This study evaluated the interaction of RUP and fat levels on growth, rumen fermentation, and blood metabolites of Holstein calves. Forty 3-d-old calves (20 females and 20 males) with a starting BW of 40.6 ± 2.8 kg were used in a completely randomized design with a 2 × 2 factorial arrangement of treatments. Within sex treatments were: (1) high RUP and low fat (HRUP-LF); (2) low RUP and high fat (LRUP-HF); (3) high RUP and low fat (HRUP-LF); and high RUP and high fat (HRUP-HF). Low-RUP starter contained 21.5%, whereas high RUP starter contained 34.3% RUP as % of CP, whereas low fat starter contained 2.9% and high starter contained 5.8% crude fat based on DM. Isonitrogenous levels in the starter grain were maintained by replacing solvent soybean meal with heat treated soybean meal while fat levels were increased by the addition of prilled fatty acids. Calves were housed individually and had ad libitum access to water and calf starter throughout the study. All calves were weaned on d 60 of age but remained in the study until d 70 for final measurements. Overall, there was no interaction between RUP and fat levels for measured variables. Starter intake tended ( = 0.09) to be greater for calves fed low fat starter during the postweaning period, although over the whole experiment and during the preweaning period, differences in starter intake were not different. Although there were no differences for most VFA concentrations, the molar proportion of butyrate tended ( < 0.08) to be greater in the rumen of calves fed low-fat starter compared to those fed high-fat starter. Serum total protein was lower ( < 0.05) and serum cholesterol was greater ( < 0.01) for calves fed high-fat starter by d 65 of life. The concentration of alanine aminotransferase was also lower ( < 0.05) for calves fed high-fat starter compared to those fed low-fat starter on d 65, and these levels tended to increase with the addition of RUP ( < 0.07). In conclusion, no effects were attributable to feeding a high-RUP starter. However, feeding a calf starter with over 3% fat appeared to decrease starter intake as growth progressed.