This report was produced by an Expert Working Group consisting of UK-based researchers with specialist knowledge of the use of animal models of bone marrow ablation and reconstitution. It aims to facilitate the implementation of the Three Rs (Replacement, Reduction and Refinement), with an emphasis on refinement. Bone marrow ablation and reconstitution procedures are done for a number of different purposes and conducted primarily in mice. These procedures can induce significant suffering, classified as ‘severe’, Category E or Category D/E under European, US and Canadian legislation respectively. Specific animal welfare issues were identified and discussed, and practical measures proposed, with the aim of reducing animal use and suffering.