ar637 ARJ Meeting abstract <p>High-affinity antibodies against β<sub>2</sub>-glycoprotein I</p> Cucnik S Kveder T Ambrozic A Borut B

Department of Rheumatology, University Medical Centre, Ljubljana, Slovenia

 Arthritis Res Ther <p>23rd European Workshop for Rheumatology Research</p> The organizer would like to thank the following companies who have generously supported the 23rd European Workshop for Rheumatology Research: Abbott, Amgen, Aventis, Merck Sharp and Dohme, Novartis, Pharmacia, Schering-Plough, Wyeth Meeting abstracts <p>23rd European Workshop for Rheumatology Research</p> Marseille, France 27 February – 2 March 2003 1478-6354 2003 5 Suppl 1 7 10.1186/ar637 24 2 2003

Background

Antibodies against β2-glycoprotein I (anti-β2GPI) are believed to be of low affinity and thus unable to bind to the free antigen in a solution.

Objective

The aim of our study was to determine the affinity of IgG anti-β2GPI, isolated by affinity chromatography.

Methods

A β2GPI affinity column was prepared by CNBr-activated agarose without spacer arms and human purified unnicked β2GPI. The IgG fraction from the protein G column was applied to the column and bound antibodies were eluted with various solutions: A) 0.1 M glycine / 0.5 M NaCl / 0.1% Tween 20 pH 2.5; B) 0.1 M glycine / 4 M NaCl / 0.1% Tween 20 pH 2.5; C) 0.1 M sodium borate pH 10 and D) 25% ethylene glycol. Eluted fractions containing anti-β2GPI antibodies were neutralised and analysed by ELISA using various binding buffers. The level of anti-β2GPI antibodies in each sample was derived from the standard curve according to the defined dilutions of monoclonal antibodies (AUG are arbitrary units of IgG monoclonals).

Results

Increased concentrations of sodium ions in the binding solution from 0.15, 0.25, 0.50, 1.11, 2.07 and 4.0 M NaCl did not completely prevent the binding between isolated antibodies and β2-GPI (79.8, 65.3, 36.1, 19.9, 12.0 and 8.1 AUG, respectively).

Conclusion

In contrast to the common opinion that all anti-β2GPI autoantibodies are of low affinity, we clearly showed that at least one subset among them was of high affinity.