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In this study we developed a microfluidic chip for the rapid capture, enrichment and detection of airborne Staphylococcus (S.) aureus. The whole analysis took about 4 h and 40 min from airborne sample collection to loop-mediated isothermal amplification (LAMP), with a detection limit down to about 27 cells. The process did not require DNA purification. The chip was validated using standard bacteria bioaerosol and was directly used for clinical airborne pathogen sampling in hospital settings. This is the first report on the capture and analysis of airborne S. aureus using a novel microfluidic technique, a process that could have a very promising platform for hospital airborne infection prevention (HAIP).