The aim of this study was to compare the effect of DL-selenomethionine (SM) with 2 routinely used Se sources, sodium selenite (SS) and seleno-yeast (SY), on relative bioavailability based on antioxidant activity and tissue Se content. Six hundred thirty 131-day-old brown laying hens were randomly assigned to 7 treatments for 168 d (24 wks) with 6 replicates of 15 hens per replicate. The SS and SY animals were supplemented a cornmeal and soybean diet that supplied a total Se 0.3 mg/kg whereas SM was added at 4 different levels to the total Se at 0.1, 0.3, 0.5 and 0.7 mg/kg. All hens fed the Se-supplemented diet showed higher glutathione peroxidase (GSH–Px) activity (P < 0.01), higher superoxide dismutase (SOD) activity (P < 0.05), lower malondialdehyde (MDA) content (P < 0.05) in plasma, and greater Se contents in egg yolks, albumen, leg muscle, breast muscle, liver, and plasma compared with those fed the control diet (P < 0.01). The organic sources (SY and SM) exhibited a greater ability to increase the GSH–Px activity (P < 0.01) and Se content in albumen (P < 0.01), leg, and breast muscles (P = 0.0099 and P = 0.0014, respectively) than the SS that was added at 0.3 mg Se/kg. The higher SM added levels increased the GSH–Px activity until the dose of 0.5mg Se/kg (P < 0.01).The greater Se concentrations in albumen, muscle and liver appeared in the higher SM-added level, as well as above the dose of 0.1 mg Se/kg (P < 0.01). In addition, hens fed the diet with SM accumulated more Se in albumen, leg, and breast muscle than those fed diets with SY (P < 0.05). These results confirmed the higher ability of organic Se sources to increase the antioxidant activity and Se deposition in egg albumen, leg, and breast muscles compared with SS, and demonstrated a significantly better efficiency of SM compared with SY for albumen and muscle Se enrichment.