| Live cell imaging of paxillin in rolling neutrophils by dual-color quantitative dynamic footprinting. | |
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MedLine Citation:
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PMID: 21418380 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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OBJECTIVE: Neutrophil recruitment to sites of inflammation involves P-selectin-dependent rolling. qDF is a useful tool to visualize the topography of the neutrophil footprint as it interacts with the substrate. However, elucidating the role of specific proteins in addition to topography requires simultaneous visualization of two fluorochromes. METHODS: To validate DqDF, mouse neutrophils were labeled with the membrane dyes DiO and DiI and perfused into microchannels coated with P-selectin-Fc. Footprints of rolling neutrophils were recorded as two separate images, one for each fluorochrome. To assess the localization of the cytoskeletal protein paxillin, we applied DqDF to DiO-stained neutrophils of mice expressing an mCherry-paxillin fusion protein. RESULTS: The footprint topographies obtained from DiO and DiI in the plasma membrane were identical. The z-coordinates of the microvilli tips obtained with the two fluorochromes in the footprint were also identical. Paxillin was found to be localized to some, but not all ridges in the neutrophil footprint. CONCLUSIONS: Our data suggest that the spectral properties of the fluorochrome do not affect the results. DqDF will be useful for simultaneous visualization of two fluorochromes in the footprint of rolling cells. |
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Authors:
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Prithu Sundd; Edgar Gutierrez; Brian G Petrich; Mark H Ginsberg; Alex Groisman; Klaus Ley |
Publication Detail:
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Type: Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: Microcirculation (New York, N.Y. : 1994) Volume: 18 ISSN: 1549-8719 ISO Abbreviation: Microcirculation Publication Date: 2011 Jul |
Date Detail:
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Created Date: 2011-07-01 Completed Date: 2011-10-24 Revised Date: 2012-09-24 |
Medline Journal Info:
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Nlm Unique ID: 9434935 Medline TA: Microcirculation Country: United States |
Other Details:
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Languages: eng Pagination: 361-72 Citation Subset: IM |
Copyright Information:
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© 2011 John Wiley & Sons Ltd. |
Affiliation:
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Division of Inflammation Biology, La Jolla Institute for Allergy and Immunology, La Jolla, California, USA. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Animals Carbocyanines / pharmacology Fluorescent Dyes / pharmacology Leukocyte Rolling / physiology* Mice Mice, Transgenic Microscopy, Fluorescence Neutrophils / cytology, metabolism* Paxillin / genetics, metabolism* Recombinant Fusion Proteins / genetics, metabolism |
| Grant Support | |
ID/Acronym/Agency:
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EB02185/EB/NIBIB NIH HHS; GM64346/GM/NIGMS NIH HHS; R01 EB002185-06A2/EB/NIBIB NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Carbocyanines; 0/Fluorescent Dyes; 0/Paxillin; 0/Pxn protein, mouse; 0/Recombinant Fusion Proteins; 40957-95-7/3,3'-dioctadecylindocarbocyanine |
| Comments/Corrections | |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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