Normal T-cell development and function in NSGAb°DR1 mice. (A) Human chimerism in blood, spleen, and bone marrow determined using human CD45 staining 20 weeks after reconstitution from a single DR1 allelically matched cord blood donor. n ≥ 6 per group pooled from 3 independent experiments. (B) Human immune cell populations as a percent of human leukocytes in spleens of reconstituted mice. cDCs, classical dendritic cells; pDCs, plasmacytoid dendritic cells; NK, natural killer. (C) Representative flow cytometry dot plots of developing T cells (left) quantified (right) showing double-positive (DP), CD4⁺, CD8⁺, double-negative (DN) and CD4⁺CD25⁺FOXP3⁺ Tregs. n ≥ 5 per group from 2 independent experiments. (D) The CDR3 region of the TCR-β locus was profiled using next-generation ultrahigh-throughput sequencing. Pie segments are displayed in different colors to highlight unique V-gene segments for NSG and NSGAb°DR1 mice. TCR-β repertoire Diversity was calculated using normalized Shannon’s entropy (P > .05). (E) Histogram of CDR3 nucleotide length in CD4⁺ T cells isolated from spleens of reconstituted NSG or NSGAb°DR1 mice. (F) The relative frequency or “evenness” of the CD4⁺ TCRβ repertoire was plotted for the top 100 clonotypes in 2 representative animals in each genotype, with each circle depicting an individual clonotype. (G) Delayed-type hypersensitivity response in OVA-immunized mice displayed as the difference in swelling between OVA-injected left footpad (lfp) and PBS-injected right footpad (rfp). NSGAb°DR1 (mismatched) refers to reconstitution using HSCs from a donor that was negative for the DRB1*01:01 allele. Data are pooled from 2 independent experiments. Bars are the mean ± standard error of the mean (SEM). Statistical analysis performed using 1-way analysis of variance (ANOVA) with Tukey’s multiple comparisons test (A), multiple Student t test corrected for multiple comparisons using Holm-Sidak method (C), nonlinear regression (E), unpaired Student t test (F-G). *P < .05, **P < .01, ***P < .001.