Photo-Switchable Cyanine Dyes for Super-Resolution Microscopy
Final Report Abstract
Super-resolution microscopy offers the unique opportunity to investigate cellular processes in living organisms. To ensure the acquisition of high-resolution images, photoactivatable dyes are required which can be turned from dark (OFF) to highly emissive (ON) states by irradiation with light. To date, several molecular switches have been invented, but most of them require ultraviolet irradiation to induce the switching process, which is a critical aspect for live cell imaging. In this project we worked on the design of organic photoswitches which can be switched between ON and OFF states by using visible light. When covalently linked to fluorophores they can quench the dye’s emission upon irradiation with visible light resulting in dark OFF states. A photochemical reaction after irradiation with visible light restores the emission of the dye and unveils the bright ON state. In this study we designed several dyads which were successfully incorporated in phospholipid membranes; lipids that are also found in cellular membranes. Moreover, we demonstrated that our samples can be switched between “ON” and “OFF” states with an exceptional high contrast. Furthermore, the samples can be used for super-resolution microscopy revealing a two-fold resolution enhancement compared to the confocal image. Despite the good results, we found that several aspects limited the resolution of our images: mechanical motion of the vesicle and poor fatigue resistance. Moreover, preliminary cell studies have shown that the dyads were randomly distributed inside the cells hampering a potential application in targeting cellular organelles or membranes. In further studies we applied a polarity-sensitive dye, which visualizes polarity changes in environments. Microscopy experiments demonstrated that the new dyad can be used in super-resolution microscopy revealing a two-fold resolution enhancement. Ongoing studies investigate how a change in polarity can be used to understand the order of lipid molecules in membranes and to localise these compounds in cellular compartments. Our results contribute to a better understanding of molecular photoswitches and how they perform in biological environments. Successful and failed experiments support scientists to modify the design of existing molecular switches towards ultra-high super-resolution images.
Publications
- Cells Undergo Major Changes in the Quantity of Cytoplasmic Organelles after Uptake of Gold Nanoparticles with Biologically Relevant Surface Coatings, ACS Nano, 2020, 14(2), 2248-2264
B. Kepsutlu, V. Wycisk, K. Achazi, S. Kapishnikov, A. J. Pérez-Berná, P. Guttmann, A. Cossmer, E. Pereiro, H. Ewers, M. Ballauff, G. Schneider, J. G. McNally
(See online at https://doi.org/10.1021/acsnano.9b09264) - Stereochemistry-Controlled Supramolecular Architectures of New Tetrahydroxy-Functionalised Amphiphilic Carbocyanine Dyes, Chemistry, 2020, 26(30), 6919-6934
B. Schade, A. K. Singh, V. Wycisk, J. L. Cuellar-Camacho, H. von Berlepsch, R. Haag, C. Böttcher
(See online at https://doi.org/10.1002/chem.201905745) - Super-resolution RESOLFT microscopy of lipid bilayers using a fluorophore-switch dyad Chem. Sci., 2020,11, 8955-8960
A. T. Frawley, V. Wycisk, Y. Xiong, S. Galiani, E. Sezgin, I. Urbančič, A. V. Jentzsch, K. G. Leslie, C. Eggeling and H. L. Anderson
(See online at https://doi.org/10.1039/d0sc02447c) - Aggregation of Amphiphilic Carbocyanines: Fluorination Favors Cylindrical Micelles over Bilayered Tubes, J. Phys. Chem. B, 2021, 125, 37, 10538-10550
A. K. Singh, B. Schade, V. Wycisk, C. Böttcher, R. Haag, H. von Berlepsch
(See online at https://doi.org/10.1002/chem.201905745)
