Generation of the NeoThy mouse model for human immune system studies.

Document Type


Publication Date



JMG, Humans, Animals, Mice, Immune System, Thymus Gland, Disease Models, Animal, Liver, Research Personnel

JAX Source

Lab Anim (NY). 2023;52(7):149-68.







This study was supported in part by the Wisconsin Alumni Research Foundation, NIH NIAID 75N93021C00004, NIH NHLBI U01HL134764, NIH NHLBI 156HL16518901 the UW Carbone Cancer Center Support Grant P30 CA014520 (M.E.B.), CA034196 (L.D.S.), AI132963 (M.A.B. and L.D.S.), OD026440 (M.A.B. and L.D.S.) and NIDDK-supported Human Islet Research Network (HIRN, DK104218 (M.A.B.). We thank J. Zellner and D. Maya for manuscript editing assistance, and K. Howard for helpful discussion.


Humanized mouse models, created via transplantation of human hematopoietic tissues into immune-deficient mice, support a number of research applications, including transplantation immunology, virology and oncology studies. As an alternative to the bone marrow, liver, thymus humanized mouse, which uses fetal tissues for generating a chimeric human immune system, the NeoThy humanized mouse uses nonfetal tissue sources. Specifically, the NeoThy model incorporates hematopoietic stem and progenitor cells from umbilical cord blood (UCB) as well as thymus tissue that is typically discarded as medical waste during neonatal cardiac surgeries. Compared with fetal thymus tissue, the abundant quantity of neonatal thymus tissue offers the opportunity to prepare over 1,000 NeoThy mice from an individual thymus donor. Here we describe a protocol for processing of the neonatal tissues (thymus and UCB) and hematopoietic stem and progenitor cell separation, human leukocyte antigen typing and matching of allogenic thymus and UCB tissues, creation of NeoThy mice, assessment of human immune cell reconstitution and all experimental steps from planning and design to data analysis. This entire protocol takes a total of ~19 h to complete, with steps broken up into multiple sessions of 4 h or less that can be paused and completed over multiple days. The protocol can be completed, after practice, by individuals with intermediate laboratory and animal handling skills, enabling researchers to make effective use of this promising in vivo model of human immune function.

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