The estuaries play an important ecological role and are the site of many human activities because of their strategic position at the interface between continental and marine waters. Seine estuary is characteristic of large anthropized estuaries. Long-term management requires better knowledge of the spatial and temporal dynamics of the estuarine food webs. Phytoplankton and microphytobenthos are the main contributor of primary production (PP) in these ecosystems and are at the basis of trophic food webs. These compartments are often reduced to the chlorophyll concentration and their productivity has never been measured along the Seine estuary. The objective of this study was to estimate the PP of these compartments along the salinity gradient. In order to access to measurements at high spatial resolution, high-frequency measurements of modulated fluorescence (PAM) were coupled to low-frequency carbon (13C) incorporation measurements. The primary production measurements have been put into perspective with the dynamics of the physical and chemical parameters and the structure of phytoplankton communities determined by different techniques (microscopy, cytometry, molecular biology). The dynamics of carbon excreted as TEP (Transparent Exopolymeric Particles) and EPS (Exopolymeric substances) were also studied for each compartment. Beyond the innovative methodologies which demonstrate the interest of high-frequency measurements in these highly dynamic ecosystems, this work provides a new insight into the phytoplankton dynamics and the specific richness of the estuarine microbiome and provides a reliable estimate of primary production.