ZCa is an engineered three-helical affinity protein that through replacement of the loop between helix two and three with an EF-hand motif has a calcium-dependent binding to its target. By using the difference of calcium concentration in the plasma and the cell along with the favorable properties of ZCa, the interest in creating therapeutics toward different diseases based on the ZCa scaffold has come about. In this project novel variants from a phage display library based on the ZCa scaffold are characterized and their ability to bind to their intended target is evaluated. The targets used in this project are IgE for its involvement in allergic reactions and IL-23 for its ability to cause autoimmune diseases. Through several different analysis methods this project shows that novel variants towards different targets are capable of calcium-dependent target binding.