We demonstrate the ability of flow cytometry to determine species specific cellular carbon and chlorophyll content in vivo by using laboratory cultures of phytoplankton encompassing a wide range of cell sizes. When applied to the large Atlantic Meridional Transect flow cytometry dataset, we reveal patterns in the species-specific phytoplankton carbon (C), chlorophyll (Chl) and C:Chl ratio. For Prochlorococcus the range of C:Chl is between 2 – 604; for Synechococcus 0.5 – 558. Using a Random Forest machine learning approach, we show that predictability of phytoplankton C:Chl, dominated by the prevalence of Prochlorococcus, is largely driven by silicate and nitrite concentration in the Atlantic Ocean.