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These downloadable slides review the biology and clinical significance of ROS1 rearrangements in non-small-cell lung cancer (NSCLC), with an emphasis on optimizing biomarker detection and translating molecular findings into treatment selection. The content compares available testing approaches, including FISH, immunohistochemistry, plasma-based testing, and DNA- and RNA-based next-generation sequencing, and highlights practical considerations for comprehensive molecular profiling and accurate identification of ROS1 fusions. It also examines guideline-based treatment recommendations and clinical efficacy and safety data for established and emerging ROS1-targeted therapies, including crizotinib, entrectinib, repotrectinib, taletrectinib, and zidesamtinib, with attention to systemic and intracranial activity, acquired resistance such as the ROS1 G2032R mutation, and outcomes after prior ROS1 TKI exposure. By integrating molecular testing strategies, evolving clinical evidence, resistance patterns, CNS efficacy, toxicity, and patient- and disease-specific factors, this activity is designed to support individualized sequencing and selection of ROS1-directed therapy in clinical practice.