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Summary: Metastasis, a complex process, is responsible for most deaths in patients with cancer. Clinically relevant research models are indispensable to advancing our understanding of metastatic mechanisms and developing new treatments. We here describe detailed protocols to establish mouse models for melanoma metastasis using the single-cell imaging system and orthotropic footpad injection. The single-cell imaging system permits the tracking and quantification of early metastatic cell survival, while the orthotropic footpad transplantation mimics aspects of the complex metastatic process.For complete details on the use and execution of this protocol, please refer to Yu et al.1,2 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.