A cutting-edge Technology

The Science Behind

Our technology is continuously fueled by the expertise detailed in our academic papers, published and discussed in international journals.

FAST AT A GLANCE

The Fluorescence-Activating and Absorption-Shifting Tag

The Twinkle Factory labeling technology enables the specific fluorescent labeling of any protein of interest (POI). It is based on the instantaneous formation of a fluorescent molecular assembly between the small (14 kDa), genetically encoded, protein tag FAST and various fluorogenic ligands (TFFluorogens).

TFFluorogens are dark in water and strongly fluoresce only when bound to FAST, enabling to detect and image FAST-tagged proteins with high contrast without the need of washing the excess of fluorogenic ligands. The labeling of FAST-tagged proteins with a TFFluorogen is non-covalent and can be reversed if necessary by washing.

Furthermore, FAST does not require molecular oxygen for being fluorescent, unlike fluorescent proteins, enabling to fluorescently label proteins in weakly oxygenated or anaerobic environments, helpful for biofilm imaging, or for any anaerobe imaging, e.g., Clostridium.

By using different TFFluorogens, the spectral properties of the FAST-tagged protein can be changed without the need to switch protein tags, providing an experimental versatility not encountered with fluorescent proteins.

The labeling of FAST-tagged proteins can be furthermore restricted to the cell-surface by using TFFluorogens unable to cross efficiently the cell membrane, enabling the study of protein trafficking at the membrane with various fluorimetric techniques, even including non-imaging flow cytometry.

The use of the Twinkle Factory labeling technology implies cloning and expression of the FAST-tagged protein, and labeling of the resulting fusion with the TFFluorogen of choice. Note that proteins of interest can be expressed with FAST as either an N- or a C-terminal fusion.

GO FURTHER

Academic Publications

Since the seminal publication in 2016 about FAST by Gautier et al. (2016) and its presentations in workshops and conferences, there has been numerous related articles by the Twinkle team, and by researchers from all over the world.

Here is a list of these publications.

By the Twinkle’s team

  • 2019. A split fluorescent reporter with rapid and reversible complementationNature Communications, 10, 2822 (2019) A. G. Tebo & A. Gautier
  • 2018. Fluorogenic probing of membrane protein trafficking. Bioconjugate Chemistry 29, 1823–1828 (2018). C. Li, A. Mourton, M.-A. Plamont, V. Rodrigues, I. Aujard, M. Volovitch, T. Le Saux, F. Perez, S. Vriz, L. Jullien, A. Joliot & A. Gautier
  • 2018. Circularly permuted fluorogenic proteins for the design of modular biosensors. ACS Chemical Biology 13, 2392–2397 (2018). A. G. Tebo, F. M. Pimenta, M. Zoumpoulaki, C. Kikuti, H. Sirkia, M.-A. Plamont, A. Houdusse & A. Gautier
  • 2018. Improved chemical-genetic fluorescent markers for protein imaging in living cellsBiochemistry 57, 5648–5653 (2018). A. G. Tebo, F.M. Pimenta, Y. Zhang & A. Gautier
  • 2018. Spying on cells with chemical-genetic hybrids (Espionner les cellules avec des hybrides chémogénétiques). L’Actualité Chimique 435, 31–35 (2018). A. Gautier
  • 2017. Dynamic multi-color protein labeling in living cells. Chemical Science 8, 5598–5605 (2017). C. Li, M.-A. Plamont, H. Sladitschek, V. Rodrigues, I. Aujard, P. Neveu, T. Le Saux, L. Jullien & A. Gautier
  • 2016. A small fluorescence-activating and absorption-shifting tag for tunable protein imaging in vivo. Proceedings of the National Academy of Sciences (PNAS) 113(3), 497–502 (2016). M.-A. Plamont, E. Billon-Denis, S. Maurin, C. Gauron, F. M. Pimenta, C. G. Specht, J. Shi, J. Querard, B. Pan, J. Rossignol, K. Moncoq, N. Morellet, M. Volovitch, E. Lescop, Y. Chen, A. Triller, S. Vriz, T. Le Saux, L. Jullien & A. Gautier

About the FAST technology, from other researchers

  • 2019. A strongly fluorescing anaerobic reporter and protein-tagging system for Clostridium organisms based on the Fluorescence-Activating and Absorption-Shifting Tag (FAST) protein. Appl. Environ. Microbiol., AEM-00622 (2019). H. E. Streett, K. M. Kalis & E. T. Papoutsakis
  • 2019. Red‐shifted substrates for FAST fluorogen‐activating protein based on the GFP‐like chromophores. Chemistry–A European Journal (2019). N. V. Povarova, S. O. Zaitseva, N. S. Baleeva, A. Y. Smirnov, I. N. Myasnyanko, M. B. Zagudaylova, … & A. S. Mishin
  • 2019. Single-molecule localization microscopy with the Fluorescence-Activating and absorption-Shifting Tag (FAST) system. ACS Chemical Biology (2019). E. M. Smith, A. Gautier & E. M. Puchner
  • 2019. Live cell super resolution imaging by radial fluctuations using fluorogen binding tags. Nanoscale 11(8), 3626-3632 (2019). M. Venkatachalapathy, V. Belapurkar, M. Jose, A. Gautier & D. Nair
  • 2018. Engineering Clostridium organisms as microbial cell-factories: challenges & opportunities. Metabolic engineering 50, 173-191 (2018). K. Charubin, R. K. Bennett, A. G. Fast & E. T. Papoutsakis
  • 2018. The inducible chemical-genetic fluorescent marker FAST outperforms classical fluorescent proteins in the quantitative reporting of bacterial biofilm dynamics. Scientific reports 8(1), 10336 (2018). A. Monmeyran, P. Thomen, H. Jonquière, F. Sureau, C. Li, M.-A. Plamont, … & N. Henry.
  • 2017. High-throughput, image-based screening of pooled genetic-variant libraries. Nature methods, 14(12), 1159 (2017). G. Emanuel, J. R. Moffitt & X. Zhuang

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