Once thought to be a mere consequence of the state of a cell, intermediary metabolism is now recognized as a key regulator of mammalian cell fate and function. In addition, cell metabolism is often disturbed in malignancies such as cancer, and targeting metabolic pathways can provide new therapeutic options. Cell metabolism is mostly studied in cell cultures , using techniques such as metabolomics, stable isotope tracing, and biochemical assays. Increasing evidence however shows that the metabolic profile of cells is highly dependent on the microenvironment, and metabolic vulnerabilities identified do not always translate to settings. Here, we provide a detailed protocol on how to perform stable isotope tracing in leukemia cells in mice, focusing on glutamine metabolism in acute myeloid leukemia (AML) cells. This method allows studying the metabolic profile of leukemia cells in their native bone marrow niche. Copyright © 2021 The Authors; exclusive licensee Bio-protocol LLC.