In the present study, the isolation of selenium (Se)-enriched bacteria from rumen fluid and hot spring water was carried out. Rumen fluid samples were taken from cannulated goats fed a basal diet and the water samples were collected from Selayang hot spring, Selangor- Malaysia. A total number of 140 Se-toler-ant isolates were obtained aerobically using an Se-enriched medium and spread plate technique. All the isolates were initially screened for the ability to transform the Se-containing medium to a red-orange cul-ture using a spectrophotometer. Twenty isolates of dark red-orange medium were selected for a screening of the highest Se-containing protein accumulating strains using the dialysis technique and icp.ms to mea-sure the Se content. Four isolates, identified as Enterobacter cloacae (ADS1, ADS7, and ADS11), and Klebsiella pneumoniae (ADS2) from rumen fluid origin, as well as, one isolate from hot spring water (Stenotro-phomonas maltophilia (ADS18)), were associated with the highest biomass organic Se-containing protein when grown in a medium enriched with 10 μg/ml sodium selenite. In addition, around 50 μg/100 μg of the absorbed inorganic Se was accumulated as an organic form. Organic Se-containing protein in all the selected strains showed antioxidant properties in the range of 0.306 to 0.353 Trolox equivalent antioxidant capacity (TEAC) mg/ml. Therefore, these strains may offer a potential source of organic Se due to their Se-tolerant nature and higher biomass organic to inorganic Se ratio.