Metastatic spread of cancer cells from the primary tumors to distant vital organs, such as lung, liver, brain, and bone, is responsible for the majority of cancer-related deaths. Development of metastatic lesions is critically dependent on the interaction of tumor cells with the stromal microenvironment. As a multifunctional paracrine signaling factor that is abundantly produced by both tumor and stromal cells, TGFβ has been well established as an important mediator of tumor-stromal interaction during cancer metastasis. Imaging the in vivo dynamic of TGFβ signaling activity during cancer metastasis is critical for understanding the pathogenesis of the disease, and for the development of effective anti-metastasis treatments. In this chapter, I describe several xenograft methods to introduce human breast cancer cells into nude mice in order to generate spontaneous and experimental metastases, as well as the luciferase-based bioluminescence imaging method for quantitative imaging analysis of TGFβ signaling in tumor cells during metastasis.