Methicillin-resistant Staphylococcus aureus (MRSA) is one of the leading causes of hospital-acquired infections worldwide, which is resistant to many antibiotics, resulting in significant mortality in societies. Vaccination is a well-known approach to preventing disease. Autolysin, a surface-associated protein in S. aureus with multiple functions, is a suitable candidate for vaccine development. As a co-adjuvant, selenium nanoparticles (SeNPs) can increase the immune system, presumably resulting in increased vaccine efficacy. The present study evaluated the immunogenicity and defense of recombinant autolysin formulated in SeNPs and Alum adjuvants against MRSA. r-Autolysin was expressed and purified by the Ni-NTA affinity chromatography. SeNPs were synthetically obtained from sodium dioxide, followed by an assessment of shape and size using SEM and DLS. Balb/c mice were injected subcutaneously with 20mg of r-autolysin formulated in Alum and SeNps adjuvants three times with the proper control group in 2 weeks intervals. Cytokine profile and isotyping ELISA were conducted to determine the type of induced immunity. Opsonophagocytosis tests assessed the functional activity of the vaccine, and the bacterial burden from the infected tissues was determined. Results showed that mice receiving SeNps and r-Autolysin had higher levels of total IgG and isotypes (IgG1 and IgG2a) and increased cytokine levels (IFN-γ, TNF-α, IL-12, and IL-4) as compared with those only receiving autolysin and PBS as a control. More importantly, mice immunized with SeNps and r-Autolysin exhibited a decrease in mortality and bacterial burden compared to the control group. We concluded that SeNps could stimulate immune responses and can be used as an adjuvant element in vaccine formulation.