Now you see me, now you don’t – labels for super-resolution microscopy in living cells

Session

Multimodal Microscopy

Authors

Dr Klaus Yserentant (2), Jonas Euchner (3), Stanimir Tashev (3), Dr Richard Wombacher (1), Prof Dirk-Peter Herten (3)

Affiliations

1. MPI for Medical Research
2. Univ. of California San Francisco
3. University of Birmingham

Keywords

fluorescent dyes, click chemistry, photo switching, single molecule spectroscopy, super-resolution microscopy

Abstract text

Over the past decade super-resolution microscopy has transformed the way we can study protein complexes within their cellular context. Aside of technical improvements, the spectroscopic properties of the fluorescence labels still play an important role in further progressing the field. In this context, we have explored the use of different quenching mechanisms to control the spectroscopic states of fluorescent labels. (1,2) In our work, we have shown that this can not only be used as an alternative approach in single molecule localization microscopy (SMLM) but furthermore to activate the label when binding to its target, thereby, improving the specificity of the fluorescence signal and enabling SMLM experiments in living cells. (3) Furthermore, I will discuss more recent work in which we engineered fluorescent labels that allow wash-free labelling experiments for fluorescence and super-resolution fluorescence microscopy. (4) Moreover, as many of these approaches use bio-orthogonal click chemistry, I will also present our recent study of the tetrazine click-reaction with trans-cyclooctene which is frequently used for bio-orthogonal labelling. (5)

References

1. Schwering M, Kiel A, Kurz A, Lymperopoulos K, Sprodefeld A, Kramer R, et al. Far-field nanoscopy with reversible chemical reactions. Angew Chem Int Ed. 2011;50(13):2940-5.
2. Brox D, Schwering M, Engelhardt J, Herten DP. Reversible chemical reactions for single-color multiplexing microscopy. Chemphyschem. 2014;15(11):2331-6.
3. 1Werther P, Yserentant K, Braun F, Kaltwasser N, Popp C, Baalmann M, et al. Live-Cell Localization Microscopy with a Fluorogenic and Self-Blinking Tetrazine Probe. Angew Chem Int Ed. 2020;59(2):804-10.
4. Werther P, Yserentant K, Braun F, Grussmayer K, Navikas V, Yu M, et al. Bio-orthogonal Red and Far-Red Fluorogenic Probes for Wash-Free Live-Cell and Super-resolution Microscopy. Acs Central Sci. 2021;7(9):1561-71.
5. Hild F, Werther P, Yserentant K, Wombacher R, Herten D-P. A dark intermediate in the fluorogenic reaction between tetrazine fluorophores and trans-cyclooctene. Biophys Rep. 2022;2(4):100084.