Cells were analyzed using Flow Cytometer (Cytomic FC500, Beckman). Tissues were trim into 1-2mm3 tissue block and put into burnisher. Then cell suspension was collected and centrifugated. To blood specimen, heparin sodium and PBS/Hanks was added into blood of individuals. Ficol was used to separate lymphocyte. Serum free medium containing 1% BSA was added into cell suspension, and then incubated on ice for 10 minutes. PE-Cy Mouse Anti-Human CD11b, FITC-Mouse Anti-Human LIN, APC Mouse Anti-Human HLA-DR, PE Mouse Anti-Human CD33 (BD Biosciences) were added to cell suspension and incubated on ice for 30 minutes. Cells were washed and resuspended in 500 mL buffer and analyzed.
Apc mouse anti human hla dr
Manufactured by BD
Sourced in United States
The APC Mouse Anti-Human HLA-DR is a laboratory equipment product used for flow cytometry analysis. It is a monoclonal antibody conjugated with allophycocyanin (APC) that binds to the HLA-DR antigen expressed on the surface of human cells.
Automatically generated - may contain errors
Lab products found in correlation
Pe mouse anti human cd33, by BD (3 mentions)
Fitc mouse anti human lin, by BD (3 mentions)
Pe cy mouse anti human cd11b, by BD (2 mentions)
Pe mouse anti human cd163, by BD (2 mentions)
Fitc mouse anti human cd68, by BD (2 mentions)
Facscanto 2 cytometer, by BD (1 mentions)
Cytomic fc500, by Beckman Coulter (1 mentions)
Histopaque 1077, by Merck Group (1 mentions)
Fitc rat anti mouse cd8a, by BD (1 mentions)
Pe rat anti mouse f4 80, by BD (1 mentions)
Apc rat anti mouse cd11b, by BD (1 mentions)
Facscalibur flow cytometer, by BD (1 mentions)
Fbs rpmi 1640, by Merck Group (1 mentions)
Gm csf, by Thermo Fisher Scientific (1 mentions)
Interleukin 4 (il 4), by Merck Group (1 mentions)
Gm csf, by Merck Group (1 mentions)
Facscelesta instrument, by BD (1 mentions)
Tnf α, by Merck Group (1 mentions)
Facscalibur ow cytometer, by BD (1 mentions)
8 protocols using apc mouse anti human hla dr
1
Multicolor Flow Cytometry for Immune Profiling
2
Modulation of MoDC Activation by α-Synuclein
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MoDCs were seeded at a density of 1 × 105 cells/ml in RPMI 1640 medium 1% FBS in 24-well cell culture plate overnight at 37°C, 5% CO2. C1 (1 µM) and Celastrol (0.25 µM) were added to pre-treat MoDCs for 1 h followed by the addition of fibrillar α-syn (1 µg/ml) for 24 h. In some cases, TNF-α (50 ng/ml) was added together with fibrillar α-syn. LPS (100 ng/ml) was used as the positive control. Afterward, MoDCs were collected and washed with fluorescence-activated cell sorter (FACS) buffer (PBS + 1% FBS) and incubated for 30 min at 4°C in 100 µl FACS Buffer with BV421 Mouse anti-Human HLA-ABC (Cat. No. 565332, BD Pharmingen), APC Mouse anti-Human HLA-DR (Cat. No. 560896, BD Pharmingen), PE-Cy7 Mouse anti-Human CD86 (Cat. No. 561128, BD Pharmingen) and FITC Mouse anti-Human CD80 (Cat. No. 555683, BD Pharmingen) antibodies. Flow cytometry was performed following standard protocols on a BD FACSCanto™ II cytometer.
3
Isolation and Characterization of MDSCs
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Peripheral blood mononuclear cells (PBMCs) from blood samples were separated by Ficoll-Hypaque density gradient centrifugation. And MDSCs were sorted from PBMCs by using CD33 labeled magnetically selection monoclonal antibodies (Miltenyi Biotech, Germany). PE mouse anti-human CD33, PE-Cy7 mouse anti-human CD11b, APC mouse anti-human HLA-DR and FITC mouse anti-human LIN (BD Biosciences, USA) were applied to examine the proportion of MDSCs by Flow Cytometry (Cytomic FC500, Beckman). WinMDI was used to analyze samples.
4
Immune Cell Isolation and Characterization
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Tissues were trim into 1-2mm3 tissue block and put into burnisher. Then cell suspension was collected and centrifugated. To blood specimen, heparin sodium and PBS/Hanks was added into blood of individuals. Ficol was used to separate lymphocyte. Serum free medium containing 1% BSA was added into cell suspension, and then incubated on ice for 10 minutes. PE-Cy Mouse Anti-Human CD11b, FITC- Mouse Anti-Human LIN, APC Mouse Anti-Human HLA-DR, PE Mouse Anti-Human CD33 (BD Biosciences) were added to cell suspension and incubated on ice for 30 minutes. Cells were washed and resuspended in 500 mL buffer and analyzed using Flow Cytometer (Cytomic FC500, Beckman).
5
Immune Cell Phenotyping from Tissues
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Immune cells from animal tissue or blood were separated by using HISTOPAQUE®‐1077(10771, Sigma), and then the dissociated lymphocytes were resuspended in PBS and stained for surface markers for 20 minutes with antibodies: PerCP‐CyTM 5.5 rat anti‐mouse CD45 (550994, BD Biosciences, 1:200), FITC rat anti‐mouse CD8a (553030, BD Biosciences, 1:500), APC rat anti‐mouse CD11b (553312, BD Biosciences, 1:200), PE rat anti‐mouse F4/80 (565410, BD Biosciences, 1:200), APC anti‐mouse CD206 (MMR) (141707, BD Biosciences, 1:200), APC mouse anti‐human HLA‐ABC (555555, BD Biosciences, 1:200), APC mouse anti‐human HLA‐DR (560896, BD Biosciences, 1:125), and BV421 rat anti‐mouse CD119 (740032, BD Biosciences, 1:200). Flow cytometry data were obtained on an LSRFortessa, and data analysis was achieved through FlowJo software (Tree Star).
6
Characterizing Macrophage Subsets
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Macrophages were harvested from 6-well plates, washed twice with PBS, made into single-cell suspensions, and then incubated with antibodies (FITC Mouse anti-Human CD68, PE Mouse Anti-human CD163, APC Mouse anti-Human HLA-DR, all from BD Biosciences, USA) for 60 min at 4 °C. The stained cells were then washed twice and resuspended in the 200 μl flow buffer for analysis on a FACS Calibur flow cytometer (BD Biosciences, USA) using FlowJo software (FlowJo, USA).
7
Differentiation of THP-1 Cells into Dendritic Cells
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THP-1 cells were cultured as reported in [25] to obtain a population of immature dendritic cells (iDCs). Briefly, THP-1 cells were seeded at a concentration of 1.75 x 10 5 cells/mL in 10% FBS-RPMI 1640 containing 100 ng/mL GM-CSF (Peprotech, RockyHill, CT, USA) and 100 ng/mL IL-4 (Sigma-Aldrich, St Louis, Mo, USA). THP-1 cells were cultured for 5 days and the medium was replaced every two days. iDCs were then cultured for supplementary 3 days in 50% of CM from siCOPZ1 or siNT TPC-1 cells, harvested 72 h after siRNA transfection. As a positive control for DC maturation, iDCs were cultured for 3 days in 10% FBS-RPMI 1640 containing 100 ng/mL GM-CSF, 200 ng/mL IL-4, 20 ng/mL TNF-α (Sigma-Aldrich) and 200 ng/mL ionomycin (Calbiochem, Merck, Germany). DCs were assessed by FACS for expression of differentiation and maturation markers [37] using the following antibodies: PE Mouse Anti-Human CD11c and BB700 Rat Anti-CD11b, BB515 Mouse Anti-Human CD80, BV650 Mouse Anti-Human CD86, BV421 Mouse Anti-Human CD83, APC Mouse Anti-Human HLA-DR (BD, Biosciences., San Jose, CA, USA). Cells were analyzed with a BD FACSCelesta™ instrument (BD Biosciences) and FlowJo software (TreeStar, Ashland, OR, USA).
8
Macrophage Immunophenotyping by Flow Cytometry
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Macrophages were harvested from 6-well plates, washed twice with PBS, made into single-cell suspensions, and then incubated with antibodies (FITC Mouse anti-Human CD68, PE Mouse Anti-human CD163, APC Mouse anti-Human HLA-DR, all from BD Biosciences, USA) for 60 min at 4°C. The stained cells were then washed twice and resuspended in the 200μl ow buffer for analysis on a FACS Calibur ow cytometer (BD Biosciences, USA) using FlowJo software (FlowJo, USA).
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