Concise reports
Use of monoclonal antibodies to detect disease associated
HLA-DRB1 alleles and the shared epitope in rheumatoid arthritis
a Reid Rheumatology
Laboratory, Burnet Clinical Research Unit, Walter and Eliza Hall
Institute of Medical Research, Parkville, Victoria, Australia
, b Tissue Typing Laboratory, Royal Melbourne
Hospital, Parkville, Victoria, Australia
Correspondence to: Dr Ian Wicks, Reid Rheumatology Laboratory, Burnet Clinical Research Unit, Walter and Eliza Hall Institute of Medical Research, PO Royal Melbourne Hospital, Victoria 3050, Australia.
Accepted for publication 4 September 1996
OBJECTIVE
To use a panel of monoclonal antibodies
(Mab) which recognise HLA class II alleles associated with rheumatoid
arthritis for fluorescence activated cell sorter (FACS) analysis of
peripheral blood mononuclear cells (PBMNC) from patients with early and
established rheumatoid arthritis and to compare these results against
DNA oligotyping of HLA class II molecules in the same patients.
METHODS27 patients (18 from an early arthritis
clinic, nine with established rheumatoid arthritis) were studied using
both techniques. PBMNC were stained with Mab which recognise the shared
epitope, the HLA-DRB1*04 molecule and its *0401, *0404 subtypes in the presence of bound peptide. Mab stained cells were analysed by FACS.
Genomic DNA was prepared from PBMNC and used for DNA oligotyping and
sequencing by standard methods.
RESULTSFACS analysis of Mab stained PBMNC gave
identical results to those obtained by DNA oligotyping in 26/27
patients. The antibodies identified the shared epitope in 14/14 cases
and the presence of an HLA-DRB1*04 molecule in 12/12 cases.
HLA-DRB1*0404 was identified in 4/4 cases. HLA-DRB1*0401 was identified
in 5/6 cases. One patient oligotyped as HLA-DRB1*0401, but consistently
failed to react with the *0401 Mab. DNA sequencing of the second exon
of the HLA-DRB1*0401 allele in this patient confirmed a normal
HLA-DRB1*0401 genotype.
CONCLUSIONSFACS analysis of PBMNC stained
with Mab recognising the shared epitope and rheumatoid arthritis
associated HLA susceptibility molecules provides a rapid, reliable, and
more accessible alternative to DNA oligotyping. The apparent
discordance between phenotypic and genetic analysis of HLA-DRB1*0401 in
one patient, may reflect variability in HLA-DRB1*0401 gene expression
or in class II peptide presentation.
© 1997 by Annals of the Rheumatic Diseases
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