The Wnt/beta-catenin signalling pathway is activated in many human hepatocellular carcinomas (HCC). Identification of beta-catenin mutation relies mostly on sequence analysis and/or immunohistochemistry. beta-catenin mutation may also be detected by analysing the expression of its target genes. The GLUL gene encoding glutamine synthetase (GS), for example, appears to be a pertinent marker. The aim of this study was to correlate GS immunostaining and beta-catenin mutations with clinicopathological features in HCC. We found that GS immunostaining had a sensitivity of 90% for the detection of beta-catenin mutations, with 98% specificity, whereas beta-catenin immunostaining had a sensitivity of 63% with 98% specificity. We used the sensitive GS marker to characterize 190 HCC cases. Sixty-eight (36%) cases displayed Wnt/beta-catenin activation. In addition to their well-differentiated pattern, these tumours exhibited significant features such as a homogeneous microtrabeculo-acinar pattern, low-grade cellular atypia, and cholestasis. As these tumours exhibited cholestasis, we hypothesized that beta-catenin acts on specific bile synthesis and/or transport pathways. In conclusion, we propose that GS immunostaining and a cholestatic pattern are relevant criteria for the identification of HCC with beta-catenin mutations. Copyright (c) 2007 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.