The NSG-CTL mouse model is a humanized mouse model that allows the generation of peripheral human immune responses, particularly CD8+ Cytotoxic T lymphocyte (CTL) responses, and serves as an effective model for studying gene-based therapies. Natural antigen-specific T cell responses in humanized mice are relatively weak and this model was developed to boost antigen specific responses, in this case to HIV, to more closely assess these responses in vivo. We have engineered human T cells that develop in these mice to express a molecularly cloned T cell receptor (TCR) specific to HIV. Cloned TCRs to any antigen can theoretically be used to study specific responses in vivo, as long as the tissue that is manipulated is of the same human leukocyte antigen (HLA) type. The modification of hematopoietic stem cells with antigen specific TCRs allows the development of mature, functional T cells in the periphery of these mice that are specific to that antigen following normal developmental processes. This model has recently been published (Kitchen et al., 2012) and this was a major modification of the Humanized Mouse BLT model published by Melkus et al. (2006). We use Non-obese diabetic (NOD)-Severe Combined Immunodeficient (SCID), common Gamma chain knockout (γc-/-)—or NSG—mice, and implant fetal thymus pieces along with genetically modified CD34+ hematopoietic stem cells (HSCs), isolated from fetal liver, under the kidney capsule to develop into a functional thymic implant. At the same time, we deplete the mouse’s bone marrow by total body irradiation and inject more modified HSCs intravenously for hematopoietic engraftment in the mouse bone marrow. This protocol outlines the procedures to process the fetal tissue, genetically transduce the HSCs using lentiviral vectors expressing a molecularly cloned T cell receptor, and perform the subcapsular kidney implant surgery.