Differential Scanning Fluorimetry
Quickly characterize protein stability for construct and formulation screening
Speed-up stability studies
With differential scanning fluorimetry (DSF), you can quickly screen through many different proteins or formulations to find the most stable ones. Instead of waiting days or weeks for an accelerated, high temp isothermal study, or even longer for a real-time stability study, DSF gets you answers in just a few hours by evaluating how protein conformation changes in response to a thermal ramp.
Make the most of fluorescence
DSF works by measuring changes in fluorescence as a protein’s conformation changes. The fluorescence can come from a protein’s own tryptophan, tyrosine, and phenylalanine residues (intrinsic fluorescence) or from an added fluorophore such as SYPRO® Orange.
See the change
For proteins, the shape of the emitted fluorescence depends on the local environment around the fluorescent amino acids. The fluorescent spectrum of a folded protein with highly non-polar hydrophobic pockets will look different from the spectrum of an unfolded protein which has the fluorescent amino acids exposed to a less polar aqueous environment.
Turn up the heat
The “difference” in “differential” scanning fluorescence comes from adding heat. With DSF, you are measuring changes in fluorescence as a protein unfolds in response to the higher temperatures of a thermal ramp.
Identify inflection points
The onset temperature (Tonset) is when proteins begin to unfold. The other inflection points (Tm1, Tm2) are the temperatures at the mid-point of unfolding events. SLS identifies the aggregation temperature (Tagg) when one of these unfolding events leads to aggregation.
Make the most of full-spectrum fluorescence detection
Full-spectrum fluorescence is about more than just having the best signal-to-noise: it’s a key tool to figure out when something is wrong with your DSF experiment and a vital part of the back-up plan. Small proteins might not have fluorescent amino acids, and buffers can also fluoresce or quench the signal from intrinsic protein fluorescence — all things that full-spectrum detection helps deal with.
DSF can be done with fluorescent dyes that track protein unfolding, which is an option only available to systems that can detect it. Or for stability work with viral vectors, dyes sensitive to nucleic acids can be used with viral vectors like AAV to detect when genomes begin leaking out of capsids.
Uncle combines DSF, static light scattering, and dynamic light scattering measurements to deliver results for protein unfolding and aggregation problems, all done on the same sample. Add in customizable analysis across full-spectrum fluorescence detection and you’ve got a flexible tool that is ready to answer almost any protein stability question. Seeing the whole story on when proteins unfold and aggregate reveals exactly what’s going on with protein samples.
Ready for more?
Biopharma scientists can now use the right tool for protein stability studies with an instrument that combines differential scanning fluorimetry (DSF), static light scattering (SLS), and dynamic light scattering (DLS). Have a question or ready to find out more?