A total number of 30 vaccine vials including 3 batches of 3 different types of live bacterial poultry vaccines (Mycoplasma red vaccine, Mycoplasma yellow vaccine and non colored Mycoplasma vaccine) were experimentally contaminated with 1,5,10 CFU of Escherichia coli reference strain (ATCC25992) /dose then subjected for sterility testing as referenced in world organization of animal health(OIE). The sterility is defined as the absence of viable microorganisms, includes viruses. It should be achieved by the use of aseptic techniques and validated sterilization methods, including heating, filtration, chemical treatments and irradiation that fit the intended purpose. Further dilution was carried out of vaccines showing negative results.Kappa index in different contamination levels in all tested vaccines after dilution was ranged between high Kappa index and almost total . In this study , it could be concluded that the applied testing method is appropriate for testing sterility of living bacterial poultry vaccines and it is fit to use