Highly pathogenic avian influenza (HPAI) clade 2.3.4.4b H5Nx viruses continue to cause episodic incursions and have been detected in more than 12 taxonomic orders encompassing more than 80 avian species, terrestrial and marine mammals, including lactating dairy cows. HPAI H5N1 spillover to dairy cattle creates a new interface for human exposure and raises food safety concerns. The presence of H5N1 genetic material in one out of five retail pasteurized milk samples in the USA has prompted the evaluation of the pasteurization processes for the inactivation of influenza viruses. Our study examined whether pasteurization could effectively inactivate HPAI H5N1 spiked into raw whole milk. First, we heated 1 mL of non-homogenized cow milk samples to attain an internal temperature of 63°C or 72°C and spiked with 6.3 log10 EID50 of clade 2.3.4.4b H5N1 virus. Complete inactivation was achieved after incubation of the H5N1 spiked raw milk at 63°C for 30 min. In addition, viral inactivation was observed in seven of eight experimental replicates when treated at 72°C for 15s. In one of the replicates, a 4.44 log10 virus reduction was achieved, which is about 1 log higher than the average viral quantities detected in bulk milk in affected areas. Therefore, we conclude that pasteurization of milk is an effective strategy for mitigation of the risk of human exposure to milk contaminated with H5N1 virus.