Journal
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
Volume 21, Issue 23, Pages -Publisher
MDPI
DOI: 10.3390/ijms21239008
Keywords
EGFR; HER2; electron microscopy; single molecule; correlative microscopy; dual labeling
Funding
- Else Kroner-Frisenius-Stiftung
- DFG [SFB1027]
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The epidermal growth factor receptor HER2 is overexpressed in 20% of breast cancer cases. HER2 is an orphan receptor that is activated ligand-independently by homodimerization. In addition, HER2 is able to heterodimerize with EGFR, HER3, and HER4. Heterodimerization has been proposed as a mechanism of resistance to therapy for HER2 overexpressing breast cancer. Here, a method is presented for the simultaneous detection of individual EGFR and HER2 receptors in the plasma membrane of breast cancer cells via specific labeling with quantum dot nanoparticles (QDs). Correlative fluorescence microscopy and liquid phase electron microscopy were used to analyze the plasma membrane expression levels of both receptors in individual intact cells. Fluorescent single-cell analysis of SKBR3 breast cancer cells dual-labeled for EGFR and HER2 revealed a heterogeneous expression for receptors within both the cell population as well as within individual cells. Subsequent electron microscopy of individual cells allowed the determination of individual receptors label distributions. QD-labeled EGFR was observed with a surface density of (0.5-5) x 10(1) QDs/mu m(2), whereas labeled HER2 expression was higher ranging from (2-10) x 10(2) QDs/mu m(2). Although most SKBR3 cells expressed low levels of EGFR, an enrichment was observed at large plasma membrane protrusions, and amongst a newly discovered cellular subpopulation termed EGFR-enriched cells.
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