The waterflea Daphnia is an interesting candidate for bioregenerative life support systems (BLSS). These animals are particularly promising because of their central role in the limnic food web and its mode of reproduction. However, the response of Daphnia to altered gravity conditions has to be investigated, especially on the molecular level, to evaluate the suitability of Daphnia for BLSS in space. In this study, we applied a proteomic approach to identify key proteins and pathways involved in the response of Daphnia to simulated microgravity generated by a two-dimensional (2D) clinostat. We analyzed five biological replicates using 2D-difference gel electrophoresis proteomic analysis. We identified 109 protein spots differing in intensity (P