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Anti hla dr percp cy5

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Anti-HLA-DR-PerCP-CY5.5 is a fluorescently-labeled antibody that binds to the HLA-DR antigen, a surface marker expressed on human antigen-presenting cells. The PerCP-CY5.5 fluorescent dye is conjugated to the antibody, allowing for detection and quantification of HLA-DR-positive cells using flow cytometry.

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Lab products found in correlation

Facsdiva software, by BD (3 mentions) Anti cd14 pacific blue, by BD (2 mentions) Lsrfortessa, by BD (2 mentions) Anti cd14 apc, by BD (2 mentions) Pe cf594 anti cd163, by BD (2 mentions) Anti cd80 bb515, by BD (2 mentions) Anti cd3 apc h7, by BD (1 mentions) Anti cd4 bv605, by BD (1 mentions) Anti p24 fitc, by Beckman Coulter (1 mentions) Anti cd19 pacific blue, by BioLegend (1 mentions) Anti ccr5 apc, by BD (1 mentions) Anti cd38 pecy7, by BioLegend (1 mentions) Anti cd11b apc, by BD (1 mentions) Anti cd11b pe cy7, by BD (1 mentions) Anti hla dr fitc, by BD (1 mentions) Alexa fluor 555 conjugated anti rabbit igg secondary antibody, by Thermo Fisher Scientific (1 mentions) Ficoll, by GE Healthcare (1 mentions) Anti cd86 bb515, by BD (1 mentions) Anti cd20 apc, by BD (1 mentions) Anti cd16 pe, by BD (1 mentions)

Spelling variants of this product

HLA-DR (247 citations) PerCP-Cy5.5 (120 citations) Anti-HLA-DR (55 citations) HLA-DR-PerCP (43 citations) HLA-DR-PerCP-Cy5.5 (29 citations) Anti-HLA-DR-PerCP (19 citations) Anti-HLA-DR- PerCP-Cy5.5 (clone G46-6, (3 citations) PerCP-Cy5.5-conjugated anti-human HLA-DR (G46-6; (3 citations) PerCP-Cy5.5-conjugated anti-human HLA-DR (3 citations)

7 protocols using anti hla dr percp cy5

1

Cellular Activation Measurement for HIV

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Cellular activation was assessed by measurement of HLA-DR and CD38, similar to previous studies (12 (link), 36 (link), 37 (link)). Staining for flow cytometry was performed both extracellularly and intracellularly. The extracellular staining cocktail consisted of LIVE/DEAD Amcyan fixable dye (Thermo Fisher Scientific, Waltham, MA, USA), anti-CD3-APC-H7, anti-CD4-BV605, anti-CD8-BV655, anti-CD14-Pacific blue (all from BD Biosciences, Franklin Lakes, NJ, USA), and anti-CD19-pacific blue (Biolegend, San Diego, CA, USA). The intracellular staining cocktail consisted of anti-CCR5-APC, anti-HLA-DR-PerCP-CY5.5 (all from BD Biosciences, Franklin Lakes, NJ, USA), anti-CD38-PE-CY7 (Biolegend, San Diego, CA, USA) and anti-p24-FITC (Beckman Coulter, Brea, CA, USA). PBMCs were collected at two time-points: day 3 (48 h post stimulation and prior to HIV infection) and day 5 (48 h post infection).
Cromarty R., Sigal A., Liebenberg L.J., McKinnon L.R., Abdool Karim S.S., Passmore J.A, & Archary D. (2019). Diminished HIV Infection of Target CD4+ T Cells in a Toll-Like Receptor 4 Stimulated in vitro Model. Frontiers in Immunology, 10, 1705.
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2

Quantification of MDSC and IL-35/IL-27 Receptors

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Peripheral blood mononuclear cells (PBMCs) were isolated from freshly obtained blood (AIH or HC) by Ficoll (GE Healthcare Life Sciences, Piscataway, NJ, USA) gradient centrifugation. Then PBMCs were stained with anti-HLA-DR-PercpCy5.5, anti-CD11b-PE-Cy7, and anti-CD33-BV421 (BD Pharmingen). Moreover, anti-gp130-APC (Biolegend) and anti-IL27RA-Alexa Flour488 (R&D systems) were used and then anti-IL12Rβ2 primary antibody was combined with Alexa Fluor 555-conjugated anti-rabbit IgG secondary antibody (Invitrogen/Life Technologies, UK). The percentages and numbers of HLA-DR−/lowCD33+CD11b+ cells (MDSCs) and expression abundance of IL-35 and IL-27 receptors on MDSC were quantified and analyzed by flow cytometry (BD LSRFortessa). For some in vitro experiments, anti-HLA-DR-FITC, and anti-CD11b-APC (BD Pharmingen) were utilized as well. 500,000 events were recorded and analyzed using the FlowJo software version 7.2.2 (Tree Star, Ashland, OR, USA).
Lian M., Zhang J., Zhao L., Chen X., Peng Y., Wang Q., Chen S, & Ma X. (2019). Interleukin-35 Regulates Immune Microenvironment of Autoimmune Hepatitis Through Inducing the Expansion of Myeloid-Derived Suppressor Cells. Frontiers in Immunology, 10, 2577.
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3

Phenotypic Analysis of Polarized HMDMs

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HMDMs were isolated, differentiated, and polarized as described above for 24 h. Polarized cells were treated with 50 nM thioA for 7 h. Cells were detached using accutase solution (Sigma-Aldrich #A6964), washed, and resuspended in FACS wash (PBS, 2.5% FCS, 0.1% sodium azide). Cells were blocked in human Fc Block (BD, #564220) for 15 min, and stained for 30 min on ice with anti-CD14-APC (BD #555399), anti-CD163-PE-CF594 (BD #562670), anti-CD80-BB515 (BD #565008), and anti-HLA-DR-PerCP-CY5.5 (BD #560652) antibodies. After washing, stained cells were resuspended in 1% paraformaldehyde in PBS prior to flow cytometric analysis on a BD LSRFortessa. Data were analyzed using BD FACSDiva software (BD Biosciences, San Jose, CA, USA). Median fluorescence intensity of singlet cells was used to quantify surface marker expression.
Dahlem C., Siow W.X., Lopatniuk M., Tse W.K., Kessler S.M., Kirsch S.H., Hoppstädter J., Vollmar A.M., Müller R., Luzhetskyy A., Bartel K, & Kiemer A.K. (2020). Thioholgamide A, a New Anti-Proliferative Anti-Tumor Agent, Modulates Macrophage Polarization and Metabolism. Cancers, 12(5), 1288.
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4

Phenotypic Analysis of Polarized Macrophages

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Flow cytometric analyses were performed as previously described [36] . Briefly, MDMs were differentiated for five to six days and polarized as described above. 2.5-5 × 105 cells were resuspended in FACSwash (PBS, 2.5% FCS, 0.1% sodium azide), blocked in human Fc Block solution (#564220, BD Biosciences, San Jose, CA, USA) for 15 minutes, and stained for 30 minutes on ice with anti-CD14-APC (#555399, BD Biosciences), anti-CD163-PE-CF594 (#562670, BD Biosciences), anti-CD80-BB515 (#565008, BD Biosciences) or anti-CD86-BB515 (#564544, BD Biosciences), and anti-HLA-DR-PerCP-Cy5.5 (#560652, BD Biosciences) antibodies. After washing, stained cells were resuspended in 1% paraformaldehyde in PBS prior to flow cytometric analysis on a BD LSRFortessa. Data were analyzed using BD FACSDiva software (BD Biosciences). Median fluorescence intensities of singlet cells were used to quantify surface marker expression.
Hoppstädter J., Dembek A., Höring M., Schymik H.S., Dahlem C., Sultan A., Wirth N., Al-Fityan S., Diesel B., Gasparoni G., Walter J., Helms V., Huwer H., Simon M., Liebisch G., Schulz M.H, & Kiemer A.K. (2021). Dysregulation of cholesterol homeostasis in human lung cancer tissue and tumour-associated macrophages. EBioMedicine, 72, 103578.
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5

Isolation of Monocyte Subsets

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Prior to infection, at 26 dpi, and at necropsy, mononuclear cells were isolated from 10 ml EDTA-treated whole blood (n = 4 animals: CM07, DB79, FD05, FB92) by density gradient centrifugation over Ficoll-Hypaque (GE Healthcare, Chicago, IL). Buffy coats were collected and washed in calcium- and magnesium-free PBS and stained for 15 min with anti-CD14-PacificBlue, anti-CD16-PE, anti-HLA-DR-PerCP-Cy-5.5, anti-CD3-FITC, and anti-CD20-APC (BD Biosciences) in PBS containing 2% FBS. Monocytes were isolated by forward and side scatter characteristics and CD14 antigen expression. HLA-DR+CD3-CD20–cells were selected to exclude B- and T-cells (Figure 2A). Nonoverlapping populations of classical, intermediate, and nonclassical monocytes were isolated based on expression of CD14 and CD16 on a BD FACS ARIA (BD Biosciences, San Jose, CA). FACS-isolated monocytes were washed in PBS to remove residual FACS buffer and vacuum pelleted to preserve RNA integrity.
Nowlin B.T., Wang J., Schafer J.L., Autissier P., Burdo T.H, & Williams K.C. (2017). Monocyte subsets exhibit transcriptional plasticity and a shared response to interferon in SIV‐infected rhesus macaques. Journal of Leukocyte Biology, 103(1), 141-155.
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6

Multicolor Flow Cytometry for MDSC Analysis

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For MDSCs characterization the following mAbs were used: anti-CD14/Pe-Cy7, anti-CD11b/APC-Cy7, anti-HLA-DR/PerCP-Cy5.5, anti-Lineage cocktail 1/FITC (BD Biosciences), anti-CD33/PE, anti-CD15/APC, anti-TGF-β/FITC (BioLegend) and anti-ARG1/AlexaFluor488, anti-IDO/AlexaFluor488, anti-IL-10/FITC (R&D Systems). For intracellular staining, the cells were fixed/permeabilized using a Cytofix/Cytoperm Fixation/Permeabilisation Kit (BD Biosciences, USA). Nonspecific staining was prevented by using FcR Blocker (Miltenyi Biotec, Bergisch Gladbach, Germany). Cells were analyzed with BD FACSCanto flow cytometer using FACS DIVA software (BD Biosciences, USA) and FCS express (De Novo Software, Los Angeles, CA). The gating strategy for MDSCs is shown in Figure 1A.
Okła K., Czerwonka A., Wawruszak A., Bobiński M., Bilska M., Tarkowski R., Bednarek W., Wertel I, & Kotarski J. (2019). Clinical Relevance and Immunosuppressive Pattern of Circulating and Infiltrating Subsets of Myeloid-Derived Suppressor Cells (MDSCs) in Epithelial Ovarian Cancer. Frontiers in Immunology, 10, 691.
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7

Isolation of Human Basophils by Flow Cytometry

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For human basophil sorting, cell suspensions were incubated at 4ºC with Fc-blocking reagent (Miltenyi Biotec) or blocking 2.4G2 mAb to FcγRIII/II (BD PharMingen) and then stained for 30 min with the following antibody combination: anti-CD45 AF700 (clone: HI30), anti-FcεRI FITC (clone: CRA1), antiCD123 PE (clone: 6H6), anti-HLA-DR PerCpCy5.5 (clone: L243), anti-CD19 eFluor450 (clone: HIB19), anti-IgD PE-Cy7 (clone: IA6–2). Basophils were sorted with a FACSAria II (BD Biosciences) after exclusion of dead cells through DAPI staining. The purity of cells sorted this way was consistently >95%.
Shan M., Carrillo J., Yeste A., Gutzeit C., Garzón D.S., Walland C., Pybus M., Grasset E.K., Yeiser A.J., Matthews D.B., van de Veen W., Comerma L., He B., Boonpiyathad T., Lee H., Blanco J., Osborne L.C., Siracusa M.C., Akdis M., Artis D., Mehandru S., Sampson H.A., Berin M.C., Chen K, & Cerutti A. (2018). Secreted IgD Amplifies Humoral T Helper-2 Responses by Binding Basophils Via Galectin-9 and CD44. Immunity, 49(4), 709-724.e8.
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