Surface hydrophobicity is a significant indicator for evaluating the surface properties of plant proteins. It plays a crucial role in determining the functionality of proteins, such as emulsification and foam stabilization abilities. This chapter provides a comprehensive analysis guideline for measuring surface hydrophobicity using two different methods: spectrofluorimetric assessment with the fluorescence probe 1-anilinonaphthalene-8-sulfonic acid (ANS1-Anilinonaphthalene-8-sulfonic acid (ANS)), and hydrophobic-interaction capacity measurement with bromophenol blue (BPB). Therefore, the surface hydrophobicity of plant proteins can be expressed by a relative index value (S0) and the bound BMP value, respectively.