Polyunsaturated FA have modulatory effects in a wide range of biological functions. The objectives for this study were (1) to determine if docosahexaenoic (DHA) or arachidonic acid (ARA) affect pre-weaning glucose and lipid metabolism, and (2) to determine if altered supply of these n-3 and n-6 polyunsaturated fatty acids (PUFA) in milk replacer modulates the immune response to vaccination. Thirty newborn Holstein heifers were blocked by date of birth and, within block, randomly assigned to 5 treatments: control (choice white grease), low dose ARA (1.6 g/d), high dose ARA (4.8 g/d), low dose DHA (1.6 g/d) and high dose DHA (4.8 g/d). Treatments provided approximately 1% added lipid (DM basis), with choice white grease added to ARA and DHA treatments. Treatments were blended into a milk replacer that was 28% CP and 20% fat (DM basis) and 1.89 L was fed 3 times/d until weaning at 63 d. Multivalent vaccines were administered on approximately d 49 and 63. Data were analyzed with mixed models including the fixed effects of treatment, time, and their interaction, and the random effect of block and heifer within block. DMI, ADG and feed efficiency were not affected by treatments, but ARA decreased hip height growth and increased the incidence of abnormal fecal consistency (P < 0.05). Glucose and insulin concentrations were greatest for control and least for the low PUFA doses (P < 0.05). Vaccination response was assessed by plasma TNF-α and haptoglobin concentrations 24-h post-vaccination, BRSV and BVD1/2 antibody titers at 2 wk, and proliferation of CD4⁺ T helper cells and CD8⁺ cytotoxic T cells at 6 to 10 d; none of these responses different by treatment (P > 0.10). The supplementation of either PUFA modulated glucose metabolism, ARA had subtle negative impacts on digestive health and skeletal growth, but neither PUFA source significantly altered the immune response to vaccination in pre-weaning calves.