Small affinity proteins based on alternative scaffolds make an attractive alternative to antibodies in anti-cancer therapy, for example for the delivery of cytotoxic payloads. They combine several advantages such as low production costs and increased tissue penetration while the possibilities to engineer the proteins are many. A major limitation of efficient delivery of cytotoxins to tumor cells is the internalization of the drug. A possible strategy to overcome this limitation is to develop binders that deliver their payload more efficiently into the targeted cells. Engineering the binder to be calcium- and pH-dependent would potentially increase intracellular delivery by dissociation in the lysosomes where the calcium concentration and pH is lower than on the cell surface and would thus allow the targeted receptor to return to the cell surface while the targeting molecule, together with the payload, is released inside the cell. We here present an engineering approach to generate calcium and pH-dependent binders based on the Z domain of Protein A. Previously, a calcium-binding loop has been successfully introduced into the domain enabling the mild purification of antibodies. A new library for calcium-regulated affinity (CaRA) has been developed based on this domain with randomized surface-exposed residues for novel target binding. The CaRA binders were selected using phage display in a newly established high-throughput workflow which has shown to be highly versatile with the potential to be integrated into any lab for the discovery of new binders. We envision that this engineering approach combined with the efficient selection procedure enables the discovery of new, promising domains for more potent anti-cancer therapy.