Tumor hypoxia is one of the hallmarks of cancer and has turned into a central issue in cancer treatment since it is associated with malignant tumor progression, resistance to therapy and increased metastatic spread. To date, many aspects of hypoxia-induced changes in tumors remain to be elucidated. This drives the need to develop multi-modal imaging workflows for discovering and identifying key biomolecules in the heterogeneous tumor microenvironment and to investigate the role of essential mineral elements in hypoxia-driven signaling pathways. The microanalytical techniques matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI – for high-throughput multiplexed molecular screening and high-precision molecular identification) and laser ablation-inductively coupled plasma-mass spectrometry imaging (LA-ICP-MSI – for targeted proteomic analysis using metal-conjugated antibodies and untargeted analysis of the metallome) will be pushed to the next level and combined into a novel multi-modal imaging approach for obtaining spatially resolved molecular and elemental information at the cellular level. This project focuses on the development of novel analytical methods, a high-throughput calibration strategy and a multi-modal imaging workflow to combine metallomic, metabolomic, proteomic, lipidomic and morphological information of the breast tumor microenvironment to achieve a more complete picture of hypoxia-driven signaling pathways.