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Nanyang Technological University

Improved Lipofuscin Models and Quantification of Outer Segment Phagocytosis Capacity in Highly Polarized Human Retinal Pigment Epithelial Cultures

2.4K Views

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10:39 min

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April 14th, 2023

DOI :

10.3791/65242-v

April 14th, 2023

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Qitao Zhang¹, Gillian Autterson¹, Jason M. L. Miller¹^,²

¹Kellogg Eye Center, University of Michigan, Ann Arbor, ²Cellular and Molecular Biology Program, University of Michigan, Ann Arbor

Summary

This protocol describes a lipofuscin accumulation model in highly differentiated and polarized human retinal pigment epithelial (RPE) cultures and an improved outer segment (OS) phagocytosis assay to detect the total OS consumption/degradation capacity of the RPE. These methods overcome the limitations of previous lipofuscin models and classical pulse-chase outer segment phagocytosis assays.

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Chapters in this video

0:00

Introduction

2:42

Preparation of Photo‐Oxidized Outer Segment Tips and Fragments

6:27

Characterization of the Lipofuscin‐Like Granule Spectrum and Composition

8:26

Assessing the Effects of Lipofuscin‐Like Granules on RPE Phagocytosis: Total Consumptive Capacity

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