The aim of the study was to investigate the antioxidant effect of seleno-amino-oligosaccharide (Se-AOS) on intestinal porcine epithelial cells (IPEC-1). MTT assay showed that Se-AOS had no effect on the viability of IPEC-1 cells up to a concentration of 9200 μg/L and Se-AOS significantly increased the viability of IPEC-1 cells compared to cells exposed to H2O2 alone. Se-AOS significantly increased the level of superoxide Dismutase (SOD) and decreased the levels of malonic dialdehyde (MDA) and lactate dehydrogenase (LDH) in IPEC-1 cells. The gene expression levels of different antioxidant enzymes dramatically increased by the pretreatment of Se-AOS compared to H2O2 treatment. In addition, the results indicated that Se-AOS up-regulated the intracellular Nrf2 and down-regulated the level of Keap1 by western blot. Taken together, these findings suggested that Se-AOS can protect IPEC-1 cells from oxidative damage through activating the Keap1/Nrf2 signaling pathway.