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Anti cd16 32 antibody 2.4g2

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The Anti-CD16/32 antibody (2.4G2) is a laboratory reagent used in immunological research. It is a rat monoclonal antibody that binds to the mouse CD16 and CD32 proteins, which are Fc receptors expressed on various immune cells. This antibody can be used to block Fc receptor-mediated functions in cell-based assays.

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Lab products found in correlation

Rbc lysis buffer, by Thermo Fisher Scientific (2 mentions) Fortessa, by BD (2 mentions) Anti gl7 gl7, by BioLegend (2 mentions) Non specific antibodies, by BD (2 mentions) Anti b220 ra3 6b2, by BioLegend (2 mentions) Facscanto 2, by BD (2 mentions) Monensin, by Merck Group (1 mentions) Mp5 20f3, by BD (1 mentions) Anti fas sa367h8, by BioLegend (1 mentions) Anti pd 1 j43, by BD (1 mentions) Anti cxcr5 2g8, by BD (1 mentions) Anti cd4 rm4 5, by BioLegend (1 mentions) Anti cd45 30 f11, by BioLegend (1 mentions) Anti siglecf e50 2440, by BD (1 mentions) Anti cd11b m1 70, by BioLegend (1 mentions) Zombie nir fixable viability kit, by BioLegend (1 mentions) Anti 1 a 1 e m5 114, by BioLegend (1 mentions) Anti cd4 gk1, by BioLegend (1 mentions) Anti cd138 281 2, by BioLegend (1 mentions) Anti ige rme 1, by BioLegend (1 mentions)

4 protocols using anti cd16 32 antibody 2.4g2

1

Flow Cytometric Analysis of Aorta Macrophages

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For blood analysis, whole blood was collected from the facial vein into EDTA treated tubes and 10 μL were used for cell count using a Nexcelom cellometer. Red blood cells (rbc) were lysed using RBC lysis buffer (ebioscience) for 5 minutes on the benchtop. Prior to staining, single cell suspensions were blocked with anti-CD16/32 antibody (2.4G2, Biolegend) diluted 1:1,000, then stained for specific antigens. Antibodies conjugated to Alexa488, Fitc, Alexa647, PE, PEcy7, APCcy7, PerCPefluor710, Pacblue, and Brilliant Violet605 were used at 1 μg/mL in 50 μL volume, including anti-CD45 (clone 30-F11), anti-CD11c (clone N418), anti-CD11b (M1/70), anti-CD64 (clone X54-5/7.1), anti-CD115 (clone AFS98), anti-Ly6G (clone 1A8), anti-Ly6C (clone HK1.4) from Biolegend. Cells were stained for 30 minutes on ice, then washed and assessed by flow cytometry using a BD x-20 or Fortessa instrument, using FACS Diva software v8. Analysis was performed using Flowjo v10 (Treestar). Gating strategies for sorting aorta macrophages were previously published in detail and blood gating strategy is outlined in Supplemental Figure 119 ,54 .
Williams J.W., Zaitsev K., Kim K.W., Ivanov S., Saunders B.T., Schrank P.R., Kim K., Elvington A., Kim S.H., Tucker C.G., Wohltmann M., Fife B.T., Epelman S., Artyomov M.N., Lavine K.J., Zinselmeyer B.H., Choi J.H, & Randolph G.J. (2020). Limited proliferation capacity of aorta intima resident macrophages requires monocyte recruitment for atherosclerotic plaque progression. Nature immunology, 21(10), 1194-1204.
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2

Flow Cytometric Analysis of Aorta Macrophages

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For blood analysis, whole blood was collected from the facial vein into EDTA treated tubes and 10 μL were used for cell count using a Nexcelom cellometer. Red blood cells (rbc) were lysed using RBC lysis buffer (ebioscience) for 5 minutes on the benchtop. Prior to staining, single cell suspensions were blocked with anti-CD16/32 antibody (2.4G2, Biolegend) diluted 1:1,000, then stained for specific antigens. Antibodies conjugated to Alexa488, Fitc, Alexa647, PE, PEcy7, APCcy7, PerCPefluor710, Pacblue, and Brilliant Violet605 were used at 1 μg/mL in 50 μL volume, including anti-CD45 (clone 30-F11), anti-CD11c (clone N418), anti-CD11b (M1/70), anti-CD64 (clone X54-5/7.1), anti-CD115 (clone AFS98), anti-Ly6G (clone 1A8), anti-Ly6C (clone HK1.4) from Biolegend. Cells were stained for 30 minutes on ice, then washed and assessed by flow cytometry using a BD x-20 or Fortessa instrument, using FACS Diva software v8. Analysis was performed using Flowjo v10 (Treestar). Gating strategies for sorting aorta macrophages were previously published in detail and blood gating strategy is outlined in Supplemental Figure 119 ,54 .
Williams J.W., Zaitsev K., Kim K.W., Ivanov S., Saunders B.T., Schrank P.R., Kim K., Elvington A., Kim S.H., Tucker C.G., Wohltmann M., Fife B.T., Epelman S., Artyomov M.N., Lavine K.J., Zinselmeyer B.H., Choi J.H, & Randolph G.J. (2020). Limited proliferation capacity of aorta intima resident macrophages requires monocyte recruitment for atherosclerotic plaque progression. Nature immunology, 21(10), 1194-1204.
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3

Flow Cytometry Analysis of Immune Markers

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Cells were stained and analysed on the FACSCanto II (Becton Dickinson, San Jose, California, USA) by using FlowJo software (TreeStar, Ashland, Oregon, USA). The following antibodies were used: anti-CD4 (RM4-5; BioLegend, San Diego, California, USA), anti-CXCR5 (2G8, BD Biosciences, San Jose, California, USA), anti-PD-1 (J43, BD Biosciences), anti-B220 (RA3-6B2, BioLegend), anti-Fas (SA367H8, BioLegend) and anti-GL7 (GL7, BioLegend). Before staining, Fc receptors were blocked with anti-CD16/32 antibody (2.4G2, BioLegend). Negative controls consisted of isotype-matched, directly conjugated, non-specific antibodies (BD Biosciences). In some experiments, single-cell suspensions of isolated glomeruli were incubated with 2 µM monensin (Sigma-Aldrich) in RPMI 1640 medium for 5 hours, and intracellular staining of SOCS3 and IL-6 and staining of podocin (NPHS2) were performed using anti-SOCS3 antibody (OTI3D3; Abcam, Cambridge, Massachusetts, USA), anti-IL-6 antibody (MP5-20F3; BD Biosciences) and anti-NPHS2 antibody (JB51-33; Hubio, Toronto, Canada).
Fukuta M., Suzuki K., Kojima S., Yabe Y., Suzuki K., Iida K., Yamada H., Makino S., Iwata A., Tanaka S., Iwamoto T., Suto A., Nakagomi D., Wakashin H., Maezawa Y., Maezawa Y., Takemoto M., Asanuma K, & Nakajima H. (2021). Suppressor of cytokine signalling 3 (SOCS3) expressed in podocytes attenuates glomerulonephritis and suppresses autoantibody production in an imiquimod-induced lupus model. Lupus Science & Medicine, 8(1), e000426.
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4

Multiparametric Flow Cytometry Analysis

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Single cell suspensions obtained from the intestinal mucosa or MLNs were stained and analyzed on FACSCanto II (Becton Dickinson, Franklin Lakes, NJ, USA) using FlowJo software v10.8.2 (TreeStar, Ashland, OR, USA). The following antibodies were used: anti-B220 (RA3-6B2; BioLegend), anti-CD38 (90; BioLegend), anti-CD45 (30-F11; BioLegend), anti-GL7 (GL7; BioLegend), anti-CD138 (281-2; BioLegend), anti-CD4 (GK1.5; BioLegend), anti-CXCR5 (L138D7, BioLegend), anti-PD-1 (29F.1A12, BioLegend), anti-Siglec-F (E50-2440, BD Biosciences, Franklin Lakes, NJ, USA), anti-CD11c (N418, BioLegend), anti-I-A/I-E (M5/114.15.2, BioLegend), anti-IgE (RME-1, BioLegend), anti-CD103 (W19396D, BioLegend), anti-CD11b (M1/70, BioLegend), anti-IL-7Rα (A7R34, BioLegend), anti-CCR6 (29-2L17, BioLegend), anti-TCRβ (H57-597, BioLegend), and anti-TCRγδ (UC7-13D5, BioLegend). Dead cells were gated out using a Zombie NIR Fixable Viability Kit (BioLegend). Before staining, Fc receptors were blocked with an anti-CD16/32 antibody (2.4G2, BioLegend). Negative controls consisted of isotype-matched, directly conjugated, nonspecific antibodies (BD Biosciences). Intracellular staining was performed using anti-Foxp3 antibody (FJK-16s; Thermo Fisher Scientific) and anti-RORγt antibody (Q31-378; BD Biosciences), as described previously [29 (link)].
Kurihara S., Suzuki K., Yokota M., Ito T., Hayashi Y., Kikuchi R., Kageyama T., Meguro K., Tanaka S., Iwata A., Goto Y., Suto A, & Nakajima H. (2024). Eosinophils Contribute to Oral Tolerance via Induction of RORγt-Positive Antigen-Presenting Cells and RORγt-Positive Regulatory T Cells. Biomolecules, 14(1), 89.
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