Gene targeting strategy for B-hCD5 mice. The exons 2~7 of mouse Cd5 gene that encode the extracellular domain were replaced by human CD5 exons 2~7 in B-hCD5 mice.

Strain specific analysis of CD5 gene expression in WT and homozygous B-hCD5 mice by RT-PCR. Mouse Cd5 mRNA was detectable in thymocytes of wild-type (+/+) mice. Human CD5 mRNA was detectable only in H/H, but not in +/+ mice.

Strain specific CD5 expression analysis in homozygous B-hCD5 mice by flow cytometry. Splenocytes were collected from WT and homozygous B-hCD5 (H/H) mice, and analyzed by flow cytometry with species-specific CD5 antibody. Mouse CD5 was detectable in WT mice. Human CD5 was exclusively detectable in homozygous B-hCD5 but not WT mice.

Strain specific CD5 expression analysis in homozygous B-hCD5 mice by flow cytometry. Spleen was collected from wild-type C57BL/6 mice (+/+) and homozygous B-hCD5 mice (H/H), and analyzed by flow cytometry with species-specific anti-CD5 antibody. mCD5 was detectable in wild-type, hCD5 was only detectable in homozygous B-hCD5 mice.

Strain specific CD5 expression analysis in homozygous B-hCD5 mice by flow cytometry. Spleen was collected from wild-type C57BL/6 mice (+/+) and homozygous B-hCD5 mice (H/H), and analyzed by flow cytometry with species-specific anti-CD5 antibody. mCD5 was detectable in wild-type, hCD5 was only detectable in homozygous B-hCD5 mice.

Frequency of leukocyte subpopulations in spleen by flow cytometry. Splenocytes were isolated from wild-type C57BL/6 mice and homozygous B-hCD5 mice (n=3, 7-week-old). A. Flow cytometry analysis of the splenocytes was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. Percentages of T cells, B cells, NK cells, dendritic cells, neutrophils, monocytes, macrophages, CD4+ T cells, CD8+ T cells and Tregs in B-hCD5 mice were similar to those in C57BL/6 mice, demonstrating that humanization of CD5 does not change the frequency or distribution of these cell types in spleen. The frequency of leukocyte subpopulations in lymph nodes and blood of B-hCD5 mice were also comparable to wild-type C57BL/6 mice (Data not shown). Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test. *P < 0.05, **P < 0.01, ***p < 0.001.

Homozygous mutation of this gene does not result in a phenotype. (Tarakhovsky A et al., 1994)