SUMMARY One hundred and thirty coded sera, 60 from patients with systemic lupus erythematosus (SLE) and 70 from patients with other autoimmune rheumatic diseases were tested for deoxyribonucleic acid (DNA) binding activity by five different types of assay. These were enzyme linked immunosorbent assay (ELISA) (distinguishing IgG and IgM anti-ssDNA and anti-dsDNA), Crithidia luciliae, a nitrocellulose filter assay, the Amersham kit, and another modified Farr assay, the radioimmunoassay (RIA) (UK). The Crithidia test was the most specific, none of the controls was positive, but the least sensitive (13% positive only). The RIA (UK) was the most sensitive (57% positive). (Sigma, Poole, Dorset) (50 ig/ml in distilled water) and incubated for one hour at 37°C. Each well was then washed three times with phosphate buffered saline (PBS) (Flow, Ayrshire, Scotland) before the addition of 50 RI of dsDNA (10 Fig/ml) or ssDNA (5 [ig/ml), both in PBS. The dsDNA was prepared by treating calf thymus DNA (Sigma) with 1 U/4g S1 nuclease (Sigma) for 30 minutes at 37°C. The ssDNA was prepared by boiling calf thymus DNA for 10 minutes and cooling on ice for 15 minutes. After overnight incubation at 4°C the plates were washed three times with PBS. These were then treated with 100 RI of

The origins and regulation of autoantibodies in SLE may involve idiotypic cell interactions. The purpose of this study was to determine if SLE patients have T cells reactive with the idiotopes of autoantibodies. Sequences of the variable regions of two DNA-binding autoantibodies (V lambda of antibody B3 and VH of 9G4) were selected according to the predicted location of their idiotypes defined previously by anti-idiotypic antibodies. The sequences were prepared as synthetic 16mer peptides (idiopeptides). Peripheral blood mononuclear cells were prepared from SLE patients (n = 28) and controls (n = 13) and put into multiple microcultures with idiopeptide for 6 days. The frequency of responding cultures was determined as those incorporating thymidine at levels above the mean plus three standard deviations of the control cultures lacking peptide. Of the 28 lupus patients, six responded to B3 idiopeptide and five to the 9G4 idiopeptide. Some patients responded to other idiopeptides, but only one normal individual responded to each reference peptide. The difference between the patient and control responses to all idiopeptides was significant by chi 2 analysis (P = 0.025). We conclude that patients with SLE show evidence of sensitisation of T cells to idiotopes of autoantibodies. Such anti-idiotypic T cells could either provide idiotype-specific help or suppression for autoantibody responses in SLE.

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The antigen-binding specificity of human hybridoma-derived monoclonal autoantibodies (mAb) was analysed with mAbs derived from the spleens of two patients with active systemic lupus erythematosus (SLE). From one patient 72 mAbs (RSP clones) and from the other 173 mAbs (RT clones) were obtained. The binding specificity of these mAbs was analysed by solid- and fluid-phase ELISA against the autoantigens ssDNA, dsDNA, cardiolipin, SmRNP, histones, Sm-D and SS-B (La) synthetic peptides, and foreign antigens including bacterial polysaccharides. In addition, antinuclear antibody activity and anti-dsDNA binding were confirmed by fluorescence staining methods. Reflecting the patient's serological profile, none of the antibodies from the RSP clones reacted with ssDNA or dsDNA but 12 reacted with cardiolipin. In addition, three mAbs reacted with H4, five with U1 RNP, two with Sm-D peptides and 12 with SS-B peptides. In contrast, from the RT fusion, nine mAbs reacted with ssDNA, HI and SS-B peptides, seven with cardiolipin, four with dsDNA, two with Sm-D peptides and one each with H2A, H3 and H4. In many cases one mAb showed reactivity with more than one antigen: for example, mAb RT 72 binds to ssDNA, dsDNA, cardiolipin, H1, H4 and an Sm-D peptide; RT 6 binds to H1, SmRNP and ubiquitinated histone H2A. However, none of the antibodies showed 'across the board' polyreactivity; indeed, the selectivity of the reactions was notable and marked variation in antibody affinity was recorded. Eight of the mAbs bound to Salmonella typhimurium and two to the Klebsiella polysaccharide K-30.(ABSTRACT TRUNCATED AT 250 WORDS)