X-ray absorption spectroscopy (XAS) is a powerful, high-resolution, local structure-determination technique that has been widely applied to studying metal sites in biological systems. The ability to simultaneously measure the geometric structure from the extended X-ray absorption fine-structure (EXAFS) region of the spectrum and the electronic/chemical structure from the pre- and near-edge XAS region has been applied to a wide range of metallo­enzymes, proteins and cofactors containing transition-metal active sites and to nonmetals that are present as ligands or other biological agents. Biological samples are temperamental to prepare, susceptible to beam-related radiation damage and are typically only available at low concentrations. Here, commonly employed methods of preparing and measuring biological samples for XAS measurements are discussed. These methods, along with advanced in-hutch instrumentation and detection modalities, aim to alleviate the challenges faced by these sensitive samples.