Description

• The B2M encodes a serum protein found in association with the major histocompatibility complex (MHC) class I heavy chain on the surface of nearly all nucleated cells. The protein has a predominantly beta-pleated sheet structure that can form amyloid fibrils in some pathological conditions. HLA-A belongs to the HLA class I heavy chain paralogues. The heavy chain is anchored in the membrane. Class I molecules play a central role in the immune system by presenting peptides derived from the endoplasmic reticulum lumen so that they can be recognized by cytotoxic T cells. They are expressed in nearly all cells.
• The B2M gene (Exon1 to Exon3) of mouse were replaced by the sequence encompassing the human B2M CDS and HLA-A*0206 gene that included leader sequence, α1 and α2 domains ligated to a fragment of the murine H-2D^b gene containing the α3, transmembrane and cytoplasmic domains.
• Human B2M and HLA-A2.6 were exclusively detectable in homozygous B-HLA-A2.6 mice.
• B-HLA-A2.6 mice provide a powerful preclinical model for in vivo evaluation of vaccines.
• Application: For example, this product is used for pharmacodynamics and safety evaluation of vaccines for cancers.

Targeting Strategy

Gene targeting strategy for B-HLA-A2.6 mice. The B2M gene (Exon1 to Exon3) of mouse were replaced by the sequence encompassing the human B2M CDS and HLA-A*0206 gene that included leader sequence, α1 and α2 domains ligated to a fragment of the murine H-2D^b gene containing the α3, transmembrane and cytoplasmic domains.

Protein Expression Analysis in Spleen and Blood

Strain specific B2M and HLA-A2.6 expression analysis in wild-type C57BL/6JNifdc mice and homozygous humanized B-HLA-A2.6 mice by flow cytometry. Splenocytes (A) and blood (B) were collected from wild-type C57BL/6JNifdc mice (+/+) and homozygous B-HLA-A2.6 mice (H/H), respectively, and analyzed by flow cytometry with species-specific anti-HLA-A2.6 (Biolegend, 311406), anti-H-2Db (Biolegend, 111513), anti-hB2M (Biolegend, 395712), and anti-mB2M (BD Biosciences, 744802) antibody. HLA-A2.6 and human B2M were exclusively detectable in homozygous B-HLA-A2.6 mice.

Frequency of Leukocyte Subpopulations in Spleen

Frequency of leukocyte subpopulations in spleen by flow cytometry. Splenocytes were isolated from wild-type C57BL/6JNifdc mice and homozygous B-HLA-A2.6 mice (female,7-week-old, n=3). A. Flow cytometry analysis of the splenocytes was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. Frequencies of T cells, B cells, NK cells, DCs, monocytes, macrophages, neutrophils and Tregs in B-HLA-A2.6 mice were similar to those in C57BL/6JNifdc mice. Percentage of CD8+ T cells in B-HLA-A2.6 mice were lower than in C57BL/6JNifdc mice, whereas the percentage of CD4+ T cells in B-HLA-A2.6 mice were higher than in C57BL/6JNifdc mice. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test.  *P < 0.05, **P < 0.01, ***p < 0.0001.

Frequency of Leukocyte Subpopulations in Blood

Frequency of leukocyte subpopulations in blood by flow cytometry. Blood were isolated from wild-type C57BL/6JNifdc mice and homozygous B-HLA-A2.6 mice (female,7-week-old, n=3). A. Flow cytometry analysis of the blood was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. Frequencies of T cells, B cells, NK cells, DCs, monocytes, macrophages, neutrophils and Tregs in B-HLA-A2.6 mice were similar to those in C57BL/6JNifdc mice. Percentage of CD8+ T cells in B-HLA-A2.6 mice were lower than in C57BL/6JNifdc mice, whereas the percentage of CD4+ T cells in B-HLA-A2.6 mice were higher than in C57BL/6JNifdc mice. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test.  *P < 0.05, **P < 0.01, ***p < 0.0001.

Frequency of Leukocyte Subpopulations in Lymph Node

Frequency of leukocyte subpopulations in lymph node by flow cytometry. Lymph node cells were isolated from wild-type C57BL/6JNifdc mice and homozygous B-HLA-A2.6 mice (female,7-week-old, n=3). A. Flow cytometry analysis of the lymph node cells  was performed to assess the frequency of leukocyte subpopulations. B. Frequency of T cell subpopulations. Frequencies of T cells, B cells, NK cells and Tregs in B-HLA-A2.6 mice were similar to those in C57BL/6JNifdc mice. Percentage of CD8+ T cells in B-HLA-A2.6 mice were lower than in C57BL/6JNifdc mice, whereas the percentage of CD4+ T cells in B-HLA-A2.6 mice were higher than in C57BL/6JNifdc mice. Values are expressed as mean ± SEM. Significance was determined by two-way ANOVA test.  *P < 0.05, **P < 0.01, ***p < 0.0001.