Fast, Bright and Reversible Rhodamine Tags for Live-Cell Imaging

  1. Julian Kompa
  2. Lars Jeremy Dornfeld
  3. Nicola Porzberg
  4. Soohyen Jang
  5. Simon Hans Lilje
  6. Claudia Catapano
  7. David Jocher
  8. Lukas Merk
  9. Silja Zedlitz
  10. Runyu Mao
  11. Jonas Wilhelm
  12. Marina Dietz
  13. Miroslaw Tarnawski
  14. Julien Hiblot
  15. Mike Heilemann
  16. Kai Johnsson
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Abstract

We present Rho-tag and SiR-tag, engineered protein tags derived from bacterial multidrug-resistance proteins that bind unsubstituted (silicon-) rhodamines with nanomolar affinity, enabling fast, reversible, and fluorogenic protein labeling. In live cells, Rho-tag labeling occurs within seconds — faster than HaloTag7 — and the tags are compatible with super-resolution methods like STED, SMLM, and MINFLUX. The high specificity of Rho-tag and SiR-tag for unsubstituted rhodamines allows their use alongside HaloTag7 and SNAP-tag. In vivo applications are demonstrated by efficient neuronal labeling in zebrafish larvae.

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