Fluorescent Protein Labeling:
The Twinkle Factory labeling technology enables the specific fluorescent protein labeling for any protein of interest (POI). It is based on the instantaneous formation of a fluorescent molecular assembly between the protein FAST-Tag and various fluorogenic ligands (TFFluorogens).
This technology displays several specificities compared to conventional fluorescent proteins commonly used for protein tagging:
FAST proteins are only 125 amino acids long (14KDa) compared to larger GFP-based (238 amino acids) or mCherry (235 amino acids). They can be fused to the N- or C-terminus of any protein of interest.
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.
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 with a TFFluorogen is non-covalent and can be reversed if necessary by washing.
The labeling of FAST-tagged proteins can be furthermore restricted to the cell surface by using TFFluorogens unable to cross efficiently the cell membrane (TFAmber-NP), hence enabling the study of protein trafficking at the membrane with various fluorimetric techniques, even including non-imaging flow cytometry.
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.
How to choose your Tag-FAST proteins?
You will find below which FAST sequence can be used with which TFFluorogens. Absorption/Emission wavelength (nm) are indicated in each case.