How to Choose Immunodeficient Mice for Human Disease Research
The lack of appropriate animal models often limits the study of human pathologies. Mice are among the most commonly used models in biomedical research, yet many human-specific pathogens, tumors, and immune responses cannot be faithfully reproduced in a normal mouse.
Since the late 1980s, when the first SCID-hu and hu-PBL-SCID models were reported, researchers have created so-called humanized mice by engrafting human cells or tissues into immunodeficient hosts. These models reproduce key aspects of human disease and immunity in vivo, opening the door to new therapeutic approaches.
Immunodeficient laboratory mice lack some or all components of a functional immune system, so they do not reject human cells. They serve as hosts that can accept human tumor cells, patient-derived tumor tissues, immune cells, or hematopoietic stem cells, allowing human genes, cells, pathologies, and drugs intended for human use to be studied in vivo.
A few tips on choosing the suitable immunodeficient mouse model are provided below.
Define Your Research Objectives
No single immunodeficient mouse model works for every situation. A model must be chosen according to the study's research aim and the constraints that aim imposes. Defining exactly which question the research needs to answer is therefore the first step in selecting an immunodeficient mouse model.
After specifying what information the animal model must provide, the next step is to determine how the model will provide it—for example, which human cells or tissues will be engrafted, how long the study will run, and which readouts will be measured. These decisions guide the rest of the model selection process.
The common strains of immunodeficient mouse models include
Nude Mice
The nude mutation was first observed by Grist in 1962 and described genetically by Flanagan in 1966; in 1968, Pantelouris showed that nude mice lack a thymus. The mutation, later transferred onto inbred backgrounds such as BALB/c, causes both hairlessness and an athymic state—the absence of a functional thymus. Its genetic basis is a mutation in the Foxn1 gene (formerly known as Whn or Hfh11). Nude mice have very few mature T cells and therefore mount poor T cell-dependent antibody responses, producing mainly IgM, while their B cells are present and their natural killer (NK) cell activity is elevated.
Nude mice are widely used for xenografting human tumor cell lines, studying mechanisms of tumor growth and malignancy, and testing treatment options.
NOD Mice
The non-obese diabetic (NOD) mouse was developed in Japan in 1980 by Makino and colleagues from the outbred ICR stock. NOD mice spontaneously develop autoimmune type 1 diabetes (T1D) and are valuable models for studying T1D, autoimmune disorders, and tolerance mechanisms. The disease shares many features with human T1D, including autoreactive CD4+ and CD8+ T cells, islet-specific autoantibodies, and similar genetic susceptibility loci.
The model is popular, although critics point out that many interventions that prevent diabetes in NOD mice have not yet achieved the same outcome in humans. Nevertheless, the striking similarities between NOD mice and the human disease explain why NOD mice are used in research so frequently.
NOD SCID Mice
NOD SCID mice carry the Prkdcscid mutation on the NOD background and lack functional T and B lymphocytes. Combined with the innate immune defects of the NOD strain, this allows better engraftment of human hematopoietic cells than earlier scid models, and the strain has been used to build models such as human HIV-1 infection.
The scid mutation impairs the recombination of antigen receptor genes and thus blocks the early development of B- and T-lineage cells. Beyond this lymphocyte defect, the NOD background further impairs NK cell, macrophage, dendritic cell, and complement function. NOD SCID mice show low "leakiness," meaning that only a small proportion of animals generate some functional T and B cells as they age. These mice are useful for studying normal and abnormal lymphocyte development, as well as for studying nonlymphoid cells in the absence of lymphocytes.
NKG Mice
NKG mice carry the Prkdcscid mutation and an Il2rg knockout on the NOD background. Loss of the common gamma chain (IL-2Rγ) blocks NK cell development, so mature T, B, and NK cells are all absent. In addition, the NOD background contributes complement deficiency and reduced macrophage phagocytosis of human-derived cells (NOD-type SIRPα binds human CD47 and delivers a "don't eat me" signal). This model can be used for transplants of peripheral blood mononuclear cells (PBMCs), patient-derived xenografts (PDXs), and hematopoietic stem cells (HSCs). Because of their high level of immunodeficiency, NKG mice have been used to study tumors, human immunity, immunotherapies, autoimmune diseases, and graft-versus-host disease (GVHD) and transplantation.
Outline Intrinsic Factors: Host and Graft
Once the experimental requirement has been identified, the next step is to outline the intrinsic factors of the model—the properties of the mouse host and of the human graft, and how the two interact. Overlooking these basic considerations early on can easily result in a model that is not fully appropriate for the research project.
On the host side, the key question is which immune components remain. Residual NK cells (as in nude and NOD SCID mice), host macrophages, and complement can reject or clear human cells and limit engraftment. The genetic background also matters: for example, the NOD background supports human cell engraftment better than many other strains.
On the graft side, different human materials have different requirements. Tumor cell lines usually grow in moderately immunodeficient hosts such as nude mice, whereas primary tumor tissues (PDX), PBMCs, and HSCs generally require more severely immunodeficient strains such as NKG mice. The type of graft also determines the time window of the study: PBMC-engrafted mice develop xenogeneic GVHD within weeks, while HSC-engrafted mice can develop multiple human immune cell lineages and support longer-term studies.
Finally, the host and graft interact. Human cells depend on cross-reactive mouse cytokines and growth factors to survive and differentiate, and some human lineages develop poorly in mice for this reason. Understanding these host–graft interactions together defines the intrinsic factors of the model.
Outline Extrinsic Factors
In addition to intrinsic parameters, extrinsic factors affect study outcomes even though they are not part of the host–graft interaction itself. They are not usually considered part of the animal model, but they are crucial for reproducibility.
The way the human cells or tissues are prepared, handled, and administered—including cell viability, dose, injection route, and any pre-conditioning such as irradiation—can strongly affect engraftment. Donor-to-donor variability in PBMCs, HSCs, or tumor samples also needs to be considered. Animal-related conditions matter too: age, sex, housing, bedding, temperature, light cycles, diet, and microbiological status can influence engraftment, the residual immune response, and the pharmacokinetics of the agents being tested. Because immunodeficient mice are highly susceptible to infection, barrier housing with sterile food, water, and caging is essential.
Extrinsic factors extend a study's experimental design and should be defined and documented (for example, following the ARRIVE guidelines) to allow comparison between studies and to improve the ability to extrapolate results to human disease. To understand and control an animal model, intrinsic and extrinsic factors must be evaluated together as part of the experimental design.
Define a Search Framework
Assessing the relevance of the intrinsic and extrinsic factors identified may require a brief preliminary literature review using freely available resources, covering published studies in immunodeficient mouse models as well as relevant human clinical data. This review helps to develop a detailed search strategy.
Based on the preliminary review, a comprehensive, systematic literature search can then be designed. Often overlooked, this step provides the information needed to make scientifically informed decisions throughout model identification and development, and helps determine which factors are truly relevant to the biological question of interest.
Because no single database contains all relevant information, a comprehensive search strategy should cover multiple sources—starting with the most relevant free resources and, where appropriate, incorporating proprietary data such as vendor model characterization data.
Identify the Right Genetic Background
Mice with different genetic backgrounds exhibit very different phenotypes and are suited to different applications. NKG mouse models are highly immunodeficient and widely used in biomedical research. One of their most important features is the NOD genetic background, which contributes innate immune defects including reduced NK cell activity, complement deficiency, impaired macrophage function, and defective antigen-presenting cells. NOD-type SIRPα also reduces macrophage phagocytosis of human cells, which is one reason why NOD-based strains support particularly efficient engraftment of human HSCs.
Research Your Model
Carefully consider the immune components of a model before choosing it. For each candidate immunodeficient model, examine the remaining function of B cells, T cells, dendritic cells, macrophages, NK cells, and complement to understand which endogenous immune responses may still occur.
In nude mice, for example, intact innate immunity—especially active NK cells—can limit the homing and survival of primary human cells. Human tissues, hematopoietic stem cells, and PBMCs engraft much more readily in models lacking the common gamma chain, such as NKG mice, in which signaling through IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 is abolished and NK cells fail to develop.
Comparing the immune components of different immunodeficient strains side by side will give an overview of how they differ as xenograft hosts and help identify the most suitable one.
Understand the Characteristics of the Model
Leakiness. Some strains carrying the scid mutation are "leaky": a proportion of the mice produce some functional T and B cells, and this increases with age. Leakiness depends on genetic background—it is relatively high in C.B-17 scid mice, low in NOD SCID mice, and negligible in NKG mice. Rag1 or Rag2 knockout mice do not show leakiness.
Lifespan. When studying long-term engraftment, such as primary tumors that take months to grow, avoid models with short lifespans. NOD SCID mice frequently develop spontaneous thymic lymphomas (associated with the endogenous retrovirus Emv30) and have a median lifespan of only about 8.5 months. NKG mice, which lack Il2rg, do not develop these thymic lymphomas and can live for more than 1.5 years, making them well suited to long-term studies.
Husbandry. Immunodeficient strains require special housing and care compared with immunocompetent strains, typically barrier facilities with sterilized food, water, bedding, and cages.
Radiosensitivity. For experiments that involve radiation therapy or irradiation conditioning, radiosensitivity is an important consideration. Scid mice carry a mutation in Prkdc, which encodes DNA-PKcs, a key enzyme in DNA double-strand break repair (non-homologous end joining); they are therefore highly sensitive to radiation. Rag1 or Rag2 knockout mice, which have normal DNA repair, tolerate higher doses of radiation and DNA-damaging chemotherapeutics and may be more suitable for such studies.
Background features. Consider important characteristics of the background strain, such as MHC haplotype, behavior, and disease susceptibility. The NOD strain, for example, develops spontaneous diabetes and has defects in NK cell, macrophage, antigen-presenting cell (APC), and complement function. Note that NOD SCID and NKG mice do not develop spontaneous diabetes, because the disease depends on T cells.
Breeding performance. Homozygous nude females have underdeveloped mammary glands, lactate poorly, and have reduced fertility, so nude colonies are usually maintained by mating heterozygous (nu/+) females with homozygous (nu/nu) males.
We hope this review has given a thorough overview of the various immunodeficient mouse models, along with helpful, practical advice on selecting and using them. To make the selection process easier, please call Cyagen at 800-921-8930 / +1 408-969-0306 or email us at inquiry@cyagen.com.
Frequently Asked Questions (FAQs)
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