Navigating metabolomics in the era of immunometabolism

Over the past decade, immunology has undergone a profound metabolic awakening. Long considered a hallmark of tumour cells, altered metabolic states are now recognised as a defining feature of immune cells, whose functions are intricately regulated by their bioenergetic and biosynthetic states. This evolving appreciation of immunometabolism has driven demand for analytical tools that can capture metabolic states in immune cells under diverse physiological and pathological conditions. Metabolomics, the comprehensive profiling of metabolites, provides a powerful means to interrogate immune cell metabolism and can offer real-time insights into pathway activity and cellular bioenergetics. However, for many immunologists, metabolomics remains a technically challenging and conceptually opaque area. In this review, we provide a simple guide for designing initial metabolomics experiments, outlining key methodological approaches and addressing common challenges along with potential solutions. Importantly, we present a step-by-step roadmap for interpreting nutrient tracing data, enabling readers to make sense of complex isotopologue datasets and draw meaningful conclusions. Not only will immunologists be better able to interpret metabolomic data from others, but by lowering the barrier to entry, this review aims to empower them to confidently incorporate metabolomics into their research, ultimately advancing our understanding of immune cell function and its metabolic regulation.