The δ15N natural abundance (‰) of the total soil N pool varies at the landscape level, but knowledge on short-range variability and consequences for the reliability of isotopic methods are poorly understood. The short-range spatial variability of soil δ15N natural abundance as revealed by the 15N abundance in spring barley and N2-fixing pea was measured within the 0.15-4 m scale at flowering and at maturity. The short-range spatial variability of soil δ15N natural abundance and symbiotic nitrogen fixation were high at both growth stages. Along a 4-m row, the δ15N natural abundance in barley reference plants varied up to 3.9 ‰, and sometimes this variability was observed even between plants grown only 30 cm apart. The δ15N natural abundance in pea varied up to 1.4 ‰ within the 4-m row. The estimated percentage of nitrogen derived from the atmosphere (%Ndfa) varied from 73-89% at flowering and from 57-95% at maturity. When increasing the sampling area from 0.01 m2 (single plants) and up to 0.6 m2 (14 plants) the %Ndfa coefficient of variation (CV) declined from 5 to 2% at flowering and from 12 to 2% at maturity. The implications of the short-range variability in δ15N natural-abundance are that estimates of symbiotic N2-fixation can be obtained from the natural abundance method if at least half a square meter of crop and reference plants is sampled for the isotopic analysis. In fields with small amounts of representative reference crops (weeds) it might be necessary to sow in reference crop species to secure satisfying N2-fixation estimates.