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Volume 3 Supplement 2

21st European Workshop for Rheumatology Research

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Heterogeneous behaviour of anti-beta2-glycoprotein I antibodies on different "high binding" microtiter plates

We recently identified anti-β2GPI antibodies in a high proportion of sera from children with atopic dermatitis (AD) and showed that these anti- β2GPI most probably recognise domain V of β2GPI by contrast to anti-β2GPI from patients with the anti-phospholipid syndrome (APS) which epitopes apparently reside in domain I or IV.

The aim of the present study was to compare the binding of IgG anti-β2GPI in AD and APS on four representative commerciallyavailable types of high binding microtiter plates.

Selected plates: Costar, Nunc, Linbro and Sumilon C. Randomly selected sera from 29 children with AD and sera from 43 SLE patients (24 with secondary APS) were tested by anti-β2GPI ELISA using affinity purified β2GPI. Assays were calibrated with the HCAL, chimeric anti-β2GPI monoclonal antibodies with human γ1 constant regions.

The calibration curves for HCAL were practically the same on all four types of plates. Sera from 7/24 APS patients with medium or high anti-β2GPI levels showed similar binding properties on all four plates, while 3/24 sera expressed values either slightly above or below the cut-off points. On the other hand, anti-β2GPI from AD sera showed very simmilar binding on Costar, Nunc and Linbro plates, while only 3/13 positive sera with the highest values on these 3 types of plates expressed low positive values for IgG anti-β2GPI on Sumilon C plates (Table 1). Except for one serum (low positive on Linbro plate) all sera from SLE patients without APS were negative on all the plates.

Table 1

Our results point to substantial differences in the binding to β2GPI coated on different microtiter plates by anti-β2GPI in AD (with signs of APS) but not by anti-β2GPI in APS. In contrast to the other plate types, Sumilon C plates coated with β2GPI bound only minimally antibodies from AD children. If such anti-β2GPI prove non-thrombogenic, we will be able to increase the specificity of detecting anti-β2GPI relevant for APS by the use of this type of microtiter plates. Alternatively, if both anti-β2GPI specificities prove thrombogenic, we will be able to increase the sensitivity of the assay system by the use of other less discriminatory types of plates.

k, slope of linear regression plot and R2 - determination coefficient: both compared to Costar. N, number of IgG anti-β2GPI positive sera in the group. *P < 0.001 (significant difference when compared with Costar, Nunc or Linbro)

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Ambrozic, A., Kveder, T., Ichikawa, K. et al. Heterogeneous behaviour of anti-beta2-glycoprotein I antibodies on different "high binding" microtiter plates. Arthritis Res Ther 3 (Suppl 2), P081 (2001).

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