Conjunctival melanoma (CM) is an ocular malignancy with a high rate of local recurrences after treatment, and can give rise to deadly metastases. The establishment of a murine model will further our understanding of this disease and allow in vivo testing of new therapies. We therefore analyzed the ability of three CM cell lines to grow orthotopically and spread to distant sites. Furthermore, we determined the characteristics of the xenografts and their metastases.Orthotopic xenografts of human CM were established by subconjunctival injection of three different CM cell lines into NOD/SCID IL2 rγnull mice. Single-cell suspensions were generated from the primary tumors and placed subconjunctivally in another set of mice, which were then screened for metastases. The presence of melanoma markers was determined on the cell lines and during tumor development.Subconjunctival injection of cultured CM cells into immunodeficient mice led to excellent subconjunctival tumor growth in all inoculated mice (n = 101) within 2 weeks; however, no metastases were found at the time of autopsy. Serial in vivo passage of primary tumor cells resulted in metastatic tumors in the draining lymph nodes (n = 21). The CM cell lines, as well as the tumor xenografts and their metastases, were positive for the melanoma markers HMB-45, S100B, and MART-1. Two cell lines and their corresponding xenografts carried a BRAF mutation, the third showed an NRAS mutation.We established a murine model for CM that shows excellent formation of metastases in a pattern that accurately resembles metastatic human CM following in vivo passaging.