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. 2013 Aug 8;8(8):e71228. doi: 10.1371/journal.pone.0071228

Figure 1. Recombinant soluble HLA-DO expression and characterization.

Figure 1

(A) Design and sequence of the recombinant soluble HLA-DO α and β chain constructs. (B) Soluble co-expressed DO/DM complex was purified using Mags.DO5 monoclonal antibody affinity column. Four replicate samples were resolved on Bis-Tris SDS-PAGE gels in decreasing protein concentrations. Gels were blotted to PVDF membranes and stained for DO (left, anti-His and anti-HA) and DM (right, anti-FLAG and anti-c-Myc) specific tags. Data shown are representative of three independent experiments. (C) SPR sensograms of DR (red trace), DM (blue trace) and co-expressed DM/DO complexes (Mags.DO5 purified) (green trace) binding to anti-HA antibody coupled chip surfaces. (D) SPR sensgrams of binding of DR (red trace), DM (blue trace) and DM/DO (Mags.DO5 purified) (green trace) to anti-His antibody coupled chip surfaces. (E) SPR sensograms showing binding of DM/DO (Ni-NTA purified) (green trace), DM (red trace), and DM/DO (Ni-NTA purified) pre-bound to soluble Mags.DO5 (blue trace) to Mags.DO5 antibody coupled chip surfaces. The SPR experiments are representative of at least two independent trials.