AimsWe aimed to evaluate the performances of the Idylla GeneFusion Assay (IGFA) designed to detect, in a single, rapid and fully automated assay,ALK,ROS1,RET,NTRK1,NTRK2andNTRK3gene fusions andMETexon 14 skipping in cancer samples.MethodsBased on a set of tumours enriched in cases with gene fusions, we applied the IGFA to tumour areas of various sizes and tumour cell contents. IGFA results were compared with those obtained with other methods (immunohistochemistry, fluorescent in situ hybridisation, DNA and RNA next-generation sequencing).ResultsWe selected 68 tumours: 49 cases with known gene fusions (8ALK, 8ROS1, 5RET, 7NTRK1, 3NTRK2and 6NTRK3ones) orMETexon 14 skipping mutations (12 cases) and 19 cases with no fusion and noMETmutation. We performed 128 IGFA tests on distinct tissue areas. The global sensitivity and specificity of the IGFA were, respectively, 62.82% and 99.2% with variations between molecular targets and tissue areas. Of note, 72.5% sensitivity and 98.79% specificity were obtained in 37 tissue areas fulfilling the manufacturer’s recommendations (ie, at least 10% of tumour cells in at least 20 mm² of tissue area). The rate of non-conclusive results was higher in small samples with low percentages of tumour cells.ConclusionsThe IGFA could contribute to the rapid detection of targetable gene fusions and mutations, especially in context of rapidly growing cancers requiring urgent therapeutic choices.