The Interaction Of Extracellular Ig-like Domains Of Mouse PTA1(CD226) With Its Ligand Mouse Tage4 (CD155)

Objective To investigate the effect of mPTAl/mCD226 on T lymphocyte differentiation and cytotoxic function in the mixed lymphocyte culture(MLC). To identify the domains of mPTA1 that interact with mPTA1 ligand mouse Tage4. Methods â‘ The eukaryotic cell expression vector p-mPTA1-Fc was constructed and expressed in CHO cells. Rabbits were immunized with protein A column-purified mPTA1-Fc fusion protein, and the mPTA-1 polyclonal antibodies were purified by mPTA1-Fc-coupled affinity column. The specificity of the purified polyclonal antibody was identified by indirect fluorescence staining and FCM analysis. Finally, the effect of mPTAl polyclonal antibody on murine T lymphocyte differentiation and cytotoxic function generated from MLC was evaluated with a standard ⁵¹Cr release assay. â‘¢The DNA fragments encoding mTage4, different domains of mPTAl and human ICAM-1 were amplified by PCR and cloned into the eukaryotic expression vector pDsRed2-N1 or pEGFP-N1 respectively, then transfected into COS-7 cells. Confocal laser scanning microscopy was used to confirm the interaction between mPTAl and mTage4. Results â‘  The mPTAl-Fc fusion protein was expressed and purified successfully. The polyclonal antibody against the extracellular region of mPTA-1 molecule could recognize the mPTA1 expression vector transfected COS-7 cells. â‘¡ Anti-mPTA1 polyclonal antibody markedly inhibited murine CTL differentiation and cytotoxicity in dose-dependent manner. â‘¢DNA sequencing...