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  A Catalytic Antibody Produces Fluorescent Tracers of Gap Junction Communication in Living Cells

Subauste, M. C., List, B., Guan, X., Hahn, K. M., Lerner, R. A., & Gilula, N. B. (2001). A Catalytic Antibody Produces Fluorescent Tracers of Gap Junction Communication in Living Cells. The Journal of Biological Chemistry, 276(52), 49164-49168. doi:10.1074/jbc.M105700200.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0014-A4D3-B Version Permalink: http://hdl.handle.net/21.11116/0000-0002-F126-3
Genre: Journal Article

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 Creators:
Subauste, M. Cecilia1, Author
List, Benjamin2, 3, Author              
Guan, Xiaojun1, Author
Hahn, Klaus M.1, Author
Lerner, Richard A. 2, Author
Gilula, Norton B.1, Author
Affiliations:
1Dept. of Cell Biology, Scripps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, California 92037, ou_persistent22              
2Dept. of Chemistry, The Scipps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, California 92037, ou_persistent22              
3Research Department List, Max-Planck-Institut für Kohlenforschung, Max Planck Society, ou_1445585              

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 Abstract: The antibody 38C2 efficiently catalyzed a retro-Michael reaction to convert a novel, cell-permeable fluorogenic substrate into fluorescein within living cells. In vitro, the antibody converted the substrate to fluorescein with ak cat of 1.7 × 10−5s−1 and a catalytic proficiency (k cat /k uncat Km ) of 1.4 × 1010 M −1(Km = 7 μm). For hybridoma cells expressing antibody or Chinese Hamster Ovarian (CHO) cells injected with antibody, incubation of the substrate in the extracellular medium resulted in bright intracellular fluorescence distinguishable from autofluorescence or noncatalyzed conversion of substrate. CHO cells loaded with antibody were 12 times brighter than control cells, and more than 85% of injected cells became fluorescent. The fluorescein produced by the antibody traveled into neighboring cells through gap junctions, as demonstrated by blocking dye transfer using the gap junction inhibitor oleamide. The presence of functional gap junctions in CHO cells was confirmed through oleamide inhibition of lucifer yellow transfer. These studies demonstrate the utility of the intracellular antibody reaction, which could generate tracer dyes in specific cells within complex multicellular environments simply by bathing the system in substrate.

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Language(s): eng - English
 Dates: 2001-06-202001-10-172001-12-28
 Publication Status: Published in print
 Pages: 5
 Publishing info: -
 Table of Contents: -
 Rev. Method: Peer
 Identifiers: DOI: 10.1074/jbc.M105700200
 Degree: -

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Title: The Journal of Biological Chemistry
  Other : JBC
Source Genre: Journal
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Publ. Info: Baltimore, etc. : American Society for Biochemistry and Molecular Biology [etc.]
Pages: - Volume / Issue: 276 (52) Sequence Number: - Start / End Page: 49164 - 49168 Identifier: ISSN: 0021-9258
CoNE: https://pure.mpg.de/cone/journals/resource/954925410826_1