Proteins are reckoned to be the key actors in a living organism. By studying proteins, one engages into deciphering a complex series of events occurring during a protein's life span. This starts at the creation of a protein, which is tightly controlled on both a transcriptional (Williams and Tyler, 2007, Curr Opin Genet Dev 17, 88-93) and a translational level (Van Der Kelen et al., 2009, Crit Rev Biochem Mol Biol 44, 143-168). During translation, a primary strand of amino acids undergoes a complex folding process in order to obtain a native three-dimensional protein structure (Gross et al., 2003, Cell 115, 739-750). Proteins take on a plethora of functions, such as complex formation, receptor activity, and signal transduction, which ultimately adds up to a cellular phenotype. Consequently, protein analysis is of major interest in molecular biology and involves annotating their presence and localization, as well as their modification state and biochemical context. To accomplish this, many methods have been developed over the last decades, and their general principles and important recent advances in large-scale protein analysis or proteomics are discussed in this review.