This study aimed to study the inhibition activity of purified bacteriocin produced from the local isolation Lactococcuslactis ssp. lactis against pathogenic bacteria species isolated from clinical samples in some hospitals Baghdad city. Screening of L. lactis ssp. Lactis and isolated from the intestines fish and raw milk was performed in well diffusion method. The results showed that L. lactis ssp. lactis (Lc4) was the most efficient isolate in producing the bacteriocin as well observed inhibitory activity the increased that companied with the concentration, the concentration of the twice filtrate was better in obtaining higher inhibition diameters compared to the one-fold concentration. The concentrated bacteriocin was purified using the gel filtration column and Sephacryl S-200. The results showed the high inhibitory activity of the purified bacteriocin after the purification against the positive and negative bacteria of the Gram stain under study compared to the one-fold concentration and two-fold before purification , The diameters of the inhibition zones after gel-filtering of the purified bacteriocin reached S. aureus, S. epidermidis, P. aeruginosa, E. coli, E. clocae and S. marcescens (23, 25, 26, 20, 22 and 28) Mm respectively. The carbohydrate content of purified bacteriocin from L. lactis ssp. lactis (Lc4) isolate was 6.02% with a molecular weight of 6310 Dalton. The results showed that purified bacteriocin retained its inhibitory activity at pH 2-10 and showed the highest inhibition at pH 4-6 and lost at pH 12. The purified bacteriocin was characterized by thermal stability. It retained its effectiveness when exposed to 40, 60, 80, 100°C for 30, 15, 5 minutes and 120°C for 15.5 minutes and lost 50% of its effectiveness when exposed to 120°C for 30 minutes. Results The purified bacteriocin was effectively retained when treated with enzyme pepsin and trypsin of 37°C for one hour and at pH 7.