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Ox40 apc

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The OX40-APC is a fluorochrome-conjugated antibody that binds to the OX40 receptor, which is expressed on activated T cells. This product is intended for use in flow cytometry applications to identify and characterize OX40-positive cells.

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2 protocols using ox40 apc

1

Splenic Immune Cell Phenotyping

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Single-cell suspensions were prepared from fresh spleen tissue and followed by lysis of red blood cell (RBC) using ACK lysis buffer. For surface staining, 1 × 106 splencytes per 100 μl were incubated for 30 min at 4 °C with the following fluorescently labeled conjugated mAbs: CD4-FITC (eBioscience, San Diego, CA), CXCR5-PE (eBioscience), CXCR5-APC (eBioscience), ICOS-PE-Cy5.5 (eBioscience), OX40-APC (eBioscience), CD40L-APC (eBioscience), PD-1-APC (eBioscience), CD19-APC (eBioscience), IL-21R-PE (BioLegend). After staining of surface markers, the cells were permeabilized with fixation/permeabilization solution (eBioscience) and subsequently intracellularly stained with Bcl-6-PE (BD Pharmingrn, San Jose, CA), the cells were then wash twice using permeabilization buffer, and eventually fixed with 4% paraformaldehyde in PBS. For APC-conjugated anti-IL-21(BD Pharmingrn), spleen cells were stimulated with PMA and Ionomycin for five hours and subsequently intracellularly stained, all procedures were in accordance with Bcl-6 staining method. In addition, Tfh (CD4+CXCR5+) subsets and B (CD19+) subsets were sorted with a FACS Aria cell sorter (BD Bioscience).
Wang Y., Lin C., Cao Y., Duan Z., Guan Z., Xu J., Zhu X.Q, & Xia C. (2017). Up-regulation of Interleukin-21 Contributes to Liver Pathology of Schistosomiasis by Driving GC Immune Responses and Activating HSCs in Mice. Scientific Reports, 7, 16682.
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2

Multiparameter Flow Cytometry Profiling

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Lymphocytes and myeloid cells were prepared as above. Cells were stained according to standard protocols with combinations of the following anti-mouse antibodies: OX40-APC (eBioscience; clone, OX86), CD4-FITC (eBioscience; clone, GK1.5), CD4-APC-Cy7 (BD Biosciences; clone, GK1.5), CD8–Pacific Orange (Life Technologies; clone, 5H10), CD8–Pacific Blue (BD Biosciences; clone, 53-6.7), TCRb–Pacific Blue (BD Biosciences; clone, H57-597), TCRb-PE (BD Biosciences; clone, H57-597), NK1.1-PE-Cy7 (eBioscience; clone, PK136), CD19–Alexa Fluor 700 (eBioscience; clone, ebio1D3), CD19-APCCy7 (BD Biosciences; clone, 1D3), CXCR5-biotin (BD Biosciences; clone, 2G8), OX40L-PE (eBioscience; clone, RM134L), F4/80-PE-Cy7 (eBioscience; clone, BM8), CD11b-PerCP-Cy5.5 (BD Biosciences; clone, M1/70), CD11c-APC (eBiosciences; clone, N418), major histocompatibility complex II–efluor 450(I-A/I-E) (eBioscience; clone, M5/114.15.2), Ly6g-PerCP-Cy5.5 (BD Biosciences; clone, 1A8), and Ly6c-AF700 (BD Biosciences, clone: AL-21). When applicable for biotinylated antibodies, cells were stained with secondary QDot 605–conjugated streptavidin (Life Technologies). Cells were analyzed using an LSR II flow cytometer (BD Biosciences) and FlowJo software (TreeStar) or sorted on an Aria III (BD Biosciences).
Publicover J., Gaggar A., Jespersen J.M., Halac U., Johnson A.J., Goodsell A., Avanesyan L., Nishimura S.L., Holdorf M., Mansfield K.G., Judge J.B., Koshti A., Croft M., Wakil A.E., Rosenthal P., Pai E., Cooper S, & Baron J.L. (2018). An OX40/OX40L interaction directs successful immunity to hepatitis B virus. Science translational medicine, 10(433), eaah5766.
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