The amino (-NH2) and sulfhydryl (-SH) groups represent two critical functional moieties frequently analyzed to elucidate protein structural alterations. Determining the abundance of free amino groups is imperative for quantifying the extent of hydrolysis, a pivotal process in protein modification. Precise control over the degree of hydrolysis is essential to achieve the desired protein properties. Sulfhydryl groups also play vital structural roles in proteins, particularly when protein characteristics change due to the rearrangement or formation of disulfide bonds resulting from various treatments. Owing to their functional significance in protein structure and their utility in protein modification processes, both amino and thiol groups are acknowledged as indispensable functional entities in numerous food proteins. In this chapter, two methods for quantifying these functional groups are outlined: the use of 2,4-dinitrobenzene-1-sulfonic acid (DNBS) for quantifying free amino groups and the use of 5,5′-dithiobis-(2-nitrobenzoic acid) (DTNB5,5′-Dithiobis-(2-nitrobenzoic acid) (DTNB)) for measuring free sulfhydryl groups. These methods are specifically designed to enhance the analysis of protein samples, contributing to a deeper understanding of their characteristics.