Individuals often respond differently to the same vaccine; some of this variation may be caused by genetic differences among animals. Our objective was to estimate heritability and identify genomic regions associated with humoral response to an Escherichia coli O157:H7 vaccine in beef cattle. Crossbred beef cattle (n = 651) were vaccinated with a commercially available E. coli O157:H7 vaccine. Serum was collected at time of initial vaccination (d 0), booster (d 21), and d 56 after initial vaccination. Total antibodies specific to siderophore receptor and porin proteins in the vaccine were quantified by enzyme-linked immunosorbent assay. Genomic DNA was isolated from whole blood and genotyped with the bovine GeneSeek Genomic Profiler-High Density 78K or 26K Single Nucleotide Polymorphism BeadChip and imputed to 777,000 SNP genotypes. Heritability was estimated by restricted maximum likelihood (REML) using both 1) pedigree and 2) genomic relationships among individuals. Fixed effects were contemporary group, calf age, sex, principal components from SNP genotype data, and pedigree-derived heterozygosity effects. Additive and dominance effects of SNPs were estimated individually while accounting for contemporary group, sex, and the top 20 principal components calculated from the genomic relationship matrix. Heritability of initial response to vaccination (d 21 -d 0) was 0.10 ± 0.175 using pedigree relationships and 0.14 ± 0.149 using genomic relationships, but neither estimate was statistically different from zero. Heritability of booster (d 56 -d 21) and overall (d 56 -d 0) responses were low and not statistically significant from zero. There were no clusters of linked SNP associated with vaccine response, but eight regionally isolated SNPs were significantly associated with initial or overall response to vaccination. Regional genetic variation for initial response to an E. coli O157:H7 vaccine was observed, although overall heritability of this response was not statistically significant from zero.