads were also trapped and manipulated to form synapses with T cells in vivo, and time-lapse imaging of synapse formation was also obtained. By providing a method to exert fine control of live pathogens with respect to immune cells, cellular interactions can be captured by fluorescence microscopy with minimal perturbation to cells and can yield powerful insight into early responses of innate and adaptive immunity.
authors.affiliation: Division of Infectious Disease, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Jenny M
authors.initials: JM
authors.lastname: Tam
authors.forename: Carlos E
authors.initials: CE
authors.lastname: Castro
authors.forename: Robert J W
authors.initials: RJ
authors.lastname: Heath
authors.forename: Michael L
authors.initials: ML
authors.lastname: Cardenas
authors.forename: Ramnik J
authors.initials: RJ
authors.lastname: Xavier
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Lang
authors.forename: Jatin M
authors.initials: JM
authors.lastname: Vyas
mesh_terms: D000818:Animals; D001232:Aspergillus fumigatus / Q000378:metabolism; D017252:CD3 Complex / Q000378:metabolism; D002176:Candida albicans / Q000378:metabolism; D002450:Cell Communication; D002478:Cells, Cultured; D006801:Humans; D019169:Jurkat Cells; D051379:Mice; D008853:Microscopy / Q000295:instrumentation* / Q000379:methods*; D018613:Microscopy, Confocal / Q000379:methods; D008954:Models, Biological; D052898:Optical Tweezers; D010587:Phagocytosis; D011108:Polymers / Q000737:chemistry; D013601:T-Lymphocytes / Q000166:cytology / Q000382:microbiology*
chemical_list: D017252:CD3 Complex; D011108:Polymers