Using the profile wall method, we determined the root-length density (RLD) of barley roots growing in large-sized biopores (diameter >2 mm) and in the bulk soil of a Haplic Luvisol down to 200 cm of soil depth. The maximum bulk density in the soil profile (1.52 g cm−3) was recorded in the Bt horizon (41–115 cm of soil depth). The proportion of RLD in biopores over the total RLD increased with increasing soil depth down to the 45–75 or 75–105 cm of soil layer but then decreased again in deeper soil. In contrast to earlier investigations, the maximum percentage of RLD in biopores recorded in this study was only 25 %. Root sampling from individually dissected biopores confirmed that roots did not predominantly grow in biopores. Results suggest that roots can use biopores as preferred pathways for growth through rather compact soil layers, whereas they can possibly leave the biopore and re-enter the bulk soil in deeper, less compact layers.