HLA-DRB1 allele differences affect DBY/HLA class II complex formation and reactivities of alloantibodies from cGVHD patients to DBY/HLA class II complexes. (A-B) Effect of HLA class II alleles on the expression levels of HLA-DR and DBY/HLA class II complexes. Different HLA-DRB1 alleles and DBY were transfected into 293T cells and analyzed via flow cytometry. Bar plots indicate the MFIs for each HLA-DR (A) and DBY complexed with HLA class II molecules (B). The MFIs are presented as the mean ± standard deviation (SD) of 3 independent experiments. (C) Affinity of plasma alloantibodies from patients with cGVHD to DBY/HLA class II complexes. Bar plots indicate the MFIs for anti-DBY/HLA class II complex antibodies. The MFIs are presented as the mean ± SD of 3 independent experiments. (D) The relationship between DBY/HLA class II complex formation and the affinity of plasma alloantibodies from patients with cGVHD binding to DBY/HLA class II complexes. Pearson correlation coefficient was calculated between the 2 groups of MFIs. R represents the correlation coefficient. The circles indicate individual HLA-DRB1 alleles. HLA-DRB1∗09:01 and HLA-DRB1∗12:01 are indicated by a green circle and a blue circle, respectively, whereas HLA-DRB1∗15:02 is indicated by a red circle.