Increasing interest in the ability of plants to take up amino acids has given rise to questions on the accuracy of the commonly used bulk method to measure and calculate amino acid uptake. This method uses bulk measurements of 13C and 15N enrichment in plant tissues after application of dual-labelled amino acids but some authors have recommended the use of compound-specific stable isotope (CSI) analysis of the plants' amino acids instead. However, there has never been a direct evaluation of both methods. We conducted a field study applying dual-labelled (13C, 15N) amino acids (glycine, valine, tyrosine and lysine) to soil of a Plantago lanceolata monoculture. Root and shoot samples were collected 24 h after label application and the isotope composition of the plant tissues was investigated using bulk and CSI measurements. Enrichment of 13C in the case of CSI measurements was limited to the applied amino acids, showing that no additional 13C had been incorporated into the plants' amino acid pool via the uptake of tracer-derived C-fragments. Compared with this rather conservative indicator of amino acid uptake, the 13C enrichment of bulk measurements was 8, 5, 1.6 and 6 times higher for fine roots, storage roots, shoot and the whole plant, respectively. These findings show that the additional uptake of tracer-derived C-fragments will result in a considerable overestimation of amino acid uptake in the case of bulk measurements. We therefore highly recommend the use of CSI measurements for future amino acid uptake studies due to their higher accuracy. Copyright © 2009 John Wiley & Sons, Ltd.