Within this chapter, various techniques and instructions for characterizing primary structure of proteins are presented, whereas the focus lies on obtaining as much complete sequence information of single proteins as possible. Especially, in the area of protein production, mass spectrometry-based detailed protein characterization plays an increasing important role for quality control. In comparison to typical proteomics applications, wherein it is mostly sufficient to identify proteins by few peptides, several complementary techniques have to be applied to maximize primary structure information and analysis steps have to be specifically adopted. Starting from sample preparation down to mass spectrometry analysis and finally to data analysis, some of the techniques typically applied are outlined here in a summarizing and introductory manner.