Crude alpha-hemolysin was produced by Staphylococcus aureus and it was extracted from the same bacteria which was isolated from clinical samples collected from skin tissue infection and sepsis.the hemolytic activity of crude alpha-hemolysin against rabbit erythrocytes was determined. The culture supernatant was treated with ammonium sulfate (75% saturation)and also hemolytic activity after addition of ammonium sulfate was measured. Skin test was determined against rabbit to detect its ability to induce delayed type hypersensitivity sensitivity and it gave positive results in compare with control and lethal toxin activity was tested in mice to reveal its ability to induce infection symptoms and mortility after 48 hour and 18 hour. so this study proved that crude alpha-hemolysin possess properties enable it to behavior as antigen and to be effective virulence factor in S. aureus.