The crucial role of atmospheric CO2 in glacial-interglacial transitions is demonstrated by recent ice-core studies that highlight the importance of accurate CO2 records for our understanding of Quaternary climate dynamics. Previous estimates of CO2 levels during the last interglacial stage (the Eemian) relied on measurements on air trapped in the Antarctic Vostok ice core. Due to uncertainties associated with in situ chemical alteration and gas diffusion, there is a need for independent estimates of past CO2 levels. Here we report 33 Eemian CO2 estimates based on the stomatal index of Salix herbacea L. leaves preserved in Greenland sediments. We reconstruct Eemian CO2 levels centered on 250–280 parts per million by volume (ppmv), in general agreement with ice- core data. Two deviating (lower) stomatal estimates may reflect diffusional smoothing of Vostok CO2 data and indicate century- scale Eemian CO2 variability.