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Blood, Vol. 106, Issue 12, 3880-3887, December 1, 2005 This Article Abstract Full Text Services Email this article to a friend Alert me to new issues of the journal Rights and Permissions Citing Articles Citing Articles via CrossRef Oxidized 2-glycoprotein I induces human dendritic cell maturation and promotes a T helper type 1 response Blood Buttari et al. 106: 3880 Supplemental materials for: Buttari et al Files in this Data Supplement: Figure S1. Determination of protein carbonyl groups (JPG, 10.8 KB) - To measure the oxidative modification of protein we determined the concentration of carbonyl groups that reacted with 2,4-dinitrophenylhydrazine (DNPH) thus forming the corresponding hydrazone, which we then analyzed spectrophotometrically (Protein carbonyl assay kit, Cayman Chemical Company, Ann Arbor, MI, USA). The concentration of protein carbonyl groups in H2O2-2-GPI was significantly higher than the level in native 2-GPI (**P < 10-4) and in ox-2-GPI. The concentration of protein carbonyl groups in ox-2-GPI was significantly higher than the level in native 2-GPI (*P < .003). The mean value of ox-2-GPI was obtained subtracting the mean value of the absorbance of the complete medium from the mean value of the absorbance of the ox-2-GPI. The carbonyl content per mg of protein (P) is expressed as DNPH per mg of protein. Results are expressed as mean ± SD from 3 different experiments. Figure S2 Flow cytometric analysis of β2-GPI binding to DCs (JPG, 95.2 KB) - Immature DCs (2 × 105) were incubated with native 2-GPI or H2O2-2-GPI (20 µg/ml) for 45 minutes on ice. After two washings in PBS containing 2% fetal calf serum cells were stained with goat anti-human 2-GPI polyclonal antibody (Affinity Biologicals) diluted 1:10. After incubation for 30 minutes at 4°C cells were stained with PE-conjugates anti-goat mAb (Sigma-Aldrich) and with FITC-conjugated mAb anti-CD1a (Becton Dickinson-PharMingen) for 30 minutes at 4°C before analysis by flow cytometry. Note that although native 2-GPI and H2O2-2-GPI both bound to DCs, H2O2-2-GPI yielded higher binding values than native 2-GPI: 80% of H2O2-2-GPI-positive DCs (panel B) versus 69% of native 2-GPI-positive DCs (panel A); 225 MFI for H2O2-2-GPI versus 154 for native 2-GPI (panel C). This Article Abstract Full Text Services Email this article to a friend Alert me to new issues of the journal Rights and Permissions Citing Articles Citing Articles via CrossRef

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