Basic Information
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Targeting strategy
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Gene targeting strategy for B-hSIRPA/hCD47/hCD3E/hCD28 mice.
The exons 2-6 of mouse Cd3e gene that encode the extracellular domain were replaced by human CD3E exons 2-7 in B-hSIRPA/hCD47/hCD3E/hCD28 mice. The exons 2-3 of mouse Cd28 gene that encode the extracellular domain were replaced by human CD28 exons 2-3 in B-hSIRPA/hCD47/hCD3E/hCD28 mice. The exon 2 of mouse Cd47 gene that encodes the IgV domain was replaced by exon 2 of human CD47 gene in B-hSIRPA/hCD47/hCD3E/hCD28 mice. The exon 2 of mouse Sirpα gene that encodes the IgV domain was replaced by exon 2 of human SIRPα in B-hSIRPA/hCD47/hCD3E/hCD28 mice.
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Protein expression analysis
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Strain specific CD3E expression analysis in homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice by flow cytometry.
Splenocytes were collected from wild type (WT) mice (+/+) and homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice (H/H;H/H;H/H;H/H), and analyzed by flow cytometry with species-specific anti-CD3E antibody. Mouse CD3E was detectable in WT mice (+/+). Human CD3E was exclusively detectable in homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice (H/H;H/H;H/H;H/H) but not in WT mice (+/+).
Strain specific CD28 expression analysis in homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice by flow cytometry.
Splenocytes were collected from wild type (WT) mice (+/+) and homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice (H/H;H/H;H/H;H/H), and analyzed by flow cytometry with species-specific anti-CD28 antibody. Mouse CD28 was detectable in WT mice (+/+). Human CD28 was exclusively detectable in homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice (H/H;H/H;H/H;H/H) but not in WT mice (+/+).
Strain specific SIRPA expression analysis in homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice by flow cytometry.
Splenocytes were collected from wild type (WT) mice (+/+) and homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice (H/H;H/H;H/H;H/H), and analyzed by flow cytometry with species-specific anti-SIRPA antibody. Mouse SIRPA was detectable in WT mice and B-hSIRPA/hCD47/hCD3E/hCD28 mice due to the cross-reactivity of antibodies. Human SIRPA was exclusively detectable in homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice (H/H;H/H;H/H;H/H) but not in WT mice (+/+).
Strain specific CD47 expression analysis in homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice by flow cytometry.
Splenocytes were collected from wild type (WT) mice (+/+) and homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice (H/H;H/H;H/H;H/H), and analyzed by flow cytometry with species-specific anti-CD47 antibody. Mouse CD47 was detectable in WT mice (+/+). Human CD47 was exclusively detectable in homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice (H/H;H/H;H/H;H/H) but not in WT mice (+/+).
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Summary
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Protein expression analysis:
Human CD3E, CD28 and CD47 were exclusively detectable on T cells of homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice (H/H;H/H;H/H;H/H) but not in WT mice (+/+), and mouse CD3E, CD28 and CD47 were detectable only in WT mice (+/+).
Human SIRPA was exclusively detectable on mCD11b+ cells of homozygous B-hSIRPA/hCD47/hCD3E/hCD28 mice (H/H;H/H;H/H;H/H) but not in WT mice (+/+), and mouse SIRPA was detectable in WT mice and B-hSIRPA/hCD47/hCD3E/hCD28 mice due to the cross-reactivity of antibodies.
