Serologic dissection of HLA-D specificities by the use of monoclonal antibodies

Abstract

To study the gene products of the HLA complex, we produced two monoclonal antibodies, termed HU-18 and HU-23. They were active in complement-dependent cytotoxicity and detected B-cell alloantigens encoded by a locus (or loci) linked to HLA. When three types of HLA-DR4 homozygous B-cell lines with different HLA-D specificities were tested for reactivity with HU-18 and HU-23, they displayed distinct reaction patterns depending on the HLA-D specificities they possessed: EBV-Wa (HLA-DYT homozygous), negative for both HU-18 and HU-23; KT2 and KOB (HLA-DKT2 homozygous), positive only for HU-18; and ER (HLA-Dw4 homozygous), positive for both. These differential reaction patterns were further confirmed by testing against a panel of 17 HLA-DR4-positive peripheral blood lymphocytes with known HLA-D specificities. Thus, these monoclonal antibodies allow us to identify HLA-DYT, HLA-DKT2, and HLA-Dw4 solely by serologic methods. This is the first clearcut serologic identification of these three HLA-DR4-associated HLA-D specificities, which have been indistinguishable by conventional serology and identified only by cellular techniques. It is hoped that immunochemical investigations using HU-18 and HU-23 will advance our understanding of the HLA-D region on a molecular level.

Interaction among human leucocyte antigen–peptide–T cell receptor complexes in cow's milk allergy: the significance of human leucocyte antigen and T cell receptor–complementarity determining region 3 loops

“…Cells were cultured for 72 h, in the presence of 1 µCi/well of [ 3 H] TdR during the final 16‐h period, and the incorporated radioactivity was measured by liquid scintillation counting. To determine restriction molecules for antigen presentation, the T cell lines were cultured with irradiated autologous PBMCs, with or without anti‐HLA class II monoclonal antibodies (mAbs) HU‐4 (anti‐HLA‐DRB1 + DRB5 IgG2a, monomorphic) [19,20], L243 (anti‐HLA‐DRB1 + DRB4 IgG2a, monomorphic, reactivity to DRB3 and DRB5 yet to be determined) [21], HU‐11 (anti‐HLA‐DQ4 + DQ5 + DQ6 IgG2a) [22], HU‐18 (anti‐HLA‐DQ7 + DQ8 + DQ9 IgG2a) [23], HU‐46 (anti‐HLA‐DQ4 IgG2a) [22], and B7/21 (anti‐HLA‐DP IgG1, monomorphic) [24]. The percentage inhibition was calculated using the following formula: [1 ‐ (value obtained with a peptide and a mAb ‐ that with the medium alone)/(value obtained with a peptide and without a mAb ‐ that with the medium alone)] × 100.…”

Evidence that HLA class II-restricted human CD4+ T cells specific to p53 self peptides respond to p53 proteins of both wild and mutant forms

“…To determine restriction molecules for antigen presentation, the T cell lines and clones were cultured with irradiated autologous PBMC, with or without saturating amounts of either anti-HLA class II mAb HU-4 (anti-HLA-DRB1 + DRB5 monomorphic) [32,33], HU-11 (anti-HLA-DQ4 + DQ5 + DQ6) [34], HU-46 (anti-HLA-DQ4) [35], PLM-16 (anti-HLA-DRB3) [36] or B7/21 (anti-HLA-DP monomorphic) [32]. Allogeneic PBMC or mouse L cells transfected with HLA class II genes were also used as APC.…”

<i>Dermatophagoides farinae</i>-1-Derived Peptides and HLA Molecules Recognized by T Cells from Atopic Individuals

“…To determine restriction molecules for antigen presentation, the T cell clones were cultured with irradiated autologous PBMC, with or with out saturating amounts of anti-HLA class II monoclonal antibodies (mAbs) HU-4 (anti-HLA-DRBl+DRB5 lgG2a, monomorphic) [10. II], L243 (anti-HLA-DRBl+DRB4 lgG2a. monomorphic, reactivity to DRB3 and DRB5 yet to be determined) [12], PLMI6 (anti-HLA-DRB3) [13], HU-11 (anti-HLA-DQ4+DQ5 t DQ6 IgG2a) [14], HU-18 (anti-HLA-DQ7+DQ8+DQ9 IgG2a) [15], HU-46 (anti-1 ILA-DQ4 IgG2a) [16], and B7/2I (anti-HLA-DP IgGl, monomorphic) [10]. At the same time, experiments were done in which PBMC of two donors with one shared haplotype and one donor with irrelevant HLA haplo types were used as APC.…”