Cd66b percp cy5.5 g10f5

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CD66b-PerCP-Cy5.5 (G10F5) is a fluorescently-labeled antibody that binds to the CD66b cell surface antigen. CD66b is expressed on the surface of human granulocytes, including neutrophils, eosinophils, and basophils. This antibody conjugate can be used to identify and distinguish these cell populations in flow cytometry applications.

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3 protocols using cd66b percp cy5.5 g10f5

Multiparameter Flow Cytometry of Hematopoietic Cells

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Single cell suspensions obtained from colony forming assays or hematopoietic organs from mice were surface stained with monoclonal antibodies. Antibodies used for analysis of human immature stem and progenitor cells from colony forming assays: CD34 PE-Cy7(581), CD38 APC (HIT2), CD10 PE/PeCy5/APC (HI10a), CD45RA PerCP-Cy5.5 (HI100), CD90 APC-Cy7 (5E10) (Biolegend). Antibodies for erythroid cells from colony assays and HEMA: CD117 PE-Cy7 (104D2), CD71 APC/APC-Cy7 (CY1G4) (Biolegend), CD235a BV421 (HIR2) (BD biosciences). Antibodies for myeloid cells from colony forming assays: CD34 PE-Cy7 (581), CD33 PE/PerCP-Cy5.5 (WM53), CD14 APC (M5E2), CD115 BV421 (g-4D2-1E4), CD15 PeCy5 (W6D3), CD66b PerCP-Cy5.5 (G10F5) (Biolegend). Antibodies for megakaryocyte populations: CD41 PerCP-Cy5.5 (HIP8), CD61 APC (VI-PL2) (Biolegend). Antibodies for overall human and human myeloid cells in xenograft assay: CD45 Biotin (HI30), CD34 PE-Cy7/APC-Cy7 (581), CD33 PE/PerCP-Cy5.5 (WM53), CD15 PeCy5 (W6D3). Antibodies for human B cells in xenograft assay: CD19 PE-Cy7 (HIB19), IgM APC-Cy7 (MHM-88), CD10 APC (HI10a) (Biolegend). Antibody to detect murine cells in xenograft assay: CD45 PE-Cy7/PerCP-Cy5.5 (30-F11) (Biolegend). Streptavidin APC/V450 (Biolegend) were used as secondary antibodies. BD LSRFortessa was used for flow cytometry. Analyses were performed using FlowJo.

Afreen S., Bohler S., Müller A., Demmerath E.M., Weiss J.M., Jutzi J.S., Schachtrup K., Kunze M, & Erlacher M. (2020). BCL-XL expression is essential for human erythropoiesis and engraftment of hematopoietic stem cells. Cell Death & Disease, 11(1), 8.

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Polychromatic Flow Cytometry Analysis

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The following fluorochrome-conjugated Abs were used for polychromatic flow cytometry analysis: CD3-Pacific-blue (UCHT1), CD4-Alexa-Fluor 700 (RPA-T4), CCR4-PE-Cy7 (1G1), CCR6-PE (11A9), CCR7-PE-Cy7 (3D12), CXCR3-PE-Cy5 (1C6) (BD Biosciences, San Diego, CA), CCR7-FITC (150503) (R&D Systems, Minneapolis, MN), CD45RA-APC-eFluor780 (HI100) (eBioscience, San Diego, CA), CD3-Alexa700 (UCHT1), CD326-BV650 (9C4), CD8-PerCP-Cy5.5 (RPA-T8), CD19-PerCP-Cy5.5 (HIB19), CD66b-PerCP-Cy5.5 (G10F5), HLADR-BV785 (L243; BioLegend), CD4-FITC (SFCI12T4D11) and HIV-p24-RD1 (FH190-1-1; Beckman Coulter, Fullerton, CA). Live/Dead Fixable Aqua Dead Cell Stain Kit (Vivid; Life Technologies, Burlington, ON) was used to exclude dead cells. Cells were analyzed by FACS using the BD-LSRII cytometer and BD-Diva (BD Biosciences, San Jose, CA), and FlowJo (^©Tree Star, Inc., Ashland, OR) softwares. All Abs were titrated for an optimal noise/signal ratio and Ab cocktails were validated by comparing single to multiple staining. Positive gates were placed based on fluorescence minus one (FMO) controls [41 (link), 69 ]

Gosselin A., Wiche Salinas T.R., Planas D., Wacleche V.S., Zhang Y., Fromentin R., Chomont N., Cohen É.A., Shacklett B., Mehraj V., Ghali M.P., Routy J.P, & Ancuta P. (2016). HIV persists in CCR6+CD4+ T cells from colon and blood during antiretroviral therapy. AIDS (London, England), 31(1), 35-48.

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Polychromatic Flow Cytometry of Immune Markers

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The following fluorochrome-conjugated antibodies (Abs) were used for polychromatic flow cytometry analysis: CD3-Pacific-blue (UCHT1), CD4-Alexa-Fluor 700 (RPA-T4), CCR4-PE-Cy7 (1G1), CCR6-PE (11A9), CCR7-PE-Cy7 (3D12), CXCR3-PE-Cy5 (1C6) (BD Biosciences, San Diego, California, USA), CCR7-FITC (150503) (R&D Systems, Minneapolis, Minnesota, USA), CD45RA-APC-eFluor780 (HI100) (eBioscience, San Diego, California, USA), CD3-Alexa700 (UCHT1), CD326-BV650 (9C4), CD8-PerCP-Cy5.5 (RPA-T8), CD19-PerCP-Cy5.5 (HIB19), CD66b-PerCP-Cy5.5 (G10F5), human leukocyte antigen - antigen D related (HLA-DR)-BV785 (L243; Biolegend ,San Diego, California USA), CD4-FITC (SFCI12T4D11), and HIV-p24-RD1 (FH190–1–1); (Beckman Coulter, Fullerton, California, USA). Live/Dead Fixable Aqua Dead Cell Stain Kit (Vivid; Life Technologies, Burlington, Ontario, Canada) was used to exclude dead cells. Cells were analyzed by fluorescence-activated cell sorting (FACS) using the BD-LSRII cytometer and BD-Diva (BD Biosciences) and FlowJo (Tree Star, Inc., Ashland, Oregon, USA) softwares. All Abs were titrated for an optimal noise/signal ratio and Ab cocktails were validated by comparing single with multiple staining. Positive gates were placed based on fluorescence minus one controls [41 (link),69 ].

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