Anti cd16 32 blocking antibody

Manufactured by BioLegend

Sourced in United States

Anti-CD16/32 blocking antibody is a laboratory reagent designed to block the binding of CD16 and CD32 receptors. It can be used to prevent non-specific Fc-receptor binding in various immunoassays and cell-based experiments.

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Lab products found in correlation

Lysis solution, by Merck Group (1 mentions) Precision count beads, by BioLegend (1 mentions) Rpmi 1640, by Thermo Fisher Scientific (1 mentions) Collagenase 1, by Thermo Fisher Scientific (1 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions) Lsr 2, by BD (1 mentions) V bottom plate, by Corning (1 mentions) Anti cd4, by Thermo Fisher Scientific (1 mentions) Fortessa, by BD (1 mentions) Anti cx3cr1 pe, by BioLegend (1 mentions) Zombie violet, by BioLegend (1 mentions) Anti cd45 fitc, by BioLegend (1 mentions) Fitc annexin 5 apoptosis detection kit with 7 aad, by BioLegend (1 mentions) Cd11b apc fire750, by BioLegend (1 mentions) Lsrfortessa, by BD (1 mentions) Facsaria 3 sorter, by BD (1 mentions) 40 μm cell strainer, by Merck Group (1 mentions) Live dead fixable near ir dead cell stain kit, by Thermo Fisher Scientific (1 mentions) Facsymphony a5 flow cytometer, by BD (1 mentions) Brilliant stain buffer, by BD (1 mentions)

Close spelling variants of this product

Anti-CD16/32 (115 citations) CD16/32 (68 citations) Anti-CD16/32 antibody (58 citations) CD16/32 antibody (44 citations) Anti‐mouse CD16/32 blocking antibody (2 citations) Anti-CD16/32 (Fcγ RII-Blocking antibody) (2 citations)

5 protocols using anti cd16 32 blocking antibody

Immunophenotyping Mouse Spleen and Kidney Cells

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The mouse spleens were firstly weighed, photographed, and then processed into single cell suspension through grinding gently. Trimmed of extra fat, kidneys were carefully minced and digested in 1640-RPMI (Gibco, Grand Island, NY) containing 10% fetal calf serum (FBS) (Gibco, Grand Island, NY) and 1 mg/mL collagenase I (Gibco, Grand Island, NY) for about 1 h at 37°C. Red blood cells (RBCs) were eliminated by lysis solution (Sigma-Aldrich, St. Louis, Missouri). Dead cells were identified using a fixable viability dyeing (Biolegend, San Diego, CA). An anti-CD16/32 blocking antibody (Biolegend, San Diego, CA) was used to reduce non-specific binding before staining with other antibodies including anti-mouse CD45 (I3/2.3), CD3e (145–2C11), CD19 (6D5), CD4 (GK1.5), CD8a (53–6.7), CD138 (281–2), F4/80 (BM8), CD11b (M1/70), CD11c (N418), MHC-II (25-9-17), CD86 (GL-1), CD40 (3/23), CD80 (16–10A1) and their isotype controls (all from Biolegend, San Diego, CA, USA). Precision count beads (Biolegend, San Diego, CA) were utilized to obtain actual cell numbers, in accordance with the manufacturer’s instructions. Stained cell suspensions were all analyzed by flow cytometry (BD LSR II, Franklin Lakes, New Jersey).

Jiang B., Zhang Y., Li Y., Chen Y., Sha S., Zhao L., Li D., Wen J., Lan J., Lou Y., Su H., Zhang C., Zhu J, & Tao J. (2022). A Tissue-Tended Mycophenolate-Modified Nanoparticle Alleviates Systemic Lupus Erythematosus in MRL/Lpr Mouse Model Mainly by Promoting Local M2-Like Macrophagocytes Polarization. International Journal of Nanomedicine, 17, 3251-3267.

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Quantifying Tumor-Infiltrating Myeloid Cells

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For flow cytometry analysis of infiltrative myeloid cells, C57/Bl6 were sacrificed 10 days after tumor implantation and 5 days after L-PRF implantation. Single cell suspension of peripheral organs was achieved by homogenizing tissue through a 70μm strainer before antibody staining. Isolation of lymphocytes and myeloid cells was performed homogenizing the tumor and patch, followed by a centrifugation of the tumors using 30%/70% discontinuous Percoll gradient centrifugation at 1000xg for 30 minutes at RT. The interface was collected, and all samples were then stained in 96 well V-bottom plates (Corning, Corning, NY). Cells were washed in PBS and stained with fixable viability dye Efluor 780 (Fisher, NJ). Cells were washed and stained in 2% FBS/PBS, incubated with anti-CD16/32 blocking antibody (Biolegend, San Diego, CA) 1:100 before staining with fluorescence conjugated antibodies. The fluorescence conjugated antibodies used were as follows: anti-CD4, anti-CD45 AmCyan, anti-CD8 BV605, anti-CD11b Pacific Blue, anti-CD11c APC, anti-CD45 PE-Cy7, and anti-CD25 Alexa-Fluor 700, and anti-Foxp3 PE (Fisher, Hampton, NH). All antibodies were purchased from Biolegend except when noted. Flow cytometry analysis was performed using the BD Fortessa (Becton Dickinson, Franklin Lakes, NJ).

Panek W.K., Pituch K.C., Miska J., Kim J.W., Rashidi A., Kanojia D., Lopez-Rosas A., Han Y., Yu D., Chang C.L., Kane J.R., Zhang P., Cordero A, & Lesniak M.S. (2018). Local Application of Autologous Platelet Rich Fibrin Patch (PRF-P) Suppresses Regulatory T cell Recruitment in a Murine Glioma Model. Molecular neurobiology, 56(7), 5032-5040.

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Isolation and Characterization of Mouse Microglia

Cited 2 times

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Mouse brains were homogenized as previously described^{10 (link),11 (link)} by gentle mechanical dissociation. Cells were then incubated in staining buffer on ice with anti-CD16/32 blocking antibody (BioLegend 101319, 1:500) for 15 min, and then with anti-mouse anti-CD11b-APC (BioLegend 101212, 1:100), anti-CD45-Alexa488 (BioLegend 103122, 1:100), and anti-CX3CR1-PE (BioLegend 149006, 1:100) for 25 min. Cell preparations for H3K27ac ChIP–seq, PLAC-seq and Hi-C were fixed with 1% formaldehyde for 10 min and quenched with 0.125 M glycine for 5 min after staining, and subsequently washed three times. Cells were washed once and filtered through a 40 μM cell strainer. Sorting was performed on a Sony MA900 or MoFlo Astrios EQ cell sorter. Microglia were defined as events that were DAPI negative, singlets and CD11b⁺CD45^lowCX3CR1⁺. Isolated microglia were then processed according to protocols for RNA-seq, ATAC-seq and ChIP–seq, Hi-C and PLAC-seq.

Fixsen B.R., Han C.Z., Zhou Y., Spann N.J., Saisan P., Shen Z., Balak C., Sakai M., Cobo I., Holtman I.R., Warden A.S., Ramirez G., Collier J.G., Pasillas M.P., Yu M., Hu R., Li B., Belhocine S., Gosselin D., Coufal N.G., Ren B, & Glass C.K. (2023). SALL1 enforces microglia-specific DNA binding and function of SMADs to establish microglia identity. Nature Immunology, 24(7), 1188-1199.

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Characterizing Wound-Infiltrating Immune Cells

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Cells isolated from skin wounds on day 3 and 6 post-injury were pre-incubated with anti-CD16/32 blocking antibody (Biolegend, clone S17011E). Surface antigens were labeled with anti-Ly6G-BV605 (clone 1A8), CD11b-APC/Fire750 (clone CBRM1/5), F4/80-PE (clone BM8), and Ly6C-Percp/cy5.5 (clone AL-21) antibodies (Biolegend, San Diego, CA, USA or BD Biosciences, San Jose, CA, USA). Next, cells were stained with FITC Annexin V Apoptosis Detection Kit with 7-AAD following manufacturer’s instructions (Biolegend). All samples were analyzed on either LSR Fortessa with HTS (BD Biosciences). Data were analyzed using FlowJo (FlowJo LLC).
For single cell RNA sequencing, single cells were pooled from skin wounds of two non-diabetic or two diabetic mice on day 6 post-injury as described above. Cells were first stained with Zombie Violet (Biolegend) to assess cell viability followed incubation with anti-CD16/32 blocking antibody. Next, cells were labeled with anti-CD45-FITC (Biolegend, clone 30-F11), Ly6G-PE/Dazzle^™ 594 (clone 1A8), and CD11b-APC/Fire750 (clone CBRM1/5). Target cells were sorted out as Zombie-CD45+CD11b+Ly6G- cells on BD FACSAria^™ III sorter (BD Biosciences).

Pang J., Maienschein-Cline M, & Koh T.J. (2021). Reduced apoptosis of monocytes and macrophages is associated with their persistence in wounds of diabetic mice. Cytokine, 142, 155516.

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Tumor Dissociation and Immune Cell Labeling

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Upon sacrifice, tumors were cut into small pieces and dissociated using PBS containing collagenase D (2 mg/ml; Roche) and DNase I (100 mg/ml; Roche) for 2 hours at 37°C. Single cells were separated from remaining tissue using a 40-μm cell strainer (Sigma-Aldrich). Single cells were pelleted and resuspended in fluorescence-activated cell sorting (FACS) buffer [PBS, 2 mM EDTA, and 2% FBS]. To block unspecific binding of antibodies, cells were incubated with anti-CD16/32 blocking antibody (BioLegend) in FACS buffer for 15 min on 4°C. Dead cells were excluded using the LIVE/DEAD Fixable Near-IR Dead Cell Stain Kit (Thermo Fisher Scientific). For immune cell labeling, single-cell suspensions were incubated with fluorophore-conjugated antibodies in Brilliant Stain Buffer (BD Biosciences) for 30 min at 4°C. Cell were washed and fixed with 1.5% paraformaldehyde (PFA). Sample acquisition and compensation was performed on a BD FACSymphony A5 flow cytometer.

Lehmann J., Caduff N., Krzywińska E., Stierli S., Salas-Bastos A., Loos B., Levesque M.P., Dummer R., Stockmann C., Münz C., Diener J, & Sommer L. (2023). Escape from NK cell tumor surveillance by NGFR-induced lipid remodeling in melanoma. Science Advances, 9(2), eadc8825.

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