MALDI-TOF MS glycan analysis of RhD-specific IgG antibodies (crude data)

<p>Series_A_09037-013_total_IgG.csv<br>MALDI-TOF MS glycan analysis of total IgG from Rhophylac, which was the starting material for the purification of Rhesus D (RhD)-specific IgG antibodies (purification A). The area under the peak is shown for each selected molecular mass (m/z) and presented as percentage of all the indicated area values. The glycan structure and abbreviation for each m/z is shown in Supplementary Figure 2 and the summary of all MALDI-TOF MS analyses is shown in Figure 1 in the main text.</p> <p>Series_A_09037-012_RhD-specific_IgG.csv<br>MALDI-TOF MS glycan analysis of RhD-specific IgG antibodies (purification A). The area under the peak is shown for each selected molecular mass (m/z) and presented as percentage of all the indicated area values. The glycan structure and abbreviation for each m/z is shown in Supplementary Figure 2 and the summary of all MALDI-TOF MS analyses is shown in Figure 1 in the main text.</p> <p>SeriesB_09037-192a total_IgG.csv<br>MALDI-TOF MS glycan analysis (measurement a of a duplicate measurement) of total IgG from Rhophylac, which was the starting material for the purification of RhD-specific IgG antibodies (purification B). The area under the peak is shown for each selected molecular mass (m/z) and presented as percentage of all the indicated area values. The glycan structure and abbreviation for each m/z is shown in Supplementary Figure 2 and the summary of all MALDI-TOF MS analyses is shown in Figure 1 in the main text.</p> <p>SeriesB_09037-192b total_IgG.csv<br>MALDI-TOF MS glycan analysis (measurement b of a duplicate measurement) of total IgG from Rhophylac, which was the starting material for the purification of RhD-specific IgG antibodies (purification B). The area under the peak is shown for each selected molecular mass (m/z) and presented as percentage of all the indicated area values. The glycan structure and abbreviation for each m/z is shown in Supplementary Figure 2 and the summary of all MALDI-TOF MS analyses is shown in Figure 1 in the main text.</p> <p>Series_B_09037-191a_RhD-specific_IgG.csv<br>MALDI-TOF MS glycan analysis (measurement a of a duplicate measurement) of RhD-specific IgG antibodies (purification B). The area under the peak is shown for each selected molecular mass (m/z) and presented as percentage of all the indicated area values. The glycan structure and abbreviation for each m/z is shown in Supplementary Figure 2 and the summary of all MALDI-TOF MS analyses is shown in Figure 1 in the main text.</p> <p>SeriesB_09037-191b RhD-specific_IgG .csv<br>MALDI-TOF MS glycan analysis (measurement b of a duplicate measurement) of RhD-specific IgG antibodies (purification B). The area under the peak is shown for each selected molecular mass (m/z) and presented as percentage of all the indicated area values. The glycan structure and abbreviation for each m/z is shown in Supplementary Figure 2 and the summary of all MALDI-TOF MS analyses is shown in Figure 1 in the main text.</p> <p> </p

FACS analyses of purified RhD-specific IgG antibodies

<p>unstained.001<br>FACS analysis with Rhesus D (RhD)-positive erythrocytes to verify the enrichment of RhD-specific IgGs from total IgG of Rhophylac. The cells in this file are unstained. The cells were gated on the main erythrocyte population in a FSC/SSC blot to exclude fragments and aggregates, analyzed in an anti-human IgG (APC) histogram and shown in the overlay histogram in Supplementary Figure 1 in the main text.</p> <p>only_secondary_antibody.004<br>FACS analysis with RhD-positive erythrocytes to verify the enrichment of RhD-specific IgGs from total IgG of Rhophylac.<br>The cells in this file are only stained with a secondary APC-coupled anti-human IgG antibody. The cells were gated on the main erythrocyte population in a FSC/SSC blot to exclude fragments and aggregates, analyzed in an anti-human IgG (APC) histogram and shown in the overlay histogram in Supplementary Figure 1 in the main text.</p> <p>260_µg_per_ml_total_IgG_from_Rhophylac.002<br>FACS analysis with RhD-positive erythrocytes to verify the enrichment of RhD-specific IgGs from total IgG of Rhophylac.<br>The cells in this file are stained with 260 µg/ml total IgG from Rhophylac and subsequently with a secondary APC-coupled anti-human IgG antibody.The cells were gated on the main erythrocyte population in a FSC/SSC blot to exclude fragments and aggregates, analyzed in an anti-human IgG (APC) histogram and shown in the overlay histogram in Supplementary Figure 1 in the main text.</p> <p>2µg_per_ml_purified_RhD_specific_IgG(from_purification_A).005<br>FACS analysis with RhD-positive erythrocytes to verify the enrichment of RhD-specific IgGs from total IgG of Rhophylac.<br>The cells in this file are stained with 2 µg/ml purified RhD-specific IgG from purification A and subsequently with a secondary APC-coupled anti-human IgG antibody. The cells were gated on the main erythrocyte population in a FSC/SSC blot to exclude fragments and aggregates, analyzed in an anti-human IgG (APC) histogram and shown in the overlay histogram in Supplementary Figure 1 in the main text.</p> <p>2µg_per_ml_purified_RhD_specific_IgG(from_purification_B).007<br>FACS analysis with RhD-positive erythrocytes to verify the enrichment of RhD-specific IgGs from total IgG of Rhophylac.<br>The cells in this file are stained with 2 µg/ml purified RhD-specific IgG from purification B and subsequently with a secondary APC-coupled anti-human IgG antibody.<br>These cells are not shown in the overlay histogram in Supplementary Figure 1 in the main text. However, the staining looks like the staining from the file before.</p> <p>2_µg_per_ml_total_IgG_from_Rhophylac.003<br>FACS analysis with RhD-positive erythrocytes to verify the enrichment of RhD-specific IgGs from total IgG of Rhophylac.<br>The cells in this file are stained with 2 µg/ml total IgG from Rhophylac and subsequently with a secondary APC-coupled anti-human IgG antibody. The cells were gated on the main erythrocyte population in a FSC/SSC blot to exclude fragments and aggregates, analyzed in an anti-human IgG (APC) histogram and shown in the overlay histogram in Supplementary Figure 1 in the main text.</p> <p> </p

Presentation_1_Sialylated Autoantigen-Reactive IgG Antibodies Attenuate Disease Development in Autoimmune Mouse Models of Lupus Nephritis and Rheumatoid Arthritis.PDF

<p>Pro- and anti-inflammatory effector functions of IgG antibodies (Abs) depend on their subclass and Fc glycosylation pattern. Accumulation of non-galactosylated (agalactosylated; G0) IgG Abs in the serum of rheumatoid arthritis and systemic lupus erythematosus (SLE) patients reflects severity of the diseases. In contrast, sialylated IgG Abs are responsible for anti-inflammatory effects of the intravenous immunoglobulin (pooled human serum IgG from healthy donors), administered in high doses (2 g/kg) to treat autoimmune patients. However, whether low amounts of sialylated autoantigen-reactive IgG Abs can also inhibit autoimmune diseases is hardly investigated. Here, we explore whether sialylated autoantigen-reactive IgG Abs can inhibit autoimmune pathology in different mouse models. We found that sialylated IgG auto-Abs fail to induce inflammation and lupus nephritis in a B cell receptor (BCR) transgenic lupus model, but instead are associated with lower frequencies of pathogenic Th1, Th17 and B cell responses. In accordance, the transfer of small amounts of immune complexes containing sialylated IgG Abs was sufficient to attenuate the development of nephritis. We further showed that administration of sialylated collagen type II (Col II)-specific IgG Abs attenuated the disease symptoms in a model of Col II-induced arthritis and reduced pathogenic Th17 cell and autoantigen-specific IgG Ab responses. We conclude that sialylated autoantigen-specific IgG Abs may represent a promising tool for treating pathogenic T and B cell immune responses in autoimmune diseases.</p