USE OF MONOCLONAL-ANTIBODIES TO DETECT DISEASE-ASSOCIATED HLA-DRB1 ALLELES AND THE SHARED EPITOPE IN RHEUMATOID-ARTHRITIS

Objective-To use a panel of monoclonal antibodies (Mab) which recognis e HLA class II alleles associated with rheumatoid arthritis for fluore scence activated cell sorter (FACS) analysis of peripheral blood monon uclear cells (PBMNC) from patients with early and established rheumato id arthritis and to compare these results against DNA oligotyping of H LA class II molecules in the same patients. Methods-27 patients (18 fr om an early arthritis clinic, nine with established rheumatoid arthrit is) were studied using both techniques. PBMNC were stained with Mab wh ich recognise the shared epitope, the HLA-DRB104 molecule and its *04 01, 0404 subtypes in the presence of bound peptide. Mab stained cells were analysed by FACS. Genomic DNA was prepared from PBMNC and used f or DNA oligotyping and sequencing by standard methods. Results-FACS an alysis of Mab stained PBMNC gave identical results to those obtained b y DNA oligotyping in 26/27 patients. The antibodies identified the sha red epitope in 14/14 cases and the presence of an HLA-DRB104 molecule in 12/12 cases. HLA-DRB10404 was identified in 4/4 cases. HLA-DRB1*0 401 was identified in 5/6 cases. One patient oligotyped as HLA-DRB104 01, but consistently failed to react with the 0401 Mab. DNA sequencin g of the second exon of the HLA-DRB10401 allele in this patient confi rmed a normal HLA-DRB10401 genotype. Conclusions-FACS analysis of PBM NC stained with Mab recognising the shared epitope and rheumatoid arth ritis associated HLA susceptibility molecules provides a rapid, reliab le, and more accessible alternative to DNA oligotyping. The apparent d iscordance between phenotypic and genetic analysis of HLA-DRB10401 in one patient, may reflect variability in HLA-DRB10401 gene expression or in class II peptide presentation.