Abstract Background Diffuse Intrinsic Pontine Glioma (DIPG) is a childhood tumor with dismal prognosis. Emerging molecular signatures have paved the way for stereotactic biopsy in selected Centers. We present our experience in DIPG stereotactic needle biopsy using the Robotic Stereotactic Assisted system (ROSA™) in a consecutive pediatric series. Methods All stereotactic biopsy procedures for DIPG performed during the last year at our Institution were considered. All procedures were carried out using the ROSA™ surgical assistant through a precoronary approach. All children underwent a post-operative CT scan to document possible surgical complications and confirm the site of biopsy. Post-operative clinical changes were recorded to test morbidity of the procedure. Results In the last year we performed 7 pontine needle biopsies. Specimens were diagnostic and useful for molecular analysis in all cases. No surgical complications were observed. One child showed a transient neurological worsening related to the biopsy that resolved within 2 weeks. The combination of the precoronary approach and use of the stereotactic ROSA™ system allowed single-session surgeries in all cases. Conclusions Pontine biopsy for DIPG is a safe procedure in selected Centers. The advantages of the single-session procedure we described might be of particular interest in the pediatric setting.