Quantifying EGFR endosomal recycling via immunofluorescence in breast cancer cells

Ana Lonic; Winona Onglao; Yeesim Khew-Goodall

doi:10.1016/j.xpro.2022.101305

Quantifying EGFR endosomal recycling via immunofluorescence in breast cancer cells

STAR Protoc. 2022 Apr 13;3(2):101305. doi: 10.1016/j.xpro.2022.101305. eCollection 2022 Jun 17.

Authors

Ana Lonic¹, Winona Onglao^{1

2}, Yeesim Khew-Goodall^{1

3

2}

Affiliations

¹ Centre for Cancer Biology, An Alliance of SA Pathology and the University of South Australia, Adelaide, SA, Australia.
² Department of Molecular and Biomedical Science, School of Biological Sciences, Faculty of Sciences, University of Adelaide, Adelaide, SA, Australia.
³ The Discipline of Medicine, Adelaide Medical School, Faculty of Health and Medical Sciences, University of Adelaide, Adelaide, SA, Australia.

Abstract

Previously published protocols for quantification of endosomal recycling are limited by the use of radioactive reagents, washing of cells in reducing buffers, or the requirement for large numbers of cells. Here, we describe a protocol for quantification of endosomal recycling using immunofluorescence that is optimized for EGFR in BT-549 breast cancer cells but could be applied to other RTKs and cell lines. Our protocol enables quick assessment of recycling and uses a relatively small number of cells. For complete details on the use and execution of this protocol, please refer to Lonic et al. (2021).

Keywords: Cancer; Cell Biology; Microscopy; Signal Transduction.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Breast Neoplasms* / metabolism
Endosomes / metabolism
ErbB Receptors* / metabolism
Female
Fluorescent Antibody Technique
Humans
Staining and Labeling

Substances

EGFR protein, human
ErbB Receptors