title: Antibodies to beta 2-glycoprotein-I: urea resistance, binding specificity, and association with thrombosis.
abstract: The aim of the present study was to evaluate the urea resistance and binding characteristics of anti-beta 2-glycoprotein I (anti-beta 2GPI) antibodies using standard anticardiolipin (aCL) and anti-beta 2GPI enzyme immunosorbent assays (ELISAs). Sera from patients with antiphospholipid syndrome (APS) (n = 22) and non-APS (n = 24), positive in a standard aCL ELISA, were tested in an anti-beta 2GPI ELISA performed in polystyrene-irradiated ELISA plates. Urea resistance aCL and anti-beta 2GPI ELISAs were performed by measuring the ability of antibodies to recognize antigen in the presence of 2 M urea. The serum dilution after urea treatment (D) expressed as a percentage of the serum dilution without urea treatment (D(o)) corresponding to the same optical density was defined as residual activity (RA = 100 D/D(o)). The higher the RA, the higher the resistance of the antibodies to urea. APS compared to non-APS sera had higher aCL binding (absorbance values ranging between 0.180 and 1.400; median, 0.717 vs 0.120-1.273; median, 0.250, respectively; P < 0.004). Six APS patients' sera had low aCL levels but they expressed RA > or = 30%. Anti-beta 2GPI antibodies were detected in 15 of 22 APS vs 3 of 24 non-APS patients (P < 0.03); RA > or = 30% was detected in 15 of 22 APS vs 1 of 23 non-APS patients (P < 0.004). Using a CL affinity column, antibodies were purified from three APS anti-beta 2GPI negative and three non-APS anti-beta 2GPI-positive patients and tested in a aCL ELISA, using highly purified bovine serum albumin (BSA) as a blocking agent (modified ELISA); reactivity was not detected in two APS and one non-APS sera. On the contrary, the reactivity of the purified antibodies was high when beta 2GPI was incubated with CL in the ELISA plates; thus some anti-beta 2GPI negative sera from APS patients recognized the CL/beta 2GPI complex, rather than CL or beta 2GPI alone. In conclusion, anti-beta 2GPI antibodies are common in the APS patients, but a number of such patients recognize the CL/beta 2GPI complex and not CL or beta 2GPI. Antibodies to either beta 2GPI or the CL/beta 2GPI complex derived from APS sera present a high resistance to urea. Anti-beta 2GPI antibodies of low urea resistance exist in a minority of non-APS patients with autoimmune disease.
authors.affiliation: Department of Pathophysiology, Medical School, National University of Athens, Greece.
authors.forename: P G
authors.initials: PG
authors.lastname: Vlachoyiannopoulos
authors.forename: C
authors.initials: C
authors.lastname: Petrovas
authors.forename: M
authors.initials: M
authors.lastname: Tektonidou
authors.forename: S
authors.initials: S
authors.lastname: Krilis
authors.forename: H M
authors.initials: HM
authors.lastname: Moutsopoulos
mesh_terms: D017152:Antibodies, Antiphospholipid / Q000097:blood* / Q000737:chemistry / Q000378:metabolism; D000918:Antibody Specificity / Q000187:drug effects; D016736:Antiphospholipid Syndrome / Q000097:blood / Q000276:immunology; D001666:Binding Sites, Antibody / Q000187:drug effects; D001667:Binding, Competitive / Q000276:immunology; D002308:Cardiolipins / Q000276:immunology; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D006023:Glycoproteins / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D013927:Thrombosis / Q000276:immunology*; D014508:Urea / Q000737:chemistry*; D053482:beta 2-Glycoprotein I
chemical_list: D017152:Antibodies, Antiphospholipid; D002308:Cardiolipins; D006023:Glycoproteins; D053482:beta 2-Glycoprotein I; D014508:Urea