Labeling of a Mutant Estrogen Receptor with an Affimer in a Breast Cancer Cell Line

Pin Ren, Christian Tiede, Sean W. Fanning, Thomas Adams, Valerie Speirs, Erik R. Nelson, Changfeng Cheng, Terry W. Moore, Geoffrey L. Greene, Darren C. Tomlinson, Paul R. Selvin* (Corresponding Author)

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

1 Citation (Scopus)


Mutations of the intracellular Estrogen Receptor alpha (ER) is implicated in 70% of breast cancers. It is therefore of considerable interest to image various mutants (L536S, Y537S, D538G) in living cancer cell lines, particularly as a function of various anti-cancer drugs. We therefore developed a small (13kD) Affimer, which, after fluorescent labeling, is able to efficiently label ER by traveling through temporary pores in the cell membrane, created by the toxin Streptolysin O. The Affimer, selected by a phage display, predominantly labels the Y537S mutant and can tell the difference between L536S and D538G mutants. The vast majority of Affimer-ERY537S is in the nucleus and is capable of an efficient, unrestricted navigation to its target DNA sequence, as visualized by single molecule fluorescence. The Affimer can also differentiate the effect of selective estrogen receptor modulators (SERMs). More generally, this is an example of a small binding reagent—an Affimer protein—that can be inserted into living cells with minimal perturbation and high efficiency, to image an
endogenous protein.
Original languageEnglish
Pages (from-to)3651-3662
Number of pages12
JournalBiophysical Journal
Issue number19
Early online date30 Jun 2022
Publication statusPublished - 4 Oct 2022


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