HD5 mAb is specific to B27₂, binds to recombinant B27₂, and blocks interaction with immunoregulatory cell receptors.

<p><b>(A)</b> ELISA results showed the specificity of HD5 and HD6 to B27_<b>2</b> homodimers when challenged for different recombinant HLA class I complexes (-A1,-B7,-B13,-C7,-B27, and-B27_<b>2</b>). Control HC10 and W6/32 antibodies were used as positive and negative control, respectively. <b>(B)</b> Direct ELISA against HLA-G and B27_<b>2</b> homodimers using HD5, HD6, HC10 and W6/32 antibodies showed specificity of HD5 and HD6 to immobilized B27_<b>2</b> homodimers but not to HLA-G homodimers. <b>(C-E)</b> Recombinant B27_<b>2</b>, B27-free-heavy chain and HLA-B27 heterotrimers were immobilized into chips for kinetic characterization by SPR. <b>C)</b> HD5 and ligand (B27_<b>2</b>) have a K^d of 0.32 nM. Immobilized free-heavy chains <b>(D)</b> or HLA–B27 heterotrimers <b>(E)</b> did not interact with HD5 and K^d values were not fitted. <b>(F)</b> Blocking competition experiments in SPR were performed by immobilizing B27_<b>2</b> to a streptavidin chip followed by injection of HD5 to form B27_<b>2</b>-HD5 complexes. Injections of LILRB2, KIR3DL2 and Pirb were assessed and binding events recorded.