1024-LB: HLA DQA and DQB epitopes: Determined by alloantibodies adsorbed and eluted from recombinant HLA DQ single antigen cell lines and the antibodies’ reaction patterns with HLA DQ single antigen beads

Aim In 2008, we defined 18 HLA DQ epitopes using mouse monoclonal antibodies and alloantibodies adsorbed and eluted (Ads/Elut) from HLA DQ homozygous cell lines. Here, we confirm 11 of the 18 epitopes and define 10 NEW epitopes of alloantibodies purified by Ads/Elut using recombinant single antigen cell lines. Methods Alloantibodies were purified from the sera of 36 kidney transplant patient by Ads/Elut using HLA DQ recombinant single antigen cell lines (rHLA-DQ). Each serum was Ads/Elut using several different cell lines to insure monoclonality. Eluted antibody specificities were determined by the HLA Class II Single Antigen (SA) beads. Epitope search software was used to identify amino acids exclusively shared among all positive DQ heterodimers (DQA or DQB) on the SA beads. These amino acids define the epitope. Results Ten new HLA DQ epitopes, 2 Alpha & 10 Beta chain, are defined (partial list in table below) and eleven previously defined DQ epitopes were confirmed. Donor Specific Antibodies (DSA) were found in almost all sera. And in most sera, we found more than one antibody positive with the same mismatched antigen, but each targets a different epitope. When anti DR or DP antibodies were present in the sera, the eluted antibodies were negative with the DR and DP beads indicating only DQ-specificities. Partial list of new DQA,B epitopes. Serum # mismatched DQ (A or B chain) Ads/Elut Cell # Cell heterodimer DQA.DQB Antigens positive with eluted antibody Epitope location (chain) Position/aa define epitope #02 DQA1*05:01 nn NA DQA4,5,6 A 40G #10 DQB1*05:02 RC893 DQA102.DQB502 DQB2,5,6 B 140A #10 DQB1*05:02 RC753 DQA301.DQB201 DQB2,5 B 77R #14 DQB1*03:01 RC883 DQA301.DQB303 DQB6,7,8,9 B 75L #22 DQB1*06:02 RC841 DQA102.DQB602 DQB6 B 87F #31 DQB1*05:01 nn na DQB1*05:01 B 125S+126Q nk = not known, nn = not needed, na = not applicable, xx = exact allele not known Conclusions Using rHLA-DQ single antigen cell lines to purify anti HLA-DQ antibodies and testing the antibodies with the HLA-DQ single antigen beads proved very effective in defining DQA and DQB epitopes.