The stem nitrate test for peppermint (Mentha piperitavar.) is a promising nitrogen (N) management tool. When used properly, this test may aid in obtaining significant savings of fertilizer costs and in the protection of groundwater quality. There are several factors related to environmental conditions, N management, and sampling procedures that have not been evaluated and may confound interpretation of test results. The objective of this study was to measure the response of stem nitrate concentrations to factors that would be expected to influence the test and develop guidelines for the collection of stem tissue. The factors considered here were i) solar radiation effects on both hourly and daily scales; ii) spatial variability; iii) differences between alternative plant materials; and iv) the temporal response of tissue nitrate concentrations to soluble N application. The most influential of these variables were the type of stem material (a 441% effect at p=3.55E‐6) and the number of stems collected to estimate the field mean concentration. It was found that the variance of the sample population and the number of stems required for a given sampling error could be greatly reduced by only collecting stems from within the plant canopy. Collecting only these stems, 30 stems were found to be adequate to estimate the field mean concentration within 10 to 15% of the true population mean (p<0.05). Statistically significant differences in stem nitrate concentrations were produced by variations in solar radiation on both hourly (p<0.05) and day length (p<0.01) scales. When measuring the diurnal response, a 17% reduction in stem nitrate concentration was observed over a nine‐hour period from 12:00 hours to 21:00 hours. On the day length scale, an 80% reduction in incoming solar radiation produced a 29% increase in stem nitrate concentrations after three days of shading. In the analysis of stem nitrate spatial variability, no discernable range of autocorrelation was detected indicating a purely random distribution of stem nitrate concentrations on the l‐150mscale. Given this finding and under the conditions of the analyses (late season with stem nitrate in excess of critical levels), it is not important that samples collected for this test fully cover the field being assessed, despite the intuitive appeal of full‐field sampling as a standard procedure. The response of stem nitrate concentrations to soluble N application was minimal, probably due to plant N status in the test plots being well above the critical deficiency content prior to application. With the data produced from these investigations, users of the peppermint stem nitrate test are presented with a method to collect data in the field whereby N management interpretations of the test can be more consistent and reliable. In addition, these results indicate the need for researchers to fully report the method of sampling employed when presenting finding for stem tissue tests.