“Humanized” mouse models of immunity refer to normal, immunocompetent mice expressing human genes via transgenesis (e.g., HLA or human immunoglobulin transgenic mice) or to immunodeficient mice engrafted with human hematopoietic and lymphoid cells or tissues. The protocols presented in this unit describe approaches for generating the latter type of humanized mice. Two basic protocols describe generating humanized mice: Basic Protocol 1 deals with hematopoietic stem cell (HSC) engraftment (human SCID repopulating cell; hu-SRC) and Basic Protocol 2 addresses engraftment with human peripheral blood mononuclear cells (PBMC) (human peripheral blood leukocyte; hu-PBL). Additionally, Alternate Protocols 1 and 2 respectively describe procedures for creating hu-SRC and hu-PBL severe combined immunodeficient (SCID) mice that differ from Basic Protocols 1 and 2 in timing and/or route of engraftment with human cells. Finally the Support Protocol outlines the steps needed to verify engraftment levels in humanized mice. These methods are complimentary, each with its individual strengths and limitations, and can be used to address different experimental questions. Creation of another type of humanized mouse, the SCID-hu mouse, a model where human fetal thymus and liver tissues are transplanted into the renal subcapsular space of immunodeficient mice, is detailed in UNIT 4.8.
The main advantage of the HSC engraftment model (hu-SRC-SCID) is that the human T and B cells develop from human stem cells engrafted in the mouse, undergo negative selection during differentiation into T and B cells, and are therefore tolerant of the mouse host. This model allows for investigation of hematopoietic lineage development and mechanisms of immune system development and the generation of primary immune responses by a naïve immune system.
The PBMC model (hu-PBL-SCID) utilizes leukocytes isolated from peripheral whole blood or spleen and allows for rapid analysis of human immune function because the transferred lymphocytes are functionally mature. This model is best suited for studies of immune function from patients with immunologic disorders, analyses of antigen recall responses, investigations of allograft rejection, and other short-term (~4-week) experiments. The sources and availability of HSC and PBMC, methods of engraftment, and evaluation of reconstitution are considered in this unit.
NOTE: Human tissues, mice engrafted with human tissues, as well as the mouse bedding and caging from humanized mice, should be considered potential biological hazards and handled with proper personal protective equipment at animal biosafety level 2 (ABSL2), in accordance with governmental and institutional biosafety guidelines.
NOTE: Immunodeficient mice should be housed in a specific pathogen free (SPF) environment, using sterile techniques and microisolator caging. See UNIT 1.2 for details on care of immunodeficient mice.