AbstractSites of acid‐phosphatase activity were found in the differentiating root protophloem ofNymphoides peltataby lead‐salt and by azo‐dye methods. Different substrates revealed different subcellular locations of the enzyme. The substrates β‐glycerophosphate (β‐GP) and naphthol ASBI phosphate revealed enzyme activity at similar sites within the sieve element. These sites included plasmodesmata, dictyosomes and small vacuoles in the cytoplasm. The substrate p‐nitrophenylphosphate (p‐NPP), however, revealed additional sites of acid‐phosphatase activity which were not detectable by either naphthol ASBI phosphate or β‐GP. For example, the inner region of the wall in mature sieve elements showed conspicuous acid‐phosphatase activity only when p‐NPP was used as substrate. The significance of the different locations of acid phosphatase within the sieve element is discussed.The convoluted ER, characteristic of immature sieve elements ofN. peltata, failed to show acid‐phosphatase activity whichever substate was used. By contrast, the stacked ER found in the parietal layer of mature sieve elements showed prominent acid‐phosphatase activity regardless of the substrate used. The demonstration of acid‐phosphatase activity in the stacked ER, and by both lead‐salt and azo‐dye methods, suggests that this organelle is a true site of acid‐phosphatase activity.The onset of acid‐phosphatase activity in the ER in later stages of sieve‐element differentiation is compatible with the view that stacked ER plays a role in the final a