This study provides extensive data on planktonic N<sub>2</sub> fixation rates across the whole Mediterranean Sea. They show that N<sub>2</sub> fixation occurs in Mediterranean waters during the stratification period, with a clear decreasing trend from the oligotrophic western basin (10–76 μmol m<sup>−2</sup> d<sup>−1</sup>) to the ultra oligotrophic eastern basin (0–0.4 μmol m<sup>−2</sup> d<sup>−1</sup>). Highest rates are measured in the less oligotrophic western basin, between the surface and 75 m-depth, where 45 to 75 % of N<sub>2</sub> fixation are found within the picoplanktonic fraction (<3 μm). While the biogeochemical impact of N<sub>2</sub> fixation in the eastern basin seems negligible, N<sub>2</sub> fixation is able to sustain up to 35 % of new primary production during the stratified period in the western basin. These data disagree with indirect estimates of N<sub>2</sub> fixation based on geochemical tracers and nutrient budgets, which indicates that N<sub>2</sub> fixation increases with increasing N:P ratios and decreasing stable N isotopic signature of particulate organic nitrogen and NO<sub>3</sub><sup>−</sup> from west to east. These results finally point out the need to assess N<sub>2</sub> fixation at a higher temporal resolution in order to better understand the diazotrophs' dynamic under contrasted biogeochemical conditions.