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Cd19 pe hib19

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CD19 PE (HIB19) is a laboratory reagent used in flow cytometry applications. It is an anti-CD19 monoclonal antibody conjugated with the fluorescent dye phycoerythrin (PE). CD19 is a cell surface marker expressed on B lymphocytes, and this reagent can be used to identify and quantify B cells in biological samples.

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Lab products found in correlation

Cd45 pe cy5, by Beckman Coulter (4 mentions) Cd3 pe cy7, by BD (4 mentions) Qiaamp dna micro kit, by Qiagen (2 mentions) Fetal bovine serum (fbs), by STEMCELL (2 mentions) Cal lyse, by Thermo Fisher Scientific (2 mentions) Anti cd3 fitc ucht1, by BD (2 mentions) Cd45 pe, by BD (2 mentions) Fortessa cytometer, by BD (2 mentions) Cd33 apc, by BD (2 mentions) Flk1 af647, by BD (2 mentions) Cd235, by BD (2 mentions) Gly a fitc 11e4b 7 6, by Beckman Coulter (2 mentions) Cd235a, by Beckman Coulter (2 mentions) Gly a pe cy7 or fitc 11e4b 7 6, by Beckman Coulter (2 mentions) Igm bv510, by BD (2 mentions) Cd4 pe cy5, by Beckman Coulter (2 mentions) Cd8 v450 rpa t8, by BD (2 mentions) Tcrαβ bv510, by BD (2 mentions) Cd31 pecam pe, by BD (2 mentions) Anti mouse igk negative beads, by BD (2 mentions)

9 protocols using cd19 pe hib19

1

Lentiviral Gene Transfer and Engraftment

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Twelve hours after lentiviral gene transfer, cells were recovered by dispase for 5 min at 37 °C, and washed by PBS three times to ensure no carry-over of virus. Cells were resuspended at 0.3 × 105 to 3.0 × 105 cells per 25 μl buffer (PBS + 2% FBS from StemCell Technologies) and kept on ice until injection. Thirty thousand to 3.0 × 105 cells were intrafemorally injected in to NOD/LtSz-scidIL2Rgnull (NSG) mice and treated with doxycycline as described below (see section on ‘Mouse transplantation’). Up to 100 μl peripheral blood was collected every 2–4 weeks, to 14 weeks. Mice were euthanized and bone marrow and thymus removed at 8–14 weeks. For transgene detection in engrafted cells, each lineage of cells was FACS-sorted from bone marrow. Myeloid cells: CD33 APC (P67.6; BD), CD45 PE-Cy5 (J33; Coulter). B cells: CD19 PE (HIB19; BD), CD45 PE-Cy5 (J33; Coulter). T cells: CD3 PE-Cy7 (SK7; BD), CD45 PE-Cy5 (J33; Coulter). Between 10,000 and 50,000 cells were isolated with 2 or 3 biological replicates for multiple cell lines (iPSCs and ESCs). The QIAamp DNA Micro kit (Qiagen) was used to collect and prepare total genomic DNA for PCR detection of transgenes. Nested PCR reaction was performed similarly to the in vitro screening described in the above section.
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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2

Quantifying Leukocyte Depletion in CLL

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Whole blood from CLL patients (90 μl) was incubated with 10 μg/ml antibody for 1 hour at 37°C. 27 After this treatment, blood was stained with anti-CD3 FITC (UCHT1) and CD19 PE (HIB19) from BD Biosciences. Cal-lyse (Invitrogen) was used to simultaneously lyse RBCs while fixing the leukocytes. Count Bright absolute counting beads were added just prior to analysis by flow cytometry in order to calculate absolute concentrations of cells. To adjust for differences in initial leukemia counts between patients, data was normalized as follows: Percent depletion = 100 × (1 – [concentration of cells in treated sample] ÷ [concentration of cells in untreated control]). Additional details are provided in the supplemental materials.
Beckwith K.A., Frissora F.W., Stefanovski M.R., Towns W.H., Cheney C., Mo X., Deckert J., Croce C.M., Flynn J.M., Andritsos L.A., Jones J.A., Maddocks K.J., Lozanski G., Byrd J.C, & Muthusamy N. (2014). The CD37-targeted antibody-drug conjugate IMGN529 is highly active against human CLL and in a novel CD37 transgenic murine leukemia model. Leukemia, 28(7), 1501-1510.
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3

Regulatory T Cell Phenotyping in Food Tolerance

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To investigate the role of T regulatory cells in natural tolerance to foods, cell surface and intracellular cytokine staining was performed using validated methods [23 (link)]. Unstimulated and PBMCs stimulated with ovalbumin and peanut extract were surface stained with markers from the indicated sources (CD3 APC-CY7 (SK7), Biolegend; CD4 V-500 (RPA-T4), BD; CD14 Alexa Fluor-700 (HCD14), Biolegend; CD19 PE (HIB19), BD; RORgT PE (AFKJS-9), eBiosciences; CD25 APC (BC96), Biolegend; and CD127 Pacific Blue (A019D5), Biolegend) and then washed, fixed, and permeabilized for intracellular staining (Foxp3 Alexa Fluor 488 (206D), Biolegend; IL-10 PE-CY7 (JES3-9D7), Biolegend; IL-5 APC (TRFK5), Biolegend) according to manufacturer’s instructions (Caltag, Carlsbad, CA). Acquisition was performed using an LSR-II (BD, San Jose, CA), and analyzed using FlowJo software (Treestar, San Carlos, CA). Data are expressed as the mean percentage of cells within the gated CD3CD4 T cells unless otherwise indicated.
Qamar N., Fishbein A.B., Erickson K.A., Cai M., Szychlinski C., Bryce P.J., Schleimer R.P., Fuleihan R.L, & Singh A.M. (2015). Naturally Occurring Tolerance Acquisition to Foods in Previously Allergic Children is Characterized by Antigen Specificity and Associated with Increased Subsets of Regulatory T cells. Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology, 45(11), 1663-1672.
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4

Quantifying Leukocyte Depletion in CLL

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Whole blood from CLL patients (90 μl) was incubated with 10 μg/ml antibody for 1 hour at 37°C. 27 After this treatment, blood was stained with anti-CD3 FITC (UCHT1) and CD19 PE (HIB19) from BD Biosciences. Cal-lyse (Invitrogen) was used to simultaneously lyse RBCs while fixing the leukocytes. Count Bright absolute counting beads were added just prior to analysis by flow cytometry in order to calculate absolute concentrations of cells. To adjust for differences in initial leukemia counts between patients, data was normalized as follows: Percent depletion = 100 × (1 – [concentration of cells in treated sample] ÷ [concentration of cells in untreated control]). Additional details are provided in the supplemental materials.
Beckwith K.A., Frissora F.W., Stefanovski M.R., Towns W.H., Cheney C., Mo X., Deckert J., Croce C.M., Flynn J.M., Andritsos L.A., Jones J.A., Maddocks K.J., Lozanski G., Byrd J.C, & Muthusamy N. (2014). The CD37-targeted antibody-drug conjugate IMGN529 is highly active against human CLL and in a novel CD37 transgenic murine leukemia model. Leukemia, 28(7), 1501-1510.
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5

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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6

Lentiviral Gene Transfer and Engraftment

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Twelve hours after lentiviral gene transfer, cells were recovered by dispase for 5 min at 37 °C, and washed by PBS three times to ensure no carry-over of virus. Cells were resuspended at 0.3 × 105 to 3.0 × 105 cells per 25 μl buffer (PBS + 2% FBS from StemCell Technologies) and kept on ice until injection. Thirty thousand to 3.0 × 105 cells were intrafemorally injected in to NOD/LtSz-scidIL2Rgnull (NSG) mice and treated with doxycycline as described below (see section on ‘Mouse transplantation’). Up to 100 μl peripheral blood was collected every 2–4 weeks, to 14 weeks. Mice were euthanized and bone marrow and thymus removed at 8–14 weeks. For transgene detection in engrafted cells, each lineage of cells was FACS-sorted from bone marrow. Myeloid cells: CD33 APC (P67.6; BD), CD45 PE-Cy5 (J33; Coulter). B cells: CD19 PE (HIB19; BD), CD45 PE-Cy5 (J33; Coulter). T cells: CD3 PE-Cy7 (SK7; BD), CD45 PE-Cy5 (J33; Coulter). Between 10,000 and 50,000 cells were isolated with 2 or 3 biological replicates for multiple cell lines (iPSCs and ESCs). The QIAamp DNA Micro kit (Qiagen) was used to collect and prepare total genomic DNA for PCR detection of transgenes. Nested PCR reaction was performed similarly to the in vitro screening described in the above section.
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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7

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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8

Multilineage Cell Identification by FACS

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Five thousand to 10,000 FACS-sorted erythroid cells (CD235a/glycophorin (GLY)-A PE-Cy7 or FITC (11E4B-7-6; Coulter)), plasmacytoid lymphocytes (CD19 PE (HIB19; BD), IgM BV510 (G20-127; BD), CD38 PE-Cy5 (LS198-4-3; Clontech)), neutrophils (CD15 APC (HI98; BD), CD31/PECAM PE (WM59; BD) and CD45 PE-Cy5 (J33; Coulter)) were cytospun onto slides (500 r.p.m. for 10 min), air dried, and stained with May-Grunwald and Giemsa stains (both from Sigma), washed with water, air dried, and mounted, followed by examination by light microscopy.
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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9

Multilineage Cell Identification by FACS

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Five thousand to 10,000 FACS-sorted erythroid cells (CD235a/glycophorin (GLY)-A PE-Cy7 or FITC (11E4B-7-6; Coulter)), plasmacytoid lymphocytes (CD19 PE (HIB19; BD), IgM BV510 (G20-127; BD), CD38 PE-Cy5 (LS198-4-3; Clontech)), neutrophils (CD15 APC (HI98; BD), CD31/PECAM PE (WM59; BD) and CD45 PE-Cy5 (J33; Coulter)) were cytospun onto slides (500 r.p.m. for 10 min), air dried, and stained with May-Grunwald and Giemsa stains (both from Sigma), washed with water, air dried, and mounted, followed by examination by light microscopy.
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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