The purpose of the study was to evaluate the incidence of postdural puncture headache in a predominantly pediatric sample before and after a transition from conventional to atraumatic spinal needles. In this retrospective cohort study, we analyzed data from 1059 lumbar puncture procedures in 181 individuals enrolled in NIH Clinical Center research protocols. Multivariate logistic regression was used to evaluate the association between postdural puncture headache and spinal needle type after adjusting for patient age, sex, and body mass index. A random effect of participant was used to accommodate repeated observations. The median age at time of procedure was 15.3 years. The overall rate of postdural puncture headache was 5.1% (54 of 1059). With conventional needles and atraumatic needles, respectively, the rate of postdural puncture headache was 7.7% (43 of 588) and 2.3% (11 of 471); (odds ratio 0.32, 95% confidence interval 0.15 to 0.68). Lumbar puncture for cerebrospinal fluid collection is an essential and common procedure in pediatric clinical care and research. Postdural puncture headache is the most common adverse event of the lumbar puncture procedure. Our data indicate that lumbar puncture is safe in pediatrics and that use of an atraumatic spinal needle further reduces the risk of postdural puncture headache.