Neutralization of cognate antibodies by HLA-Fc in clinical samples. (A) Changes in anti-A2 and -B7 reactivities in sera treated with A2Fc or B7Fc relative to PBS treatment. Clinical samples reactive to A2 (sera 01-02), or B7 (sera 05-07), or both (sera 03-04) were incubated with PBS, A2Fc, or B7Fc before testing by the single-antigen bead assay. The changes in mean fluorescence intensity (MFI) values after A2Fc or B7Fc treatment compared to PBS treatment are represented as a heatmap for each bead. Each row represents a bead specificity; the left and right panels show HLA-A- and HLA-B-specific beads, respectively. Red and blue hues indicate increased and decreased MFI, respectively. (B-C) Neutralization of anti-A2 and anti-B7 reactivities in titrated serum samples #03 (B) and #07 (C) by cognate HLA-Fc. Serial 1:5 dilutions were performed on the sera before incubation with PBS (green), A2Fc (red), or B7Fc (blue), followed by the single-antigen bead assay. The A2-specific reactivity in (B) was represented by MFI values from A∗02:01, A∗02:03, and A∗02:06 beads; The B7-specific reactivity in (C) was represented by B∗07:02, B∗42:01, and B∗81:01 beads (serum #07 is cross-reactive to these 3 antigens). Broken lines at 5000 MFI indicate the threshold associated with positive lymphocyte crossmatching.