The subsets of T lymphocytes14 (link),15 (link) and monocytes12 (link),13 (link) at baseline and at following timepoints during PU-H71 treatment were monitored by flow cytometry. Peripheral blood (PB) cells from healthy donor was used as a control. Cells were isolated by the Ficoll-Paque method from PB and bone marrow (BM) aspirates. Cells were labeled with anti-CD45 APC-H7, anti-CD33 BV650 (BD Biosciences, clone WM53, cat. 303430), anti-CD56 AlexaFluor700 (BioLegend, clone HCD56, cat. 318316), anti-CD3 BV711 (BioLegend, clone SK7, cat. 344838), anti-CD4 PE-Cy5 (BioLegend, clone OKT4, cat. 317412), anti-CD8 BV605 (BioLegend, clone SK1, cat. 344742), anti-CD19 PerCP/Cy5.5 (BioLegend, clone HIB19, cat. 302230), anti-CD14 PE (BD Biosciences, clone M5E2, cat. 555398), anti-CD16 BV785 (BioLegend, clone 3G8, cat. 302046), anti-CD45RA PE-Cy7 (BioLegend, clone HI100, cat. 304126) and anti-CCR7 Alexa Fluor 647 (BD Biosciences, clone 150503, cat. 560816). After washing, flow cytometry evaluation was performed on BD LSRFortessa. Data was analyzed using FlowJo software.
Anti cd4 pe cy5
Manufactured by BioLegend
Anti-CD4-PE-Cy5 is a fluorescently-labeled antibody that binds to the CD4 surface molecule. CD4 is a glycoprotein expressed on the surface of T helper cells, which play a crucial role in the immune response. The PE-Cy5 conjugate provides a specific fluorescent signal for the detection and analysis of CD4-positive cells using flow cytometry.
Automatically generated - may contain errors
Lab products found in correlation
Anti cd56 alexafluor700, by BioLegend (1 mentions)
Anti cd3 bv711, by BioLegend (1 mentions)
Anti cd8 bv605, by BioLegend (1 mentions)
Anti cd19 percp cy5, by BioLegend (1 mentions)
Anti cd14 pe, by BD (1 mentions)
Anti cd16 bv785, by BioLegend (1 mentions)
Anti cd45ra pe cy7, by BioLegend (1 mentions)
Anti ccr7 alexa fluor 647, by BD (1 mentions)
Lsrfortessa, by BD (1 mentions)
Anti cd19 fitc, by BD (1 mentions)
Anti cd3 pe cy7, by BD (1 mentions)
Anti cd8 pe cy5, by BD (1 mentions)
Anti cd69 fitc, by BD (1 mentions)
Anti cd11c pe cy 5, by BD (1 mentions)
Anti cd86 pe cy7, by BD (1 mentions)
Anti hla dr pe, by BD (1 mentions)
Anti cd123 pe cy 7, by BD (1 mentions)
Anti cd20 pe, by BD (1 mentions)
Anti cd279 pd 1 apc, by BD (1 mentions)
Anti cd11b pe, by BD (1 mentions)
4 protocols using anti cd4 pe cy5
1
Multiparametric Flow Cytometry Analysis of Immune Cell Subsets
2
Lymph Node Immunophenotyping by Flow Cytometry
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The following extracellular, anti-human monoclonal antibodies were used for lymph node immunophenotyping: anti-CD3 PE-Cy 7, anti-CD8+PE-Cy 5, anti-CD4 FITC, anti-CD62L PE-Cy 5, anti-CD69 FITC, anti-CD152 (CTLA-4) PE, anti-CD11c PE-Cy 5, anti-CD86 PE-Cy 7, anti-HLA-DR PE, anti-CD3 FITC, anti-CD14 FITC, anti-CD16 FITC, anti-CD19 FITC, anti-CD123 PE-Cy 7, anti-CD141 PE, anti-CD68 PE-Cy 5, anti-CD20 PE, anti-CD56 PE, anti-CD279 (PD-1) APC, anti-CD11b PE, anti-CD64 FITC (BD Pharmingen, San Jose, CA).
The human monoclonal antibodies, anti-CD4 PE-Cy 5 and anti-CD25 PE, were purchased from Biolegend (San Diego, CA) and used in conjunction with intracellular staining with anti-FoxP3 FITC for the quantification of regulatory T cells. Cell death was determined by propidium iodine staining (eBioscience, San Diego, CA USA). Four-color flow cytometry was performed on a Guava 8HT flow cytometer (EMD Millipore, Billerica, MA USA) capturing 25,000 events for all samples. Results were analyzed using Guava Soft Incyte (EMD Millipore, Billerica, MA USA).
The human monoclonal antibodies, anti-CD4 PE-Cy 5 and anti-CD25 PE, were purchased from Biolegend (San Diego, CA) and used in conjunction with intracellular staining with anti-FoxP3 FITC for the quantification of regulatory T cells. Cell death was determined by propidium iodine staining (eBioscience, San Diego, CA USA). Four-color flow cytometry was performed on a Guava 8HT flow cytometer (EMD Millipore, Billerica, MA USA) capturing 25,000 events for all samples. Results were analyzed using Guava Soft Incyte (EMD Millipore, Billerica, MA USA).
3
Multiparametric Flow Cytometry Analysis
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The following anti-human fluorochrome-conjugated antibodies were used: Alexa Fluor 700-conjugated antibody to human CD3 (UCHT1); anti-IL-7Rα-BV421 (A019D5), anti-CD4-PE-Cy5 (OKT4), anti-CD45RA-APC-Cy7 (HI100), anti-Fas-BV650 (DX2), anti-CCR7-BV785 (G043H7), anti-CD31-Alexa Fluor 488 (WM59), anti-TCR γ/δ-Percp-Cy5.5 (B1), anti-CD25-BV650 (BD96; BioLegend), anti-CD19-BV711 (SJ25C1), anti-CD14-BV711 (MφP9), anti-CD3-BV510 (HIT3a), anti-CD8-Alexa Fluor 700 (RPA-T8; all from BD Biosciences); anti-CD19-PE-eFluor610 (HIB19); anti-CD14-PE-eFluor610 (61D3), and anti-CD56-APC (CMSSB; Thermo Fisher). Simultaneously, dead cells were stained using a Live/Dead Fixable Red Dead Cell Stain Kit (Thermo Fisher). The cells were fixed and permeabilized using a Foxp3 Staining Buffer Set (Thermo Fisher), and then intracellular molecules were stained (4°C, 20 minutes) using anti-Bcl-2-Alexa Fluor 488 (100; BioLegend), anti-Ki-67-PE-Cy7 (20Raj1), and anti-Foxp3-PE (236A/E7; Thermo Fisher). Detailed information about the multicolor flow cytometry panel used in this study is shown in supplemental Table 4 .
4
Flow Cytometry Immunophenotyping of BAL Cells
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Cells in the BAL compartment were incubated in human gamma globulin to block Fc receptors and prevent nonspecific binding of antibodies. Cells were washed and incubated in anti-CD3ε-Brilliant Violet 510 (BioLegend, San Diego, CA), anti-CD4-PE-Cy5 (BioLegend), anti-CD193-Alexa Fluor 647 (BD Biosciences, San Jose, CA), anti-Ly6G-V450 (BD Biosciences), and anti-Siglec-F-PE-CF594 (BD Biosciences) on ice for 30 minutes. Controls included unstained cells, isotype controls, and single color controls. Cells were washed and fixed with stabilizing fixative (BD Biosciences) and data were acquired with a BD LSR Fortessa and analyzed with FlowJo v10.1r5 (Ashland, OR).
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