mouse anti human iga2 hrp  (SouthernBiotech)

Journal: European Journal of Immunology

Article Title: Anti‐SARS‐CoV‐2 Spike IgA2 Induces Inflammation by Human Macrophages

doi: 10.1002/eji.70068

Figure Lengend Snippet: IgA induces proinflammatory cytokine production by alveolar‐like macrophages. Macrophages were stimulated with immune complexes made from pooled serum (ps) IgA and TLR3 agonist Poly(I:C) and analyzed for their cytokine production ( A ). Cytokine concentrations are depicted on the left y ‐axis, and the fold increase of cytokine production upon double stimulation with psIgA and Poly(I:C) over single stimulation with Poly(I:C) is depicted on the right y ‐axis ( n = 14–19). After Preincubation with FcαRI blocking antibodies ( B ) or Syk inhibitor entospletinib ( C ), macrophages were stimulated with psIgA and Poly(I:C) and analyzed for their cytokine production ( n = 4–8). Symbols represent one individual donor, and bars indicate mean values. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 (two‐way ANOVA).

Article Snippet: Mouse‐anti‐human IgA2 clone A9604D2 , Southern Biotech , Cat. 9140‐05.

Techniques: Blocking Assay

Journal: European Journal of Immunology

Article Title: Anti‐SARS‐CoV‐2 Spike IgA2 Induces Inflammation by Human Macrophages

doi: 10.1002/eji.70068

Figure Lengend Snippet: Proinflammatory cytokine production is mostly induced by IgA2. Anti‐spike IgA1 and IgA2 antibody titers in sera of mild and severe COVID‐19 patients were measured by ELISAs ( A ). Each dot represents a single donor ( n = 5–25); statistical significance was determined using unpaired t‐tests. Upon stimulation with anti‐spike IgA1 or ‐IgA2 and Poly(I:C), alveolar‐like macrophages were analyzed for cytokine production ( B ). Each dot or line represents a donor ( n = 14), and the average of multiple donors is reflected by bars (two‐way ANOVA). The ratio of protein production by IgA2 over IgA1 is presented in ( C ). The cells were preincubated with FcαRI blocking antibodies ( D ) or Syk inhibitor entospletinib ( E ) to examine the dependency on both the receptor and the signaling molecule. The cytokine concentration of IL‐6 is depicted in the left panel ( n = 5–8, two‐way ANOVA), and other cytokines in Figure . Right panels show the cytokine reduction upon pretreatment as compared with double stimulation with poly(I:C) and IgA (paired t‐tests). Transcriptional regulation was analyzed by qPCR ( F ). mRNA levels were corrected to the housekeeping genes RACK1, HPRT1, and UBB. For every gene of interest, the fold increase compared with unstimulated cells, T = 0, is presented for a representative example. The area under the curve of these mRNA expression graphs is depicted in ( G ). Each dot or line represents a donor ( n = 4), and the average of multiple donors is reflected by bars. For each cytokine, the ratio of protein production over mRNA expression upon IgA2/1 stimulation, in the presence of Poly(I:C), was calculated ( H ). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Mouse‐anti‐human IgA2 clone A9604D2 , Southern Biotech , Cat. 9140‐05.

Techniques: Blocking Assay, Concentration Assay, Expressing

Journal: European Journal of Immunology

Article Title: Anti‐SARS‐CoV‐2 Spike IgA2 Induces Inflammation by Human Macrophages

doi: 10.1002/eji.70068

Figure Lengend Snippet: IgA2‐induced inflammation is more dependent on glycolysis and results in higher mitochondrial activity than IgA1. Alveolar‐like macrophages co‐stimulated with Poly(I:C) and anti‐spike IgA were tested for their dependency on glycolysis, the pentose phosphate pathway, and fatty acid metabolism. Cells preincubated with metabolic inhibitors were stimulated and analyzed for their production of IL‐6 ( A ) and other cytokines (Figure ). Each dot represents a single donor, and bars reflect the mean cytokine production ( n = 4–7), statistically tested by two‐way ANOVA. The cytokine‐reduction rates compared with co‐stimulation with Poly(I:C) and IgA are shown in ( B ). Data of single donors (dots) with mean + SD are analyzed by paired t‐tests ( n = 4–7). Samples were also examined for L‐lactate production ( C ) and mitochondrial SDH activity ( D ). Each dot or line represents a donor, and bars represent mean values per condition ( n = 9–10). Data were tested for statistical significance by two‐way ANOVA, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. SDH, succinate dehydrogenase.

Article Snippet: Mouse‐anti‐human IgA2 clone A9604D2 , Southern Biotech , Cat. 9140‐05.

Techniques: Activity Assay

Journal: European Journal of Immunology

Article Title: Anti‐SARS‐CoV‐2 Spike IgA2 Induces Inflammation by Human Macrophages

doi: 10.1002/eji.70068

Figure Lengend Snippet: Metabolic reprogramming by IgA2 is characterized by enhanced basal glycolysis and mitochondrial alterations. Extracellular flux assays (Seahorse) were performed on macrophages that were first stimulated with Poly(I:C) and anti‐spike IgA1 and anti‐spike IgA2 for 3 h. During the assay, glucose, oligomycin (OM), FCCP with pyruvate (FCCP + pyr), and rotenone with Antimycin A and 2‐DG (ROT + AA + 2‐DG) were injected at different timepoints. The proton efflux rate (PER) was measured over time ( A ), along with Basal ( B ) and Max PER, and the glycolytic reserve ( C ). The oxygen consumption rate (OCR) ( D ) with calculated proton leak ( E ), glucose‐reducing OCR ( F ), and glucose‐independent OCR ( G ). In panels A and D, each dot represents the mean + SD of four donors. Panels B, C, E–G present the calculated parameters of these donors (dots) and the mean values (bars) per condition, analyzed by one‐way ANOVA. These OCR and PER values are shown for basal ( H ), max ( I ), and combined in ( J ), presented as mean + SD. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Mouse‐anti‐human IgA2 clone A9604D2 , Southern Biotech , Cat. 9140‐05.

Techniques: Injection