After 8-y of elevated CO2, we previously detected greater amounts of total soil nitrogen, suggesting that rates of ecosystem N flux into or out of tallgrass prairie had been altered. Denitrification and associative N fixation rates are the two primary biological processes that are known to control N loss and accumulation in tallgrass prairie soil. Therefore, our objective was to assess the natural abundance of plant and soil 15N isotopes as a cumulative index of potential change in efflux or influx of N into and out of the tallgrass prairie after 8-y of exposure to elevated CO2. Aboveground plant delta 15N values of Andropogon gerardii were close to zero and more positive as a result of elevated CO2, but whole-soil values at the 5–30 cm depth were significantly reduced (6.8 vs 7.3; P<0.05) under elevated CO2-chamber (EC) relative to ambient CO2- chamber (AC). Total, aboveground plant biomass, root-in-growth, extractable N, microbial biomass N, and soil pools collectively exhibited a range of delta 15N values from −2.8 to 7.3. Measurements of surface soil 15N indicate that a change in N inputs and outputs has occurred as a result of elevated atmospheric CO2. In addition to possible changes in denitrification and N2 fixation, other sources of N such as the re-translocation of N to the surface from deeper soil layers are needed to explain how soil N accrues in surface soils as a consequence of elevated CO2. Our results support the notion that C accrual may promote N accrual, possibly driven by high plant and microbial N demand amplified by soil N limitation.