Gene targeting strategy for B-hTLR4 mice. The exons 1~3 of mouse Tlr4 gene were replaced by human TLR4 exons 1~3 in B-hTLR4 mice.

Strain specific analysis of TLR4 mRNA expression in wild-type C57BL/6 mice and B-hTLR4 mice by RT-PCR. Spleen RNA were isolated from wild-type C57BL/6 mice (+/+) and homozygous B-hTLR4 mice (H/H), then cDNA libraries were synthesized by reverse transcription, followed by PCR with mouse or human TLR4 primers. Mouse Tlr4 mRNA was detectable only in wild-type C57BL/6 mice. Human TLR4 mRNA was detectable only in homozygous B-hTLR4 mice but not in wild-type mice.

Strain specific TLR4 expression analysis in wild-type and homozygous B-hTLR4 mice by flow cytometry. Peritoneal cells were collected from wild-type C57BL/6 mice (+/+) and homozygous B-hTLR4 mice (H/H). Flow cytometry analysis of the peritoneal macrophages were detected with anti-TLR4 antibody. Mouse TLR4 was detected on peritoneal macrophages in wild-type mice. Human TLR4 was detected on peritoneal macrophages in homozygous B-hTLR4 mice.