I am using fluorescence spectroscopy to determine stability parameters for human acidic fibroblast growth factor 1. (FGF1). The fraction unfolded for FGF1 is determined from the ratio of emission at 350 nm to 308 nm corresponding to tryptophan and tyrosine emissions respectively. So, what I need is the fraction unfolded, do I still need to subtract blank from the 350 nm and 308 nm fluorescence reading separately at each denaturant concentration?