Display omitted •There are approved targeted drugs for NSCLCs carrying EGFR, BRAF and MET mutations.•NSCLC testing guidelines include the analysis of ALK, ROS1, RET and NTRK gene fusions.•NSCLC emerging targets are KRAS G12C substitutions and HER2 mutations and amplifications.•The above markers can be analysed by DNA/RNA analysis, while PD-L1 testing requires IHC.•There is a paucity of predictive markers for squamous NSCLC and for cytotoxic therapy. Molecular testing has become a mandatory component of the non-small cell lung cancer (NSCLC) management. The detection of EGFR, BRAF and MET mutations as well as the analysis of ALK, ROS1, RET and NTRK translocations have already been incorporated in the NSCLC diagnostic standards, and the inhibitors of these kinases are in routine clinical use. There are emerging biomarkers, e.g., KRAS G12C substitutions and HER2 activating alterations, which are likely to enter NSCLC guidelines upon the approval of the corresponding drugs. In addition to genetic examination, NSCLCs are usually subjected to the analysis of PD-L1 protein expression in order to direct the use of immune checkpoint inhibitors. Comprehensive NSCLC testing for multiple predictive markers requires the analysis of distinct biological molecules (DNA, RNA, proteins) and, therefore, the involvement of different analytical platforms (PCR, DNA sequencing, immunohistochemistry, FISH). There are ongoing efforts aimed at the integration of multiple NSCLC molecular assays into a single diagnostic pipeline.