OBJECTIVE To investigate the accuracy of laboratory results of urine samples collected from cotton balls as compared with paired samples collected from catheterization, Preemie Pampers, and Newborn Pampers. METHODS Urine samples were collected from 30 infants by aspirating urine from a cotton ball placed over the meatus. An additional urine sample was obtained from the same infants by aspiration from the diaper. The samples were tested by labstik analysis and results were compared by t-test. An additional sample of 20 infants had urine collected by cotton ball and catheterization for detection of Group B strep antigen and the results were correlated. RESULTS Samples obtained from cotton balls and Preemie Pampers yielded equivalent results. Significant differences were found in the pH and specific gravity of Newborn Pampers and Preemie Pampers. No redness nor skin breakdown was observed with the cotton ball application. Three positive Group B strep results were detected by both methods of urine collection. The remainder of the samples were negative by both methods. CONCLUSIONS Urine samples collected by the cotton ball method were accurate for pH and specific gravity and were atraumatic to the skin of newborns. The sample size of positive Group B strep antigen was small (n = 3) and results should be replicated with further research.