This chapter focuses on the subtractive modification of the Edman degradation. For successful coupling and subsequent degradation, certain precautions should be observed that (1) the peptide should be free of salts of acids and bases since they will interfere with pH control in the coupling step and (2) the solvents and reagents should be free of even traces of aldehydes because they will form Schiff bases with amino groups and block the reaction with phenylisothiocyanate. Under ideal conditions, successive application of the Edman degradation should cause an integral decrease of a single amino acid residue at each step. Side reactions occur leading to the accumulation of products that will not react with phenyl isothiocyanate or undergo cyclization. As a result, the remaining peptides will show a non-integral decrease of the amino acids which are being removed. The main advantage of the subtractive method is that it provides quantitative information about the yield at each stage of the degradation. This can be very important when difficulties in the degradation arise which prevent the continuation of the Edman procedure.