Methicillin-resistant Staphylococcus aureus (MRSA) has rapidly emerged worldwide, affecting both healthcare and community settings, and intensive livestock industry. The efficient control of MRSA strongly depends on its adequate laboratory detection. This guideline provides recommendations on the appropriate use of currently available diagnostic laboratory methods for the timely and accurate detection of MRSA in patients and healthcare workers. Herewith, it aims to standardise and improve the diagnostic laboratory procedures that are used for the detection of MRSA in Dutch medical microbiology laboratories.