Staphylococcus aureus, including methicillin-resistant S. aureus (MRSA), is a major bacterial pathogen associated with nosocomial and community-acquired S. aureus infections all over the world. A rapid detection assay for staphylococcal gene of nuc and mecA is needed. In this study, a rapid identification assay based on the loop-mediated isothermal amplification (LAMP) method was established. PCR and LAMP assays were used to detect Staphylococcus aureus and other related species for nuc and mecA. With optimization of the primers and reaction temperature, the LAMP successfully amplified the genes under isothermal conditions at 62 °C within 60 min, of which the results were identical with those of the conventional PCR methods. The detection limits of the LAMP for nuc and mecA were 1.47 and 14.7 pg/μl DNA per tube, respectively, by naked eye inspections, while the detection limits of the PCR for nuc and mecA were 14.7 pg/μl and 147 pg/μl DNA, respectively. Finally, The LAMP method was then applied to clinical blood plaque samples. The LAMP and PCR demonstrated identical results for the plaque samples with the culture assay. Together, the LAMP offers an alternative detection assay for nuc and mecA with a great advantage of the sensitivity and rapidity.