Ultraviolet Photostability Improvement for Autofluorescence Correlation Spectroscopy on Label-Free Proteins
The natural autofluorescence of proteins in the UV is appealing to get the detailed information from single molecule data without requiring a potentially disturbing external fluorescent label. However, proteins feature significantly lower autofluorescence brightness and photostabilities than conventional fluorescent dyes. This issue has largely prevented so far the detection of label-free proteins in the ultraviolet.
In a recent article in the Journal of Physical Chemistry Letters, we use a dedicated combination of oxygen scavengers and reducing agents to promote the protein photostability, reduce the photobleaching probability and improve the net UV autofluorescence signal.
- This is the first time that different photostability improvement strategies are reported and quantitatively assessed for label-free proteins in the ultraviolet range.
- We show that the underlying photochemical concepts initially derived for organic fluorescent dyes are still valid for protein autofluorescence in the UV, and therefore appear to be quite general.
- Fluorescence correlation spectroscopy (FCS) is demonstrated on label-free streptavidin proteins containing only 24 tryptophan residues, 6.5× less than the current state-of-the-art.
Also freely available on ArXiv 2002.09761