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Human alteration of the nitrogen cycle has stimulated research on nitrogen cycling in many aquatic and terrestrial ecosystems, where analyses of nitrate (NO3 −) by standard laboratory methods are common. A recent study by Colman et al. (Biogeochemistry 84:161–169, 2007) identified a potential analytical interference of soluble iron (Fe) with NO3 − quantification by standard flow-injection analysis of soil extracts, and suggested that this interference may have led Dail et al. (Biogeochemistry 54:131–146, 2001) to make an erroneous assessment of abiotic nitrate immobilization in prior 15N pool dilution studies of Harvard Forest soils. In this paper, we reproduce the Fe interference problem systematically and show that it is likely related to dissolved, complexed-Fe interfering with the colorimetric analysis of NO2 −. We also show how standard additions of NO3 − and NO2 − to soil extracts at native dissolved Fe concentrations reveal when the Fe interference problem occurs, and permit the assessment of its significance for past, present, and future analyses. We demonstrate low soluble Fe concentrations and good recovery of standard additions of NO3 − and NO2 − in extracts of sterilized Harvard Forest soils. Hence, we maintain that rapid NO3 − immobilization occurred in sterilized samples of the Harvard Forest O horizon in the study by Dail et al. (2001). Furthermore, additional evidence is accumulating in the literature for rapid disappearance of NO3 − added to soils, suggesting that our observations were not the result of an isolated analytical artifact. The conditions for NO3 − reduction are likely to be highly dependent on microsite properties, both in situ and in the laboratory. The so-called “ferrous wheel hypothesis” (Davidson et al., Glob Chang Biol 9:228–236, 2003) remains an unproven, viable explanation for published observations. [ABSTRACT FROM AUTHOR]