Abstract: The present invention concerns a novel in vitro assay for determining the invasive ability of brain tumour cells, in particular brain tumour stem cells. This new assay, called 3D invasion assay, is based on the analysis and comparison of the area of neurospheres grown in vitro into a Matrigel extracellular-like matrix. The invention also concerns the use of said assay for diagnosing metastatic brain tumour in subjects, for determining its prognosis, as well as for the monitoring, for the stratification, for identifying subjects at risk of metastasis, and/or predicting the metastasis-associated risk in subjects diagnosed with brain tumour.