Anti hla dq

To determine HLA I-restricted epitopes from CD8⁺ T cells, a B cell panel containing autologous and allogeneic BCL was used. BCL were pulsed with the specific peptide for 1 h at 10 μg/mL, washed, and then cocultured with CD8⁺ T cell clones in an IFN-γ ELISpot assay. To determine HLA II-restricted epitopes from CD4⁺ T cell clones or lines, autologous BCL were blocked with 10 μg/mL of either anti-HLA-A/B/C (BD Biosciences), anti-HLA-DP (Abcam), anti-HLA-DQ (BioLegend), anti-HLA-DR (BioLegend), or anti-HLA-DP/DQ/DR (BioLegend) antibodies for 30 min and then pulsed with the specific peptide for 1 h at 10 μg/mL. Pulsed BCL (1 × 10³ cells/well) were then washed 3 times with R10, mixed with CD4⁺ T cells (1 × 10⁴ to 2 × 10⁴ cells/well), and tested in IFN-γ ELISpot assay in the presence of 10 μg/mL of anti-HLA antibodies.

Neutralizing antibodies against proinflammatory cytokines (anti-human IFN-γ (eBioscience, 14-7318-85) and anti-human IL-17A (eBioscience, 16-7178-85)) were used during allogeneic antigen stimulation. Anti-HLA-ABC (BioLegend, 311412), anti-HLA-DR (BioLegend, 307612), and anti-HLA-DQ (BioLegend, 361502) antibodies were used to block ADSC surface antigens. All of these antibodies were added at a concentration of 1 μg/ml every 7 days when exchanging the medium.