β‐catenin plays an important role in the Wnt signaling pathway by activating T‐cell factor (Tcf)/lymphoid enhancer factor (Lef)‐regulated gene transcription. The level of β‐catenin is regulated through GSK‐3β phosphorylation of specific serine and threonine residues, all of which are encoded for in exon 3 of the β‐catenin gene (CTNNB1). Mutations altering the GSK‐3β phosphorylation sites lead to cellular accumulation of β‐catenin and constitutive transcription of Tcf/Lef target genes. Such mutations have previously been found in melanoma cell lines. In our study, primary melanomas and their corresponding metastases were screened for CTNNB1 exon 3 mutations using single‐strand conformation polymorphism and nucleotide sequence analysis. One of 31 primary tumors and 1 of 37 metastases, both originating from the same patient, had a TCT to TTT mutation at codon 45, changing serine to phenylalanine. Immunohistochemical analysis revealed membranous localization of β‐catenin in a majority of the samples. The mutated primary tumor and metastasis, however, displayed widespread cytoplasmic and nuclear expression of β‐catenin. An additional 30% of the primary tumors showed focal cytoplasmic and nuclear staining. Thus, β‐catenin exon 3 mutations are rare in primary as well as metastatic melanomas and do not explain the abnormal cytoplasmic and nuclear localization of β‐catenin found in a relatively large fraction of primary melanomas. © 2001 Wiley‐Liss, Inc.