Background/objectives: The emergence of highly pathogenic avian influenza A (H5N1) virus infections in dairy cattle in the United States revealed a novel mammalian host and a potential transmission pathway involving raw milk and dairy production systems. Sustained circulation of H5N1 in dairy herds is of concern because ongoing viral adaptation in mammals may increase the risk of efficient mammalian transmission and subsequent zoonotic spread. In response to this emerging threat, we developed a chimpanzee adenovirus (ChAd)-vectored vaccine expressing the haemagglutinin 5 antigen (H5HA) from the dairy cattle isolate A/dairy cattle/Texas. Methods: Lactating dairy cows were vaccinated by either intramuscular (N = 3) or intranasal administration (N = 3). H5HA clade 2.3.4.4b antibodies in cow’s milk and sera were evaluated by ELISA. Hemagglutination and neutralisation capacity were also evaluated. Results: IgG and IgA H5HA-specific antibodies were detected in serum and milk from parenterally vaccinated animals, demonstrating the induction of a systemic immune response. The neutralising antibody responses elicited were only detected in serum of cows vaccinated via the intramuscular route. Conclusions: These preliminary findings support the feasibility of ChAd-vectored vaccination as a strategy to induce humoral immunity in cattle against emerging H5N1 influenza A viruses. Cross-reactive antibody responses against both A/dairy cattle/Texas/24-008749_001/2024 and A/Ibis/Egypt/RLQP-229S/2022 support the capacity of the vaccine to recognise antigenically related H5N1 clade 2.3.4.4b viruses circulating in mammalian and avian reservoirs. Further studies of vaccine efficacy and the immunological mechanism of protection should now be undertaken with the aim of reducing viral transmission and milk-associated shedding in dairy herds.