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Home List of Titles Quantifying subcellular distribution of fluorescent fusion proteins in cells migrating within tissues
Please use this identifier to cite or link to this item: http://hdl.handle.net/1959.3/94886
- Quantifying subcellular distribution of fluorescent fusion proteins in cells migrating within tissues
- Melichar, Heather J.; Li, Ou; Herzmark, Paul; Padmanabhan, Raghav K.; Oliaro, Jane; Ludford-Menting, Mandy J.; Bousso, Philippe; Russell, Sarah M.; Roysam, Badrinath; Robey, Ellen A.
- The movement of proteins within cells can provide dynamic indications of cell signaling and cell polarity, but methods are needed to track and quantify subcellular protein movement within tissue environments. Here we present a semiautomated approach to quantify subcellular protein location for hundreds of migrating cells within intact living tissue using retrovirally expressed fluorescent fusion proteins and time-lapse two-photon microscopy of intact thymic lobes. We have validated the method using GFP-PKCζ, a marker for cell polarity, and LAT-GFP, a marker for T-cell receptor signaling, and have related the asymmetric distribution of these proteins to the direction and speed of cell migration. These approaches could be readily adapted to other fluorescent fusion proteins, tissues and biological questions.
- Publication type
- Journal article
- Immunology and Cell Biology, Vol. 89, no. 4 (May/Jun 2011), pp. 549-557
- Publication year
- FOR Code(s)
- 1107 Immunology
- Cells migration; Fluorescent fusion proteins; Subcellular distribution; Tissues
- Nature Publishing Group
- Publisher URL
- Copyright © 2011.
- Additional information
- Supported by the NIH (AI064227, AI065537, AI065831 and EB005157), the HFSP, the National Health and Medical Research Centre, and the California Institute for Regenerative Medicine through a postdoctoral fellowship (Training Grant Number T1-00007).
- Peer reviewed