For class switching assays, cell suspensions were stained with fluorochrome-conjugated anti-IgG1, anti-IgG3 (BD-Biosciences), anti-IgG2b (BioLegend), or anti-IgA (Southern Biotech). Samples were acquired on a LSRFortessa cell analyzer (BD-Biosciences). Analysis of B cell development and differentiation was performed using anti-CD21/CD35-FITC, anti-IgD-FITC, anti-IgM-PE, anti-IgM-FITC, anti-CD43-PE (BD-Biosciences) and anti-CD23-PE, anti-CD3-PE and anti-CD19-APC (BioLegend) antibodies. For cell proliferation analysis by cell tracking dye dilution, primary B cells were pulsed with 2 μM carboxyfluorescein succinimidyl ester (CFSE) (Invitrogen) or 5 μM CellTrace Violet (Thermofisher) for 10 min at 37C. CFSE/CellTrace covalently labels intracellular molecules, and each cell division halves the signal intensity. For cell cycle analysis, CH12 cells were collected, fixed, and permeabilized using Fixation/Permeabilization Solution (included in BrdU Flow Kit, BD-Biosciences) according to the manufacturer’s instructions. BrdU pulse and staining was performed by using BrdU Flow Kit (BD-Biosciences) according to the manufacturer’s instructions.
Anti igg2b
Manufactured by BioLegend
Sourced in United States
Anti-IgG2b is a lab equipment product that can be used to detect and measure the presence of IgG2b, a specific type of antibody, in various samples. It functions as a detection reagent for immunoassays, such as ELISA, and can be used for research purposes.
Automatically generated - may contain errors
Lab products found in correlation
Mouse recombinant il 4, by Merck Group (2 mentions)
Lipopolysaccharides (lps), by Thermo Fisher Scientific (2 mentions)
Lipopolysaccharides (lps), by Merck Group (2 mentions)
Tgf β, by Thermo Fisher Scientific (2 mentions)
Tgf β, by R&D Systems (2 mentions)
Anti igg3, by BD (2 mentions)
Recombinant murine il 5, by Thermo Fisher Scientific (2 mentions)
Celltrace violet, by Thermo Fisher Scientific (1 mentions)
Anti igm fitc, by BD (1 mentions)
Anti cd43 pe, by BD (1 mentions)
Anti igd fitc, by BD (1 mentions)
Anti igm pe, by BD (1 mentions)
Lsrfortessa cell analyzer, by BD (1 mentions)
Brdu flow kit, by BD (1 mentions)
Streptavidin conjugated horse radish peroxidase, by Thermo Fisher Scientific (1 mentions)
α cd40, by BioLegend (1 mentions)
Acd40, by BioLegend (1 mentions)
5 protocols using anti igg2b
1
Assaying B cell development and proliferation
2
ELISA Protocol for Chromatin-Binding Antibodies
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Chromatin was prepared as previously described [26 (link), 27 (link)]. Plates were coated with chromatin at 3 μg/ml overnight at 4 °C. The plates were then washed and incubated with blocking buffer for 2 h at room temperature (RT). The plates were washed again and then 1/500 diluted serum samples were added in duplicate and incubated for 2 h at RT. Biotin-labeled anti-IgG, anti-IgG1, anti-IgG2a, anti-IgG2b, and IgG3 (BioLegend) were used as capture antibodies, and streptavidin-conjugated horseradish peroxidase (HRP) (Thermo Scientific) was used as the detective antibody. Tretramethylbenzidine (TMB, 1×; eBioscience) was then added and incubated for 30 min at RT. Sulfuric acid (2 N) was used as a stop solution and optical density (OD) values were then measured at 450 nm and 570 nm.
Sera antichromatin and anti-dsDNA antibodies of SLE patients were measured with ELISA kits from Inova Diagnostics Company. The experiments were performed according to the manufacturer’s instructions.
Sera antichromatin and anti-dsDNA antibodies of SLE patients were measured with ELISA kits from Inova Diagnostics Company. The experiments were performed according to the manufacturer’s instructions.
3
Induction of Class-Switched Ig Isotypes
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CH12 cells were stimulated to undergo CSR to IgA by treatment with 1–5 µg/ml αCD40 (BioLegend), 5 ng/ml TGFβ (R&D Systems), and 5 ng/ml mouse recombinant IL-4 for 48 h. B lymphocytes were stimulated to undergo class switching with 5 µg/ml LPS (Sigma-Aldrich) and 5 ng/ml mouse recombinant IL-4 (Sigma-Aldrich) for CSR to IgG1; 5 µg/ml LPS only for CSR to IgG3; 5 µg/ml LPS, 10 ng/ml BAFF (PeproTech), and 2 ng/ml TGFβ for CSR to IgG2b; or 5 µg/ml LPS, 10 ng/ml BAFF, 2 ng/ml TGFβ, and 1.5 ng/ml recombinant murine IL-5 (PeproTech) for CSR to IgA. For class switching analysis, cell suspensions were stained with fluorochrome-conjugated anti-IgG1, anti-IgG3 (BD Biosciences), anti-IgG2b (BioLegend), or anti-IgA (Southern Biotech).
4
Multiparameter Immune Phenotyping
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Anti-IgG1, anti-IgG2a, and anti-IgG2b Abs were used as isotype controls and purchased from BioLegend (San Diego, CA, USA). Allophycocyanin (APC)-Cy7-anti-T cell receptor (TCR)-β (H57-597), PerCP5.5-anti-CD4 (GK1.5), BV785-anti-CD8α (53–6.7), phycoerythrin (PE)/cy7-anti-interferon-gamma (IFN-γ) (B27), BV 785-anti-CD11c (N418), Alexa Fluor 647-anti-CD40 (HM40-3), BV 605-anti-CD80 (16-10A1), PE/Cy7-anti-CD86 (GL-1), PerCP5.5-anti-MHC class I (AF6-88.5), and PE-anti-MHC class II (M5/114.15.2) were obtained from BioLegend.
5
Induction of Class-Switch Recombination
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CH12 cells were stimulated to undergo CSR to IgA by treatment with 1-5 µg/mL aCD40 (BioLegend), 5 ng/ml TGFb (R&D Systems) and 5 ng/ml of mouse recombinant IL-4 for 48 h. B lymphocytes were stimulated to undergo class switching with 5 µg/ml LPS (Sigma-Aldrich) and 5 ng/ml of mouse recombinant IL-4 (Sigma-Aldrich) for CSR to IgG1; 5 µg/ml LPS only for CSR to IgG3; 5 µg/ml LPS, 10 ng/ml BAFF (PeproTech) and 2 ng/ml TGFb for CSR to IgG2b; or 5 µg/ml LPS, 10 ng/ml BAFF, 2 ng/ml TGFb and 1.5 ng/ml recombinant murine IL-5 (PeproTech) for CSR to IgA. For class switching analysis, cell suspensions were stained with fluorochrome-conjugated anti-IgG1, anti-IgG3 (BD-Biosciences), anti-IgG2b (BioLegend), or anti-IgA (Southern Biotech).
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