The trial was conducted to see if an endoscopic transcortical intraventricular biopsy was effective and safe in children with solitary thickened pituitary stalk lesions. Between 1994 and 2018, 16 juvenile patients (8 males and 8 females) underwent endoscopic transcortical intraventricular biopsies at a single institution to establish an accurate pathological diagnosis and choose the best therapy options. Before surgery, the pituitary stalks had a median diameter of 5.31± 1.67 mm (range 3.86–9.17 mm). Overall, an endoscopic intraventricular biopsy had a diagnostic yield of 93.8%. Germinoma (n = 10, 62.5%), Langerhans cell histiocytosis (n = 4, 25%), pilocytic astrocytoma (n = 1, 6.3%), and pituicytoma (n = 1, 6.3%) were among the histopathological diagnoses. After the operation, two patients acquired new diabetes insipidus, and three patients developed new hypothyroidism. In this series, there were no postoperative neurological impairments. In managing isolated pituitary stalk lesions in pediatric patients, neuroendoscopic biopsy via the transcortical intraventricular corridor was proven safe and aided in a reliable histological diagnosis. For open biopsies, it can be considered a less invasive option to transcranial or endonasal pathways.