Engineered affinity proteins have, together with antibodies and antibody derivatives, become indispensable tools in many areas of life science and with an increasing number of applications. The need for high-throughput methods for generation of these different affinity proteins is evident. Today, combinatorial protein engineering is the most successful strategy to generate novel affinity proteins of non-immunoglobulin origin. In this approach, high-complexity combinatorial libraries are constructed from which affinity proteins are isolated using appropriate selection methods, thus circumventing the need for detailed knowledge of the protein structure and the binding mechanism that is necessary in more rational approaches. Since the introduction of the phage display technology, several alternative selection systems have been developed for this purpose. This review presents briefly some of the more commonly used affinity proteins, and gives an overview of the different methods and challenges related to the generation of library diversity and the selection methods available for the isolation of affinity proteins with desired properties. (C) 2012 Society of Chemical Industry