Differential Scanning Fluorimetry measures protein thermal unfolding by monitoring changes in fluorescence emission of a sample upon heating. This allows the determination of protein thermostability and complex formation even with weakly binding ligands by thermal shift assay. Differential Scanning Fluorimetry is therefore ideally suited for screening of optimum buffer conditions like pH, buffer composition and ionic strength. The technique is applicable to any biological sample, from soluble proteins to integral membrane proteins.
What are the sample requirements?
- Sample limits: From 1 kDa to 1 MDa
- Sample volume per experiment: 10 μl/capillary or 10 ul diluted to 25 ul per well
- Protein with intrinsic fluorescence (tryptophan and tyrosine) or use of dye
- Sample concentration depends on equipment:
- 0.1 mg/ml to 150 mg/ml for capillaries
- Typically 1 mg/ml for multi-well plates
What other specific considerations are relevant?
- Typical temperature ranges are 10oC to 95oC
- Close-to-native analysis possible with no labelling.
- Protein samples need to be pure as impurities also will contribute to the measurement.
Partners offering this technique
MOSBRI reference partner site for this technique: