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Methods Cell Biol. 2016;131:21-90. doi: 10.1016/bs.mcb.2015.07.001. Epub 2015 Sep 2.

Live-cell imaging of neurofilament transport in cultured neurons.

Author information

1
Department of Neuroscience, The Ohio State University, Columbus, OH, USA.

Abstract

Neurofilaments, which are the intermediate filaments of nerve cells, are space-filling cytoskeletal polymers that contribute to the growth of axonal caliber. In addition to their structural role, neurofilaments are cargos of axonal transport that move along microtubule tracks in a rapid, intermittent, and bidirectional manner. Though they measure just 10nm in diameter, which is well below the diffraction limit of optical microscopes, these polymers can reach 100 μm or more in length and are often packed densely, just tens of nanometers apart. These properties of neurofilaments present unique challenges for studies on their movement. In this article, we describe several live-cell fluorescence imaging strategies that we have developed to image neurofilament transport in axons of cultured neurons on short and long timescales. Together, these methods form a powerful set of complementary tools with which to study the axonal transport of these unique intracellular cargos.

KEYWORDS:

Axonal transport; Electroporation; Fluorescence microscopy; Fluorescent fusion protein; Kymograph analysis; Lipofection; Live-cell imaging; Magnetofection; Microinjection; Neurofilament; Photoactivation; Photobleaching; Primary cell culture; Time-lapse imaging; Transfection

PMID:
26794508
PMCID:
PMC6007976
DOI:
10.1016/bs.mcb.2015.07.001
[Indexed for MEDLINE]
Free PMC Article

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