Our objective of the study was to effect of probiotics on the immune response of Hanwoo heifers. Lipopolysaccharide (LPS) challenge was used for investigating physiological response of dietary probiotics. A completely random design was used (4 pens; 2pens/treatment; 5 heifers/pen). After the cattle fed probiotic for 5 months, 16 heifers were transported and acclimated to environmentally controlled chambers. Heifers were fitted with indwelling jugular catheters prior to 24 hours of the LPS challenge. Blood samples were collected at 30-min intervals from -1 to 6 h (0 h; 1μg/kg BW of LPS from Escherichia coli O111:B4). Glucose, non-esterified fatty acid (NEFA), albumin, triglyceride, total protein, phosphorus concentrations, plasma CBC (WBC, RBC, Platelet, Neutrophils, Lymphocytes, Eosinophils, Basophil, Hemoglobin), and pro-inflammatory cytokines (TNFα, IL6, IL1b) were determined from blood samples. Response to the LPS challenge over time was analyzed by ANOVA with the MIXED procedure of SAS. Overall ADG and serum compositions did not differ between probiotic or control diet for 5 months (P >0.05). Pre-LPS NEFA concentration did not differ (P >0.05), but probiotic treated heifers was decreased at 2 hours after LPS challenge. NEFA concentration was decreased at 2 hours after LPS challenge in probiotic treated group (P <0.05). Serum triglyceride was peaked at 0.5 h after LPS challenge in of probiotic treated heifers (P <0.05). There was no difference at CBC test between treatment pre- and post - LPS challenge except red blood cell (RBC). Plasma RBC concentration was increased from 0.5h to 3h post-LPS challenge in probiotic treated heifers. These data suggest that probiotic diet did not directly altered immune response to Hanwoo heifers but indirectly regulated lipid metabolism of Hanwoo heifers at the LPS challenge.