Throughout their circumtropical distribution, bonefish (Albula spp.) play a vital role in local economies as a highly prized sport fish. Recent interest in stock enhancement to sustain bonefish fisheries has led to the recognition that there currently are no data on how to live capture large numbers of adults (potential broodstock), transport them to captive facilities and how to handle them to ensure high survival. The objective of this study was to develop strategies for the capture and relocation of wild bonefish to a marine research holding facility to enable basic research and explore the potential for culturing bonefish for stock enhancement. Bonefish Albula vulpes (Linnaeus, 1758) were captured as they entered or left tidal creeks on Eleuthera, The Bahamas using seine nets and then transported by boat or truck to the laboratory. The relocation process evoked secondary stress responses at the metabolic, osmoregulatory and haematological levels as indicated by changes in blood glucose, lactate, haematocrit and ion values, relative to control fish. Physical and behavioural disturbances were also observed in bonefish that were unable to acclimate to laboratory conditions. Successful laboratory acclimation and long-term holding of wild bonefish was achieved through an adaptive learning process, whereby we identified a series of strategies and handling techniques to facilitate the acclimation of wild adult bonefish to captivity. This knowledge will enable future laboratory research on bonefish and is a prerequisite to the culture of this highly prized sport fish, and other sub-tropical and tropical marine species.