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Understanding the links between intraspecific trait variability and environmental gradients is an important step toward unravelling the mechanisms that link species performance to environmental variation. Here, we performed a comparative, experimental study to investigate variability of cellular traits in three prokaryotic and three eukaryotic freshwater phytoplankton species along gradients of temperature and nitrogen:phosphorus ratio (N:P) in laboratory microcosms. Temperature and N:P strongly affect phytoplankton growth and are changing due to climate change and eutrophication. Metabolic theory and allometric scaling predict that smaller organisms should be favored at higher temperatures through improved metabolic uptake partly due to greater surface area to volume ratios. In addition, chlorophyll a (chl a) concentration should increase due to higher chlorophyll synthesis in response to light limitation at higher cell densities. We found that cell volume both increased and decreased with temperature, whereas intermediate N:P yielded higher growth rates and more extreme conditions yielded bigger cell volumes. Species growth responses to these gradients were distinct and not related to phylogenetic differences. Meaningfully coupled traits like the chl a fluorescence and cell volume shifted consistently and can improve our understanding of individual cell responses to abiotic drivers. This study showed that intraspecific trait variability of freshwater phytoplankton harbors potential for short term acclimation to environmental gradients. Finally, the high trait variability in some species has strong implications for their ecology and the accuracy of predictions where responses may differ when based on mean or fixed trait values.