The drug-resistant cell line EMT6/MTXR was derived from the mouse mammary tumour cell line EMT6/P by growth in increasing concentrations of methotrexate. EMT6/MTXR did not hyperexpress P-glycoprotein and did not show cross-resistance to other drugs of the multi-drug resistance (MDR) phenotype. It is recommended to culture the cells without drug on resuscitation until first passage or growth has been fully established.