The availability of the complete genome sequence of Arabidopsis thaliana and of large collections of insertion mutants paved the way for systematic studies of gene functions in this organism, thus requiring adapting biochemical and physiological tools to this model plant. For physiological analysis of photosynthesis, methods combining high level of chloroplast purity and preservation of the photosynthetic activity were missing. Here, we describe a rapid method (less than 1 h) to obtain Percoll-purified and photosynthetically active chloroplasts from Arabidopsis leaves retaining almost 90% of the Vmax of photosynthesis measured in the starting leaves from plants grown under a light intensity of 150 μmol photon m−2 s−1 and 80% of their initial photosynthetic rate after 3 h of storage.