IOVS
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

This Article
Right arrow Full Text (PDF)
Right arrow Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by McLaren, M. J.
Right arrow Articles by Li, C. Y.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by McLaren, M. J.
Right arrow Articles by Li, C. Y.

Investigative Ophthalmology & Visual Science, Vol 34, 317-326, Copyright © 1993 by Association for Research in Vision and Ophthalmology

ARTICLES AND REPORTS

Double fluorescent vital assay of phagocytosis by cultured retinal pigment epithelial cells

MJ McLaren, G Inana and CY Li
Department of Ophthalmology, Bascom Palmer Eye Institute, University of Miami School of Medicine, Florida 33101.

PURPOSE. The goal of this study was to develop the first vital assay system for in vitro analysis of phagocytosis of rod outer segments (ROS) by normal retinal pigment epithelial (RPE) cells and for study of the phagocytic defect in RPE of the Royal College of Surgeons (RCS) rat with inherited retinal degeneration. Required features included ability to directly visualize and quantitate the phagocytic process in living RPE cultures, and capability for subsequent quantitative analysis after fixation of the cells at any chosen time after incubation with ROS. METHODS. A double fluorescent method was designed, based on the process of phagosome-lysosome fusion. For vital staining of lysosomes, confluent cultures of rat RPE cells were incubated with sulforhodamine (SR), a red fluorescent lysosomotropic dye. SR-stained cultures were challenged with isolated rat ROS tagged with fluorescein isothiocyanate (FITC), a green fluorescent probe. RESULTS. This method was used to observe all phases of the phagocytic process in the living cells and the kinetics of ROS binding, ingestion, and phagosome-lysosome fusion were determined. Control studies showed no differences in binding, ingestion, or digestion of unstained versus FITC-stained ROS. Additionally, the phagocytic defect in dystrophic RCS rat RPE cells was confirmed using this technique. CONCLUSIONS. This relatively simple new method is useful in that it uses inexpensive, readily available reagents, it enables real-time analysis of phagocytosis experiments, and it does not require termination of the cultures for analysis of phagocytic ability.


This article has been cited by other articles:


Home page
 J. Biol. Chem.Home page
S. Qin and G. W. De Vries
{alpha}2 but Not {alpha}1 AMP-activated Protein Kinase Mediates Oxidative Stress-induced Inhibition of Retinal Pigment Epithelium Cell Phagocytosis of Photoreceptor Outer Segments
J. Biol. Chem., March 14, 2008; 283(11): 6744 - 6751.
[Abstract] [Full Text] [PDF]

Home page
 JGPHome page
A. L. Kindzelskii, V. M. Elner, S. G. Elner, D. Yang, B. A. Hughes, and H. R. Petty
Toll-Like Receptor 4 (TLR4) of Retinal Pigment Epithelial Cells Participates in Transmembrane Signaling in Response to Photoreceptor Outer Segments
J. Gen. Physiol., July 26, 2004; 124(2): 139 - 149.
[Abstract] [Full Text] [PDF]

Home page
 IOVSHome page
G. T. Higgins, J. H. Wang, P. Dockery, P. E. Cleary, and H. P. Redmond
Induction of Angiogenic Cytokine Expression in Cultured RPE by Ingestion of Oxidized Photoreceptor Outer Segments
Invest. Ophthalmol. Vis. Sci., April 1, 2003; 44(4): 1775 - 1782.
[Abstract] [Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Copyright © 1993 by the Association for Research in Vision and Ophthalmology