Oxytocin is a peptide hormone with high affinity to both Zn²⁺ and Cu²⁺ ions compared to other metal ions. This affinity makes oxytocin an attractive recognition layer for monitoring the levels of these essential ions in biofluids. Native oxytocin cannot differentiate between Cu²⁺ and Zn²⁺ ions and hence it is not useful for sensing Zn²⁺ in the presence of Cu²⁺. We elucidated the effect of the terminal amine group of oxytocin on the affinity toward Cu²⁺ using theoretical calculations. We designed a new Zn²⁺ selective oxytocin-based biosensor that utilizes the terminal amine for surface anchoring, also preventing the response to Cu²⁺. The biosensor shows exceptional selectivity and very high sensitivity to Zn²⁺ in impedimetric biosensing.