Optimized methods for imaging membrane nanotubes between T cells and trafficking of HIV-1.

Stefanie Sowinski, Juha-Matti Alakoskela, Clare Jolly, Daniel M Davis

Journal: Methods (San Diego, Calif.) 2011;53(1):27-33

PMID: 20382227

Full Text on Elsevier Science View on PubMed

Abstract

A wide variety of cell types, including immune cells, have been observed to frequently interact via transient, long-distance membrane connections. However, considerable heterogeneity in their structure, mode of formation and functional properties has emerged, suggesting the existence of distinct subclasses. Open-ended tunneling nanotubes allow for the trafficking of cytoplasmic material, e.g. endocytic vesicles, or the transmission of calcium signals. Closed-ended membrane nanotubes do not seamlessly connect the cytoplasm between two interacting cells and a junction exists within the nanotube or where the nanotube meets a cell body. Recent live cell imaging suggested that membrane nanotubes between T cells could present a novel route for HIV-1 transmission. Here, we describe detailed protocols for observing membrane nanotubes and HIV-1 trafficking by live cell fluorescence microscopy.

Address: Gladstone Institute of Virology and Immunology, San Francisco, CA 94158-2216, USA. [email protected]

DOI: 10.1016/j.ymeth.2010.04.002

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MeSH Terms: Cell Culture Techniques, Cell Membrane Structures, Cell Tracking, Fibronectins, Fixatives, Green Fluorescent Proteins, HIV-1, Humans, Immunological Synapses, Jurkat Cells, Microscopy, Confocal, Pseudopodia, Recombinant Fusion Proteins, T-Lymphocytes, gag Gene Products, Human Immunodeficiency Virus