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Cd33 apc

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CD33-APC is a lab equipment product that is used for the detection and analysis of CD33 antigen expression on cells. It contains anti-CD33 antibodies conjugated with the fluorescent dye allophycocyanin (APC).

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Lab products found in correlation

Cd19 pe, by BD (4 mentions) Cd34 pe cy7, by BD (4 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (3 mentions) Cd34 pe, by BioLegend (3 mentions) Fetal bovine serum (fbs), by Atlanta Biologicals (3 mentions) Qbsf 60, by Quality Biological (3 mentions) Cd10 pacific blue, by BioLegend (3 mentions) Cd19 pe cy5, by BioLegend (3 mentions) Cd3 pe cy7, by BD (3 mentions) Cd38 pe cy7, by BD (3 mentions) Cd14 percp cy5, by BD (3 mentions) Eclipse ts100 inverted microscope, by Nikon (2 mentions) Methocult h4434 classic, by STEMCELL (2 mentions) Cd3 apc, by BD (2 mentions) Cd235a fitc, by BD (2 mentions) Hla dr pe, by BD (2 mentions) Cd11b fitc, by BD (2 mentions) Lsrii flow cytometer, by BD (2 mentions) Cd45 fitc, by BD (2 mentions) Cd3 ecd, by Beckman Coulter (2 mentions)

Spelling variants of this product

Anti-CD33-APC (8 citations) APC‐CD33 clone WM53 (4 citations) APC-conjugated anti-CD33 antibody (2 citations) APC-conjugated CD33 (2 citations)

21 protocols using cd33 apc

1

CD34+ Hematopoietic Progenitor Differentiation

Cited 4 times
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Unstimulated and TLR-Stimulated CD34+ hematopoietic progenitors were plated at 10,000 cells/well for liquid, stromal cell-free B lineage cultures, or 1000 cells per sample in Methocult H4434 Classic (Stemcell Technologies), according to the manufacturer’s directions, as previously described [4 (link),5 (link),18 (link),19 (link)]. For B lineage liquid culture, cells were grown in QBSF®60 (Quality Biological, Inc.) supplemented with 10% FBS (Atlanta Biologicals), 100U Penicillin/Streptomycin (Gibco), 10% hASC conditioned media (LaCell LLC), 10 ng/ml stem cell factor, 10 ng/ml granulocyte stimulating factor, 5 ng/ml FLT-3 ligand, and 5 ng/ml IL-7 (as above). Half of the media was replaced once a week. After 4 weeks, cells were harvested, counted and assessed by flow cytometry using CD34-PE, CD10-Pacific Blue, CD19-PE-Cy5 (BioLegend), CD33-APC (BD Biosciences), and goat anti-ARID3a with a FITC-conjugated anti-goat secondary. Methocult cultures were incubated for 14 days and colonies were analyzed visually for monocytic, granulocytic and erythrocytic lineage cell colonies and counted using a Nikon Eclipse TS100 inverted microscope. Methocult cultures were harvested with warm 1X PBS, washed twice with warm PBS and once with ice cold PBS and then assessed by flow cytometry for the following surface markers: CD34, CD33, CD14, CD16, and CD41 with intracellular ARID3a.
Ratliff M.L., Shankar M., Guthridge J.M., James J.A, & Webb C.F. (2020). TLR engagement induces ARID3a in human blood hematopoietic progenitors and modulates IFNα production. Cellular immunology, 357, 104201.
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2

Phospho-specific Flow Cytometric Analysis of PBMC Signaling

Cited 1 time
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Freshly thawed PBMCs were resuspended in serum-free, antibiotic-free RPMI 1640 media. Cells were distributed (0.5 x 106cells in 100 μL per tube) to Falcon polystyrene round-bottom tubes (12 x 75 mm; Corning Incorporated, Durham, NC) and treated with IL-6 or IFN-γ (100 ng/mL) (R&D Systems, Minneapolis, MN) for 15 min at 37°C before subjecting them to phospho-specific flow cytometric analysis as described previously [7 (link)]. Briefly, after stimulation, cells were fixed by incubating in 2% PFA (BD Cytofix Fixation Buffer; BD Biosciences) for 10 min at 37°C and pelleted. They were then permeabilized by resuspending with vigorous vortexing in 300 μL ice-cold methanol. Cells were washed in staining media. Fluorophore-specific MAbs were added and incubated for 30 min at RT. The following markers were analyzed: CD3-PE/Cy7, CD4-APC/Cy7, CD45RO-PerCP/Cy5.5, CD33-APC, STAT1 (pY701)-AF488, STAT3 (pY705)-AF488, (BD Biosciences, San Diego, CA); and CD45-PE (R&D Systems, Minneapolis, MN). The cells were washed with staining media and pelleted. Finally, the samples were resuspended in 250 μL of staining media and analyzed.
Camargo J.F., Bhimji A., Kumar D., Kaul R., Pavan R., Schuh A., Seftel M., Lipton J.H., Gupta V., Humar A, & Husain S. (2015). Impaired T Cell Responsiveness to Interleukin-6 in Hematological Patients with Invasive Aspergillosis. PLoS ONE, 10(4), e0123171.
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3

Investigating Leukemia Cell-Derived EV Effects

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In the differentiation experiment, LSCs were cultured with Iscove’s Modified Dulbecco’s Medium (IMDM) medium (Gibco, Carlsbad, CA, USA) containing 10% FBS, 100 ng/mL recombinant human stem cell factor (rhSCF), 100 ng/mL rhFlt3 and 100 ng/mL for recombinant human thrombopoietin (rhTPO) with 5% CO2 at 37° C for 24 h. After culture, LSCs were treated with 1 μg/mL AML cell-derived EVs for 7 days. Next, LSCs were collected and labeled with CD19-PE (555413, BD Biosciences, San Jose, CA, USA), CD33-APC (561817, BD Biosciences, San Jose, CA, USA), CD3-APC (561811, BD Biosciences, San Jose, CA, USA), and CD235a-FITC (559943, BD Biosciences, San Jose, CA, USA) for 30 min. Cells were then detected using a FACS Calibur (BD Biosciences, San Jose, CA, USA) and analyzed with the Cell Quest software (BD Biosciences, San Jose, CA, USA) [33 (link)].
Chen L., Guo Z., Zhou Y., Ni J., Zhu J., Fan X., Chen X., Liu Y., Li Z, & Zhou H. (2021). microRNA-1246-containing extracellular vesicles from acute myeloid leukemia cells promote the survival of leukemia stem cells via the LRIG1-meditated STAT3 pathway. Aging (Albany NY), 13(10), 13644-13662.
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4

Immunophenotyping of Hematological Cells

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For analysis of surface markers, cells were stained in PBS containing 2% BSA. The following fluorescent‐labeled antibodies (purchased from BD Biosciences) were used: CD13 PE (652820), CD33 APC (652807), MPO FITC (652821), and CD45 PerCP (652803). Flow cytometry data were collected using Canto‐I (BD Biosciences) cytometers and analyzed with FlowJo V10 software.
Xu Y., Yu Q., Wang P., Wu Z., Zhang L., Wu S., Li M., Wu B., Li H., Zhuang H., Zhang X., Huang Y., Gan X, & Xu R. (2022). A Selective Small‐Molecule c‐Myc Degrader Potently Regresses Lethal c‐Myc Overexpressing Tumors. Advanced Science, 9(8), 2104344.
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5

Flow Cytometry Immunostaining Analysis

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Cell surface and intracellular immunostaining analyses were performed using an Accuri C6 Flow Cytometer. NK cells and T cells were stained with the dye-conjugated mouse mAbs to human CD56-PE-Cy5 (Beckman Coulter), CD3-PE (eBioscience), CCR7-FITC (R&D Systems), granzyme B-PE (Invitrogen), and CD16-FITC, CD8-PE-Cy5, CD45RA-FITC, CD45RO-PE, and CD57-FITC (BD Biosciences). MDSCs were stained for CD11b-FITC, CD14-PE, CD33-APC, CD34-PE-Cy5, CD11c-PE, HLA-DR-PE, DC-SIGN-FITC, CD80-FITC, CD86-FITC, and CD83-PE (BD Biosciences and eBioscience), as well as IDO-A488 (R&D Systems), NOS2-PE (Santa Cruz Biotechnology), and COX1-FITC/COX2-PE (BD Biosciences). The corresponding mouse antibody isotype controls IgG1-FITC, IgG2b-FITC, IgG1-PE, IgG2a-PE, IgG1-PE-Cy5, IgG1-APC, and IgG1-A488 (BD Biosciences) were used, as appropriate. Before staining, the cells were treated for 20 min at 4°C in PBS buffer containing 2% human serum, 0.5% BSA, 0.1% NaN3, and 1 μg/ml of mouse IgG (Sigma-Aldrich) to block non-specific binding. Cell permeabilization for intracellular staining was performed using the Foxp3 Fix/Perm Buffer Set (eBioscience), according to the manufacturer’s protocol. Cells were stained for 40 min at 4°C followed by washing with PBS buffer containing 0.5% BSA and 0.1% NaN3, then fixed and stored in 4% paraformaldehyde until analysis.
Wong J.L., Obermajer N., Odunsi K., Edwards R.P, & Kalinski P. (2016). Synergistic COX2 induction by IFNγ and TNFα self-limits type-1 immunity in the human tumor microenvironment. Cancer immunology research, 4(4), 303-311.
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6

Evaluating AML Therapy in NOD/SCID Mice

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NOD/SCID mice were bred and housed at the UHN/Princess Margaret Hospital (PMH; Toronto, Ontario, Canada) animal facility, and all studies were performed in accordance with guidelines approved by the UHN/PMH Animal Care Committee. Eight- to 12-week-old female NOD/SCID mice (10 per cohort) were sublethally irradiated (275 cGy) and interperitoneally injected with anti-CD122 antibody the day before intrafemoral transplantation. Freshly thawed primary AML samples harvested from patients’ peripheral blood were transplanted at cell doses of 5e6/mouse. At day 21, post transplantation, AGS67E and an isotype control ADC were dosed by i.v. injection at 1.5 mg/kg, QW for a total of 4 doses. Mice were sacrificed 7 days after the last treatment to assess the efficacy of AGS67E determined by the human AML engraftment in the injected right femur and non-injected bone marrow (left femur and two tibias). AML outgrowth was evaluated by flow cytometry using the following antibodies: CD45-FITC (BD), CD33-APC (BD), CD34-PE-Cy5 (Beckman Coulter), CD3-ECD (Beckman Coulter), CD38-PE-Cy7 (BD), and AGS67C-Biotin. Secondary detection of biotinylated antibodies utilized streptavidin–PE. Samples were analyzed using an LSRII flow cytometer (BD).
Pereira D.S., Guevara C.I., Jin L., Mbong N., Verlinsky A., Hsu S.J., Aviña H., Karki S., Abad J.D., Yang P., Moon S.J., Malik F., Choi M.Y., An Z., Morrison K., Challita-Eid P.M., Doñate F., Joseph I.B., Kipps T.J., Dick J.E, & Stover D.R. (2015). AGS67E, an Anti-CD37 Monomethyl Auristatin E Antibody–Drug Conjugate as a Potential Therapeutic for B/T-Cell Malignancies and AML: A New Role for CD37 in AML. Molecular cancer therapeutics, 14(7), 1650-1660.
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7

Quantifying AML Myeloid Blast and LSC Populations

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PBMCs from AML patients were isolated and incubated with a cocktail of CD45-FITC (BD), CD33-APC (BD), CD34-PE-Cy5 (Beckman Coulter), CD3-ECD (Beck-man Coulter), CD38-PE-Cy7 (BD), and either AGS67C-Biotin or isotype-Biotin antibodies. Secondary detection for biotinylated antibodies was accomplished with Streptavidin-PE. An LSRII flow cytometer (BD) was used for acquisition. Within the CD45+ population, two distinct subpopulations were defined, CD33+/CD3−/CD20− (myeloid blasts) and CD33+/CD3−/CD34+/CD38− [leukemic stem cells (LSC)]. Analysis was done with FlowJo version 9.5.4. MFIR for each AML sample was calculated by dividing the AGS67C MFI by the isotype MFI.
Pereira D.S., Guevara C.I., Jin L., Mbong N., Verlinsky A., Hsu S.J., Aviña H., Karki S., Abad J.D., Yang P., Moon S.J., Malik F., Choi M.Y., An Z., Morrison K., Challita-Eid P.M., Doñate F., Joseph I.B., Kipps T.J., Dick J.E, & Stover D.R. (2015). AGS67E, an Anti-CD37 Monomethyl Auristatin E Antibody–Drug Conjugate as a Potential Therapeutic for B/T-Cell Malignancies and AML: A New Role for CD37 in AML. Molecular cancer therapeutics, 14(7), 1650-1660.
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8

Quantifying CXCR4 Expression in AML and CD34+ Cells

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CXCR4 expression was determined in AML and CD34+ cells treated with IMiDs. A total of 0.2 × 106 (for cell lines) or 1 × 106 (for primary cells) cells were incubated with 10 µM of IMiDs (the CXCR4 antagonist AMD3100 was used as a positive control). Seven days later (48 h–72 h for primary AML cells) treated cells were stained with anti-human CD45-APC.Cy7, CD33-APC, CD34-PECy7 and CD184-BV421 (BD Biosciences) and the mean fluorescence intensity (MFI) of CD184 within the blast population (CD45+CD33+) was FACS-quantified using a FACS Canto II cytometer. The expression of the CXCR4 ligand SDF1 was analyzed in BM-MSC (0.5 x 106) 48 h after treatment with 10 µM IMiDs. BM-MSCs were trypsinized and stained with anti-human CXCL12/SDF-1 (R&D Systems) using the Cell Permeabilization Kit (FIX&PERM®).26
Lopez-Millan B., Diaz de la Guardia R., Roca-Ho H., Anguita E., Islam A.B., Romero-Moya D., Prieto C., Gutierrez-Agüera F., Bejarano-Garcia J.A., Perez-Simon J.A., Costales P., Rovira M., Marín P., Menendez S., Iglesias M., Fuster J.L., Urbano-Ispizua A., Anjos-Afonso F., Bueno C, & Menendez P. (2018). IMiDs mobilize acute myeloid leukemia blasts to peripheral blood through downregulation of CXCR4 but fail to potentiate AraC/Idarubicin activity in preclinical models of non del5q/5q- AML. Oncoimmunology, 7(9), e1477460.
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9

Multiparametric Flow Cytometry Immunophenotyping

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Cells grown in EHT culture or harvested animal tissues were stained with the following antibody panels. Haemogenic endothelium panel: CD34 PE-Cy7 (8G12; BD), FLK1 AF647 (89106; BD), CD235a/glycophorin (GLY)-A FITC (11E4B-7-6; Coulter), and CD43 PE (1G10; BD). HSPC panel: CD38 PE-Cy5 (LS198-4-3; Clontech), CD34 PE-Cy7, and CD45 PE (HI30; BD). Lineage panel: CD235a/glycophorin (GLY)-A PE-Cy7 or FITC (11E4B-7-6; Coulter), CD33 APC (P67.6; BD), CD19 PE (HIB19; BD), IgM BV510 (G20-127; BD), CD4 PE-Cy5 (13B8.2; Coulter), CD3 PE-Cy7 (SK7; BD), CD8 V450 (RPA-T8; BD), TCRαβ BV510 (T10B9; BD), TCRγδ APC (B1; BD), CD45 PE-Cy5 (J33; Coulter), CD15 APC (HI98; BD), and CD31/PECAM PE (WM59; BD). All stains were performed with fewer than 1 × 106 cells per 100 μl staining buffer (PBS + 2% FBS) with 1:100 dilution of each antibody, for 30 min at 4 °C in the dark. Compensation was performed by automated compensation with anti-mouse Igk negative beads (BD) and cord blood MNC stained with individual antibodies. All acquisitions were performed on a BD Fortessa cytometer. For detection of engraftment, human cord-blood-engrafted mouse marrow was used as a control to set gating; sorting was performed on a BD FACS Aria II cell sorter.
Sugimura R., Jha D.K., Han A., Soria-Valles C., da Rocha E.L., Lu Y.F., Goettel J.A., Serrao E., Rowe R.G., Malleshaiah M., Wong I., Sousa P., Zhu T.N., Ditadi A., Keller G., Engelman A.N., Snapper S.B., Doulatov S, & Daley G.Q. (2017). Haematopoietic stem and progenitor cells from human pluripotent stem cells. Nature, 545(7655), 432-438.
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10

CD34+ Hematopoietic Progenitor Differentiation

Cited 4 times
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Unstimulated and TLR-Stimulated CD34+ hematopoietic progenitors were plated at 10,000 cells/well for liquid, stromal cell-free B lineage cultures, or 1000 cells per sample in Methocult H4434 Classic (Stemcell Technologies), according to the manufacturer’s directions, as previously described [4 (link),5 (link),18 (link),19 (link)]. For B lineage liquid culture, cells were grown in QBSF®60 (Quality Biological, Inc.) supplemented with 10% FBS (Atlanta Biologicals), 100U Penicillin/Streptomycin (Gibco), 10% hASC conditioned media (LaCell LLC), 10 ng/ml stem cell factor, 10 ng/ml granulocyte stimulating factor, 5 ng/ml FLT-3 ligand, and 5 ng/ml IL-7 (as above). Half of the media was replaced once a week. After 4 weeks, cells were harvested, counted and assessed by flow cytometry using CD34-PE, CD10-Pacific Blue, CD19-PE-Cy5 (BioLegend), CD33-APC (BD Biosciences), and goat anti-ARID3a with a FITC-conjugated anti-goat secondary. Methocult cultures were incubated for 14 days and colonies were analyzed visually for monocytic, granulocytic and erythrocytic lineage cell colonies and counted using a Nikon Eclipse TS100 inverted microscope. Methocult cultures were harvested with warm 1X PBS, washed twice with warm PBS and once with ice cold PBS and then assessed by flow cytometry for the following surface markers: CD34, CD33, CD14, CD16, and CD41 with intracellular ARID3a.
Ratliff M.L., Shankar M., Guthridge J.M., James J.A, & Webb C.F. (2020). TLR engagement induces ARID3a in human blood hematopoietic progenitors and modulates IFNα production. Cellular immunology, 357, 104201.
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