Hi, Could someone refer me to some papers on using ANS to estimate amount of hydrophobic surface in a protein? I tried the classic way by STEP 1: titrating my protein into 1uM ANS till saturation to get the molar fluorescence activity of ANS, then STEP 2: titrate ANS into protein to get amount of fluorescene at plateau so that I can estimate how much ANS is bound per protein molecule. However, for this protein I am dealing with, I can never get the STEP 1 done because the fluorescence signal never saturates. It keeps going up. Are there other ways of getting around this? Thanks, Huiming Li
