Bovine mastitis continues to be the most detrimental factor for profitable dairying. Recent research conducted within our laboratory has identified a genetic marker in the CXCR2 gene associated with mastitis susceptibility. The objective of the present study was to evaluate the migratory ability of neutrophils from cows with different CXCR2 +777 genotypes. Neutrophils isolated from peripheral blood of 30 Holstein cows were tested for in vitro migration and adhesion molecule expression. Cows with the CC or GC genotype at CXCR2 +777 showed significantly lower neutrophil migration to recombinant human interleukin-8 (rhIL-8) than cows with the GG genotype (P < 0.05). Cows with the CC genotype at CXCR2 +777 also showed decreased neutrophil migration to zymosan-activated serum compared to these same cows (P < 0.05). Decreased upregulation of CD18 expression was observed after stimulation with rhIL-8 in cows expressing the CXCR2 +777 CC genotype compared to cows expressing the GG genotype (P < 0.05). A similar trend was observed for CD11b (P < 0.10). However, no difference in CD62 downregulation was observed with respect to genotype. These results provide initial evidence for a phenotypic association between a single nucleotide polymorphism and neutrophil function in dairy cows, as well as potential insight into specific mechanisms affected in cows more susceptible to mastitis.