The neuropeptide substance P is a possible candidate as histamine-releasing factor in some patients with chronic ordinary urticaria (COU), particularly in those with evidence of a nonautoantibody circulating histamine-releasing factor. In this study, serum substance P levels were measured by enzyme immunoassay in 117 COU patients, 40 atopic subjects and 24 normal subjects. In vivo and in vitro assays for histamine-releasing factors, autologous serum skin test (ASST) and basophil histamine release (BHR) assay, respectively, were performed in all COU patients. Mean serum substance P concentration was not significantly different in COU patients and in normal subjects; however, significantly higher levels were detected in atopic subjects than in COU patients (P < 0.003). ASST and BHR assays allowed us to distinguish different subsets of COU patients. Mean serum substance P concentration did not vary significantly in the different subsets of patients. Nevertheless, interestingly three patients with positive ASST and negative BHR assay showed very high substance P levels. These results suggest that substance P does not play an important role as histamine-releasing factor in COU in general but only in occasional patients in whom it might act as a trigger of urticarial symptoms.