Specificity of anti-Ap^1-17 IgG for CENP-A documented by human recombinant CENP-A inhibition of anti-Ap^1-17 IgG binding to KLH-Ap^1-17.

<p>Anti-Ap^1-17 Abs from pt4 (A) and pt14 (B) were diluted in PBS-T20 at the lowest concentration giving 80%–100% of maximal A₄₉₀ in binding assay, and pre-incubated with an equal volume of PBS containing 2-fold serial dilutions of CENP-A (○), CENP-B (•), and CENP-B-derived peptide CBp^1-13 (▪). Following a 2-h incubation, the mixture was added to microtiter plate wells coated with KLH-Ap^1-17. After a 4-h incubation and three washes, bound IgG was detected with HRP-conjugated anti-human IgG (Fc portion) and <i>o</i>-phenylenediamine. Inhibition by Ap^1-17 peptide (□) and by Qp-1a (X) were included as positive and negative controls, respectively. Results are expressed as percentage of binding inhibition. The data are representative of 2 experiments.