IUHSCT allows for robust donor engraftment in homozygous Fancd2^–/– mice with WT competitive advantage over time. (A) Generation of Fancd2^–/–, Fancd2^+/−, and WT fetuses and in utero transplantation schema. Parental FA carriers (Fancd2^+/− CD45.2) were mated to generate time-dated FA fetuses. BM from WT (CD45.1) mice was isolated and CD117 enriched by magnetic separation. At E13.5-14.5, fetuses were transplanted via intrahepatic injection with 1 × 10⁶ CD117⁺ selected HSCs. Pups were genotyped and subsequently monitored (WT, n = 5; Fancd2^+/−, n = 10; and Fancd2^–/–, n = 3). Peripheral blood and BM donor chimerism was assessed by flow cytometry (% CD45.1) at various time points after IUHSCT. Multilineage donor chimerism was examined in (B) granulocytes and (C) lymphocytes, including B cells, CD4 helper T cells, and CD8 cytotoxic T cells. (D) LT-HSC chimerism was also measured. (E) HSC and progenitor functionality and resistance to DNA damaging agent mitomycin C was further assessed by CFU assays (n = 3) and (F) secondary transplantation in lethally irradiated WT mice (n = 4). Data represent mean ± standard error of the mean (SEM). Comparisons were performed using analysis of variance (ANOVA) with Tukey multiple comparison test, and a P value of <.05 was considered significant. ∗∗∗∗P < .0001; ∗∗∗P < .001; ∗∗P < .01; ∗P < .05. CFU, colony-forming unit; ns, nonsignificant.