TABLE I.

Overview of the advantages and disadvantages of the different thermal unfolding methods applied in drug discovery.

Technique	Advantages	Disadvantages
DSC	No dyes, labels, Abs required. Direct measurement of heat capacity and enthalpy of unfolding	High protein consumption, low throughput.
DSF	High throughput, lower sample consumption. Can be automated	Requires dye binding. Potential issues with dye binding perturbing the equilibrium. Temperature dependence of fluorescence signal. May detect aggregation of dye.
nanoDSF	No dye required. Ability to take the ratio of 2 wavelengths to remove temperature dependence of fluorescence effect	Requires tryptophan / tyrosine content. More sensitive to compound fluorescence.
CETSA	Measurement in a physiological setting. A range of detection methodologies available	Complexity of interpretation.