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Fluorescent low density lipoprotein for observation of dynamics of individual receptor complexes on cultured human fibroblasts.

Publication ,  Journal Article
Barak, LS; Webb, WW
Published in: J Cell Biol
September 1981

The visible wavelength excited fluorophore 3,3'-dioctadecylindocarbocyanine iodide (Dil[3]) was incorporated into human low density lipoprotein (LDL) to form the highly fluorescent LDL derivative dil(3)-LDL. Dil(3)-LDL binds to normal human fibroblasts and to human fibroblasts defective in LDL receptor internalization but does not bind to LDL receptor-negative human fibroblasts at 4 degrees C or 37 degrees C. It is internalized rapidly at 37 degrees C by normal fibroblasts and depresses the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoA reductase) in a manner similar to that of LDL. It is prevented from binding to the LDL receptor by an excess of unlabeled LDL or by heparin sulfate. Identical distributions of dil(3)-LDL are observed on cells by either indirect immunofluorescence with fluorescein-labeled antibody or directly by dil(3) fluorescence. Upwards of 45 molecules of dil(3) are incorporated per molecule of LDL without affecting binding to the receptor. This labeling renders individual molecules visible by their fluorescence and enables the derivative to be used in dynamic studies of LDL-receptor motion on living fibroblasts by standard fluorescence techniques at low LDL receptor density. Observations with this derivative indicate that the LDL-receptor complex is immobilized on the surface of human fibroblasts but, when free of this linkage, undergoes a Brownian motion consistent with theory.

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Published In

J Cell Biol

DOI

ISSN

0021-9525

Publication Date

September 1981

Volume

90

Issue

3

Start / End Page

595 / 604

Location

United States

Related Subject Headings

  • Temperature
  • Receptors, LDL
  • Receptors, Cell Surface
  • Quinolines
  • Microscopy, Fluorescence
  • Lipoproteins, LDL
  • Hydroxymethylglutaryl CoA Reductases
  • Humans
  • Heparin
  • Fluorescent Dyes
 

Citation

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Barak, L. S., & Webb, W. W. (1981). Fluorescent low density lipoprotein for observation of dynamics of individual receptor complexes on cultured human fibroblasts. J Cell Biol, 90(3), 595–604. https://doi.org/10.1083/jcb.90.3.595
Barak, L. S., and W. W. Webb. “Fluorescent low density lipoprotein for observation of dynamics of individual receptor complexes on cultured human fibroblasts.J Cell Biol 90, no. 3 (September 1981): 595–604. https://doi.org/10.1083/jcb.90.3.595.
Barak, L. S., and W. W. Webb. “Fluorescent low density lipoprotein for observation of dynamics of individual receptor complexes on cultured human fibroblasts.J Cell Biol, vol. 90, no. 3, Sept. 1981, pp. 595–604. Pubmed, doi:10.1083/jcb.90.3.595.

Published In

J Cell Biol

DOI

ISSN

0021-9525

Publication Date

September 1981

Volume

90

Issue

3

Start / End Page

595 / 604

Location

United States

Related Subject Headings

  • Temperature
  • Receptors, LDL
  • Receptors, Cell Surface
  • Quinolines
  • Microscopy, Fluorescence
  • Lipoproteins, LDL
  • Hydroxymethylglutaryl CoA Reductases
  • Humans
  • Heparin
  • Fluorescent Dyes