Glioblastoma (GBM) is an aggressive and malignant form of glioma, with a dismal 5% five-year survival rate. The treatment for glioblastoma has largely remained unchanged for over a decade, with the last advance being the addition of a chemotherapeutic agent to a patient’s treatment regimen alongside radiotherapy. Immunotherapy provides a new therapeutic option to enhance cancer survival rates. One such therapy is Chimeric Antigen Receptor (CAR) T cell therapy, which has been incredibly successful in treating haematological cancers. EGFRvIII is a tumour specific mutation of EGFR, occurring in a subset of GBM as well as breast, ovarian and colon cancer. We have identified and generated two EGFRvIII-specific scFv and have successfully generated specific and functional EGFRvIII-specific CAR T cells. Here, we provide both in vitro and in vivo data demonstrating therapeutic EGFRvIII-specific CAR T cells which have been used to successfully treat EGFRvIII-expressing glioblastoma in tumour bearing mice.