Microbial nitrogen cycling was investigated along a 79 km transect into the Greenland Ice Sheet (GrIS) in early August 2010. The depletion of dissolved nitrate and production of ammonium (relative to icemelt) in cryoconite holes within 7.5 km of the ice sheet margin suggested microbial uptake and ammonification respectively. Nitrogen fixation (<4.2 μmoles C2H4 m 2 day 1 to 16.3 μmoles C2H4 m 2 day 1) was active in some cryoconite holes at sites up to 5.7 km from the ice sheet margin, with nitrogen fixation inversely correlated to concentrations of inorganic nitrogen. There may be the potential for the zone of nitrogen fixation to progressively extend further into the interior of the GrIS as the melt season progresses as reserves of available nitrogen are depleted. Estimated annual inputs of nitrogen from nitrogen fixation along the transect were at least two orders of magnitude lower than inputs from precipitation, with the exception of a 100 m long marginal debris-rich zone where nitrogen fixation could potentially equal or exceed that of precipitation. The average estimated contribution of nitrogen fixation to the nitrogen demand of net microbial growth at sites along the transect ranged from 0% to 17.5%.