It has been hypothesized that bacterial debris may accumulate in tissues of the reticuloendothelial system (RES) serving as an inflammatory stimulus for human disease. In support of this hypothesis, muramic acid (Mur), a component of bacterial peptidoglycan (PG), has previously been reported to be present in culture-negative human spleen. High-performance liquid chromatography (HPLC) was employed in these analyses, and a peak was detected at the retention time of Mur. However, HPLC is best used as a screening technique, and it is vital that these tentative observations be reexamined by the state-of-the-art approach (gas chromatography-tandem mass spectrometry [GC-MS(2)]). Indeed, in the present work using GC-MS(2), Mur was not detected in six out of seven human spleens previously examined by HPLC. However, Mur was categorically detected at minute concentrations, 50 ppb, in one spleen. In conclusion, since Mur is not generally found in culture-negative human spleen, in future studies, these tissues can serve as negative controls. The study of Mur levels in inflammation (e.g., reactive arthritis) could prove important in testing the hypothesis that bacterial debris persisting in tissues could serve as a depot inciting diseases of unknown etiology.