This study evaluated the relationships between immune response, disease resistance, and bovine leukocyte antigens, BoLA, in Holstein cows following intramammary challenge with Staphylococcus aureus. This investigation was to determine whether immune responsiveness differed between these cows and whether differences were related to expression of class I BoLA antigens, which might explain the increased resistance or susceptibility to S. aureus mastitis. Antibody responses to S. aureus in milk and serum, total IgG1 in milk, and blastogenic responses with and without concanavalin A were evaluated. The CA42 allele, previously associated with increased risk of infection, was relatively uninformative for the immune response parameters examined. Other alleles, such as W3, were associated with higher milk antibody responses to S. aureus and higher milk IgG1 postchallenge. Alleles W7, W4, and W26 were associated with lower milk IgG1 and lower antibody titers in serum postchallenge. The association reported between allele CA42 and increased risk of S. aureus mastitis did not relate to lower antibody or blastogenic responses by these cows; however, indicators of innate resistance were not examined. In addition, the different associations detected between milk and serum emphasize the importance of considering mucosal immunity and unique immunological compartments when searching for relevant genetic markers of disease resistance.