Basic Information
-
Targeting strategy
-
Gene targeting strategy for B-NDG hB2M/HLA-A11.1 mice plus. The promoter and signal peptide sequences of HLA-A2.1, the coding sequences of human B2M gene and the genomic sequences of HLA-A11.1 gene (from the first coding base on exon1 to the last base of the 3‘ UTR) were inserted into mouse Hipp11 locus of B-NDG mice. Thus, a B-NDG background mouse strain with human B2M/HLA genes was obtained. The exons 2-8 of mouse H2-K1 and H2-D1 genes were knocked out in B-NDG H2-K1/H2-D1 KO mice. B-NDG hB2M/HLA-A11.1 mice plus was developed by cross-mating the B2M/HLA humanized mice and B-NDG H2-K1/H2-D1 KO mice together.
-
Protein expression analysis
-
Strain specific B2M expression analysis in wild-type B-NDG mice and homozygous B-NDG hB2M/HLA-A11.1 mice plus by flow cytometry. Splenocytes, blood cells and bone marrow were collected from wild-type B-NDG mice and homozygous B-NDG hB2M/HLA-A11.1 mice plus. Protein expression was analyzed with anti-mouse B2M antibody (Biolegend, 154504) and anti-human B2M antibody (Biolegend, 395712) by flow cytometry. Mouse B2M was detectable in wild-type mice but not in homozygous B-NDG hB2M/HLA-A11.1 mice plus. Human B2M was exclusively detectable in homozygous B-NDG hB2M/HLA-A11.1 mice plus but not in wild-type mice.
Strain specific HLA-A expression analysis in wild-type B-NDG mice and homozygous B-NDG hB2M/HLA-A11.1 mice plus by flow cytometry. Splenocytes, blood cells and bone marrow were collected from wild-type B-NDG mice and homozygous B-NDG hB2M/HLA-A11.1 mice plus. Protein expression was analyzed with anti-mouse H-2Kb/H-2Db antibody (Biolegend, 114613) and anti-human HLA-A,B,C antibody (Biolegend, 311406) by flow cytometry. Mouse H2-K1/H2-D1 was detectable in wild-type mice but not in homozygous B-NDG hB2M/HLA-A11.1 mice plus. Human HLA-A was exclusively detectable in homozygous B-NDG hB2M/HLA-A11.1 mice plus but not in wild-type mice.
-
Frequency of leukocyte subpopulations
-
Frequency of leukocyte subpopulations in bone marrow, spleen and blood by flow cytometry. Bone marrow, splenocytes and blood cells were isolated from B-NDG mice and homozygous B-NDG hB2M/HLA-A11.1 mice plus (male,7-week-old, n=3). A. Flow cytometry analysis of the immune cells was performed to assess the frequency of leukocyte subpopulations. Frequencies of T cells, B cells, NK cells, DCs, neutrophils, monocytes and macrophages in B-NDG hB2M/HLA-A11.1 mice plus were similar to those in B-NDG mice, demonstrating that humanization of HLA-A11.1 does not change the frequency or distribution of these cell types in bone marrow, spleen and blood when compared with those in B-NDG mice. Values are expressed as mean ± SEM.
