The antimicrobial activity of Cefoperazone-Sulbactam was studied by microbiological assays in order to determine their potency by diffusion gel method. Bacillus subtilis ATCC 6633 was established as a microbiology model, and comparison study of antibiotic samples using USP standard, generic and originals was executed. Procedures were established in the laboratory, capable of handling experiential test for assaying various execution biomass conditions such as bacteria growing, incubation times and inhibition zones as a response to various concentrations of antibiotic. Also, the bio-assay`s methodology was validated. This study revealed all antibiotic samples behave in similar ways, through in vitro methods. We therefore conclude that all of the samples are pharmaceutical equivalents and the products can be used in the antimicrobial therapy.