Cd33 apc

Manufactured by Beckman Coulter

CD33-APC is a fluorochrome-conjugated antibody used for the detection and identification of cells expressing the CD33 antigen. CD33 is a transmembrane protein expressed on the surface of myeloid progenitor cells, monocytes, and granulocytes. The APC (Allophycocyanin) fluorochrome allows for the visualization and quantification of CD33-positive cells in flow cytometry analysis.

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Lab products found in correlation

Cd14 v450, by BD (3 mentions) Cd11b pe, by Beckman Coulter (3 mentions) Hla dr pc7, by Beckman Coulter (3 mentions) Cd15 fitc, by Beckman Coulter (3 mentions) Lsr 2 flow cytometer, by BD (2 mentions) Facscanto, by BD (1 mentions) Anti cd11c apc clone b ly6, by BD (1 mentions) Cyan adp, by Agilent Technologies (1 mentions) Anti cd1a pe, by BD (1 mentions) Moflo sorter, by Beckman Coulter (1 mentions) Anti cd33 fitc, by Beckman Coulter (1 mentions) Anti cd1d pe, by BD (1 mentions) Facscelesta, by BD (1 mentions) 96 well u bottom plate, by Corning (1 mentions) Cd41 pe cy5, by Beckman Coulter (1 mentions)

5 protocols using cd33 apc

Quantifying Myeloid-Derived Suppressor Cells

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PBMC from each patient at the pre-treatment and post-Cycle 1 time points were analyzed for myeloid derived suppressor cells (MDSC) as previously described [25 (link)]. Specific antibodies included CD15-FITC (Beckman Coulter), CD33-APC (Beckman Coulter), HLA-DR-PC7 (Beckman Coulter), CD11b-PE (Beckman Coulter), and CD14-V450 (BD Biosciences). Single color staining was performed for compensation. All samples were run on a BD LSR II flow cytometer and were subsequently analyzed with FlowJo software (TreeStar). MDSC were defined as cells positive for CD33 and CD11b and lacking HLA-DR with subsets expressing CD15 or CD14 representing granulocytic and monocytic MDSC, respectively, as discussed in figure legends.

Duggan M.C., Jochems C., Donahue R.N., Richards J., Karpa V., Foust E., Paul B., Brooks T., Tridandapani S., Olencki T., Pan X., Lesinski G.B., Schlom J, & Carson WE I.I.I. (2016). A phase I study of recombinant (r) Vaccinia-CEA(6D)-TRICOM and rFowlpox-CEA(6D)-TRICOM vaccines with GM-CSF and IFN-α-2b in patients with CEA expressing carcinomas. Cancer immunology, immunotherapy : CII, 65(11), 1353-1364.

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Flow cytometric analysis of CD1 expression

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CD1 expression was determined using mAbs anti–CD1a-PE (clone HI149; BD), anti–CD1b-PE (clone 0249; Santa Cruz Biotechnology, Inc.), anti–CD1c-PE (clone L161; Santa Cruz Biotechnology, Inc.), and anti–CD1d-PE (clone 42.1; BD). Leukemia blasts were identified using anti–CD19-FITC or CD19-PerCPCy5.5 (both clone HIB19) and anti–CD14-FITC (clone HCD14; all from BioLegend); anti–CD3-FITC (clone UCHT1), anti–CD45-APC (clone HI30), and anti–CD117-PerCpCy5.5 (clone 2B8; all from BD); anti–CD7-PC5 (clone 8H8.1), anti–CD10-PC5 (clone ALB1), anti–CD33-FITC or CD33-PC5 or anti–CD33-APC (all clone D3HL60-251), anti–CD34-FITC or CD34-PC5 or anti–CD34-APC (all clone 581), and anti–CD45-PC7 (clone J33; all from Beckman Coulter). cDCs were sorted using anti-CD64 FITC (clone 22; Beckman Coulter) and anti–CD11c-APC (clone B-ly6; BD). Samples were acquired on CyAn ADP (Dako) or FACSCanto (BD) flow cytometers. Cell sorting experiments were performed using a MoFlo sorter (Beckman Coulter). Dead cells were excluded on the basis of propidium iodide and DAPI staining. All data were analyzed using FlowJo software (Tree Star). Relative fluorescent intensity (RFI) of CD1c expression on leukemia cells was calculated dividing the mean CD1c fluorescence intensity by the mean fluorescence intensity of the corresponding isotype control mAb.

Lepore M., de Lalla C., Gundimeda S.R., Gsellinger H., Consonni M., Garavaglia C., Sansano S., Piccolo F., Scelfo A., Häussinger D., Montagna D., Locatelli F., Bonini C., Bondanza A., Forcina A., Li Z., Ni G., Ciceri F., Jenö P., Xia C., Mori L., Dellabona P., Casorati G, & De Libero G. (2014). A novel self-lipid antigen targets human T cells against CD1c+ leukemias. The Journal of Experimental Medicine, 211(7), 1363-1377.

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Multicolor Flow Cytometry Immunophenotyping

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Cells from the RF and BM of transplanted mice were stained in 96-well U-bottom plates (Corning, 351177) for 45 min at 4 °C. The following antibodies were used (all from BD Biosciences, unless stated otherwise): CD45 APC-Cy7 (1:100, 348795560566), CD45 A700 (1:100, 560566), CD33 APC (1:100, 340474), and CD19 V450 (1:100, 560353), CD41 PE-Cy5 (1:200, Beckman Coulter, 6607116), GlyA PE (1:100, Beckman Coulter, IM2211U), and CD3 FITC (1:100, 349201). For NSGW41 mice at 12 weeks, CD3 FITC staining was omitted. Cells were analyzed on the FACSCelesta (BD Biosciences).

Wagenblast E., Azkanaz M., Smith S.A., Shakib L., McLeod J.L., Krivdova G., Araújo J., Shultz L.D., Gan O.I., Dick J.E, & Lechman E.R. (2019). Functional profiling of single CRISPR/Cas9-edited human long-term hematopoietic stem cells. Nature Communications, 10, 4730.

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MDSC Phenotyping in PBMC

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PBMC were analyzed for the presence of MDSC as previously described [21 (link)]. MDSC were defined as cells positive for CD33, CD11b and lacking HLA-DR with subsets expressing CD15 or CD14 representing granulocytic and monocytic MDSC, respectively (Fig. 1). Notably, the current method for phenotyping M-MDSC (CD33+/HLADR-/CD14+/CD11b+) compares favorably to methods employed by other investigators (e.g., CD14+/HLADRlow/−) with respect to the percentages of M-MDSC obtained [22 (link)]. Specific antibodies included CD15-FITC, CD33-APC, HLA-DR-PC7, CD11b-PE (all Beckman Coulter), and CD14-V450 (BD Biosciences). All samples were run on a BD LSR-II flow cytometer and data was analyzed with FlowJo software (Tree Star, Inc.).

Wesolowski R., Duggan M.C., Stiff A., Markowitz J., Trikha P., Levine K.M., Schoenfield L., Abdel-Rasoul M., Layman R., Ramaswamy B., Macrae E.R., Lustberg M.B., Reinbolt R.E., Mrozek E., Byrd J.C., Caligiuri M.A., Mace T.A, & Carson WE I.I.I. (2017). Circulating myeloid derived suppressor cells increase in patients undergoing neo-adjuvant chemotherapy for breast cancer. Cancer immunology, immunotherapy : CII, 66(11), 1437-1447.

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Phenotypic Characterization of MDSC

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PBMC were analyzed for the presence of MDSC as previously described (12 (link)). Briefly, MDSC were defined as cells positive for CD33, CD11b and with low to no expression of HLA-DR with subsets expressing CD15 or CD14 representing granulocytic and monocytic MDSC, respectively (Figure 1). Specific antibodies included CD15-FITC, CD33-APC, HLA-DR-PC7, CD11b-PE (all Beckman Coulter), and CD14-V450 (BD Biosciences). All samples were analyzed on a BD LSR II flow cytometer and the data were subsequently analyzed with FlowJo software. Cells were then categorized into specific subsets: early MDSC (CD14-/CD15-), monocytic MDSC (CD14+/CD15+), granulocytic MDSC (CD14-/CD15+). Notably, there is a sizeable population within the gated cells that were double positive (CD14+/CD15+) that are not traditionally described as MDSC. As such, these cells were analyzed separately.

Sun S.H., Benner B., Savardekar H., Lapurga G., Good L., Abood D., Nagle E., Duggan M., Stiff A., DiVincenzo M.J., Suarez-Kelly L.P., Campbell A., Yu L., Wesolowski R., Howard H., Shah H., Kendra K, & Carson W.E. (2021). Effect of Immune Checkpoint Blockade on Myeloid-Derived Suppressor Cell Populations in Patients With Melanoma. Frontiers in Immunology, 12, 740890.

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