Objective To screen the aptamers of Staphylococcus aureus (S. aureus cells) and lay the foundation for establishing a specific, sensitive and rapid detection method for S. aureus cells.Methods Cell-SELEX was performed using SELEX (Systematic Evolution of Exponentially Enriched Ligands) technique for screening, targeting S. aureus cells. Results Four sequences showed significantly higher binding to S. aureus cells, distinguishing them from control bacteria of other important microbial species: Escherichia coli,Candida albicans, Streptococcus pyogenes, Streptococcus pneumoniae, S. typhimurium, Staphylococcus pseudintermedius and Inactivated S. aureus cells. Apt-SA-1(Kd=56.0±3.0 nM), Apt-SA-6(Kd =86.0±3.2 nM ), Apt-SA-25(Kd=89.3±2.9 nM) and Apt-SA-32(Kd=90.2 ± 5.6 nM), with Apt SA-1 having the highest affinity. The four aptamers showed high affinity and specificity for S. aureus cells. Conclusion The four aptamers obtained in this study were able to specifically recognize S. aureus cells and provide a theoretical basis for the development of radiopharmaceuticals capable of recognizing the source of S. aureus cells infection and other aptamer-based bacterial diagnostic methods