In this study, about 312 clinical samples were collected from different health institutions in Babylon governorateIraq, during the period from 1/10/2021 to 12/1/2022, by taking swabs from Burns (50 samples), wounds (55 samples), nose swabs (50 samples), abscesses (50 samples), diabetic foot (52 samples), and samples from the various inflammatory site such as urine (55 samples), various phenotypic, microscopic, and biochemical tests were conducted on it to detect about the presence of S. aureus among the isolates, these tests proved the return of (105 isolates out of 312 samples) isolated from different sites at clinical samples (Percentage of 33.65%) to the genus S. aureus, Zinc oxide nanoparticles were also chemically synthesized by the sol-gel method. The diagnosis of the nano-material was confirmed by conducting four specialized analyzes to reveal the properties of the synthesized material, by (XRD, AFM, FTIR, FESEM), the synthesized nanomaterial was also tested on Staphylococcus aureus bacteria to know its antibacterial activity and its effectiveness in inhibiting the growth of Staphylococcus aureus bacteria, used statistical program SPSS version 26 with biological part by calculating least significant differences with one way ANOVA.