Purpose: Glioma are highly malignant brain tumors that exhibit strong angiogenesis (Ref. 1,2). Hence neoangiogenesis has become a main treatment target. This requires imaging methods to assess vascularization status, treatment effects and disease progression. However until now mapping tumor vascularization has been difficult. We have developed a combined magnetic resonance and optical vascularization “tool kit” (MR-UM) to study neoangiogenesis in mouse glioma models. We use in vivo magnetic resonance (MR) imaging and correlative ultramicroscopy (UM, Ref.3) of ex vivo cleared whole brains to track neovascularization at single vessel resolution. Methods: 105 Gl-261 glioma cells were implanted intracranially in black6 WT mice. Longitudinal in vivo magnetic resonance imaging (experimental 9.4 Tesla) was performed pre and post contrast using high resolution T2* sequences (80µm isotropic resolution). USPIO (CLIO-FITC) or conventional Gd contrast agents (0.2 mmol/kg gadovist©) were used. Dynamic contrast enhanced imaging was performed to evaluate permeability. Correlative ultramicroscopy using selective plane illumination microscopy (SPIM) of ex vivo cleared whole brains (3DISCO, Ref. 4) was performed to validate MR imaging. For optical detection of the microvasculature lectin-FITC or lectin-texas red (0.12 mg/kg) that bind to endothelial glyocoproteins were administered iv. VEGF inhibition (10mg/kg) was done to assess treatment responses by MR-UM. Results: T2* MR imaging allows the identification of single sprouting vessels in glioma development and the quantification of neovessels over time (weeks 2-4 post tumor implantation). First tubular neovessels can be delineated two weeks after tumor inoculation. After contrast administration single tubular hypointense neovessels run both in the center and in the periphery of the developing glioma. Within one week and following the angiogenic switch the entire tumor core is filled with hypointense neovessels (~3.9 fold increase of neovessels from week 2 to 3). Using a variety of innovative MR sequences morphological (T2*) and functional aspects (vessel permeability, DCE) of angiogenesis are assessed. Vascular endothelial growth factor (VEGF) inhibition leads to partial vascular normalization with decreased permeability and vessel density (Fig. 1). Resolution of MRI is however limited to the higher µm range. To further resolve the tumor microvasculature we performed correlated UM of fluorescently labeled microvessels in 3DISCO cleared whole brains. UM resolved typical features of abnormal tumor vessel morphology such as increased caliber, density and tortuousness with a spatial resolution of ~5 µm (Fig. 1). Conclusions: UM serves as a complimentary imaging technique that in combination with dedicated MR sequences (MR-UM) allows for precise mapping and high-resolution quantification of tumor neoangiogenesis and treatment responses. Detection of vessel signals at high field can be performed by Gd or USPIO contrast agents.