Antigen-Binding Diversity of Human Hybridoma Autoantibodies Derived from Splenocytes of Patients with SLE

Abstract

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)

Epigenetic Histone Code and Autoimmunity

“…This result was in good agreement with other data showing that autoantibodies to ubiquitin are detectable by ELISA in about 80% of lupus patients and in 0-13% only of patients with SS, RA, juvenile chronic arthritis, SSc, sarcoidosis, and normal individuals [31]. Human monoclonal antibodies reacting either with ubiquitin or with the UH2A branched peptide were also successfully generated by fusion of a human cell line with splenocytes from a young girl with active SLE who required splenectomy for severe thrombocytopenia [32]. Furthermore, using rabbit antibodies to UH2A peptide, it was possible to visualize UH2A deposits in renal biopsies from lupus patients [33].…”

β2‐Glycoprotein i reactivity of monoclonal anticardiolipin antibodies from patients with the antiphospholipid syndrome

“…Monoclonal aCL from MRL-lpr mice, from normal human subjects, and from SLE patients can be classified into 3 categories in terms of DNA binding: those that bind exclusively to ssDNA, those that bind to both dsDNA and ssDNA, and those without DNA binding (26,33,34,36,39). In this study, monoclonal aCL from patients with the APS did not react with either dsDNA or ssDNA, independent of p2-GPI.…”

Phospholipid binding specificities and idiotype expression of hybridoma derived monoclonal autoantibodies from splenic cells of patients with systemic lupus erythematosus.

“…We reported a detailed analysis of the antigen binding profile of 19 human MAbs from a selection of 245 clones derived from the spleens of two patients with SLE. 10 We have now extended this study and analysed the binding specificity of these aPL antibodies to a variety of negatively charged and neutral phospholipids, and examined their functional effects in anticoagulation assays.…”