For phenotyping human circulating monocytes, peripheral blood mononuclear cells (PBMCs), which contain lymphocytes and monocytes, but no neutrophils, were obtained from the anticoagulated peripheral blood of TBI patients or patients who underwent cranioplasty by centrifugation with Ficoll-Paque Plus (Sigma-Aldrich, MO, USA). PBMCs were stained with anti-human CD16 (Clone: 3G8)-FITC (BioLegend, CA, USA, Cat: 302006), CD14 (Clone: MΦP9)-PE (Becton Dickinson, NJ, USA, Cat: 347497), and CXCR2 (Clone: 5E8/CXCR2)-APC (BioLegend, Cat: 320710) fluorescent monoclonal antibodies (mAb). For phenotyping monocytes in the CSF, 10 mL of anticoagulated CSF was obtained from TBI or non-TBI patients. Nucleated cells were collected by centrifugation at 150 g for 10 min, and then stained with anti-human CD11b (Clone: ICRF44)-FITC (BioLegend, Cat: 301330), CD14 (Clone: MΦP9)-PE (Becton Dickinson, Cat: 347497), and CXCR2 (Clone: 5E8/CXCR2)-APC (BioLegend, Cat: 320710) fluorescent mAb. To analyze surface calreticulin (CRT) expression, SH-SY5Y cells were stained with anti-human CRT (Clone: 1G6A7)-APC (Novus Biologicals, CO, USA, Cat: NBP1-47518APC). The isotype controls were analyzed in parallel. The samples were acquired on a FACSCalibur flow cytometer (BD Biosciences, NJ, USA) and analyzed using FlowJo software (FlowJo, OR, USA).
Anti human cd11b
Manufactured by BioLegend
Sourced in United States
Anti-human CD11b is a laboratory product used for the detection and analysis of CD11b, a cell surface molecule expressed on various immune cells, including monocytes, macrophages, and granulocytes. It can be used for applications such as flow cytometry and immunohistochemistry.
Automatically generated - may contain errors
Lab products found in correlation
Anti human cd16, by BioLegend (1 mentions)
Ficoll paque plus, by Merck Group (1 mentions)
Facscalibur flow cytometer, by BD (1 mentions)
Software, by FlowJo (1 mentions)
Fluorescein isothiocyanate (fitc), by BioLegend (1 mentions)
Anti human cd29, by BioLegend (1 mentions)
Apc conjugated anti human cd45, by BioLegend (1 mentions)
Anti human cd90, by BioLegend (1 mentions)
Ki 67, by Miltenyi Biotec (1 mentions)
Cytoflex, by BD (1 mentions)
Cd11b, by BioLegend (1 mentions)
Ifn γ, by BioLegend (1 mentions)
Cd206, by BioLegend (1 mentions)
Annexin 5 fitc apoptosis detection kit, by Dojindo Laboratories (1 mentions)
Anti cd11b bv421, by BioLegend (1 mentions)
Rbc lysis buffer, by Thermo Fisher Scientific (1 mentions)
Cytoflex s flow cytometer, by Beckman Coulter (1 mentions)
Mca2260, by Bio-Rad (1 mentions)
Nod cg prkdcscid il2rgtm1wjl szj nsg, by Jackson ImmunoResearch (1 mentions)
Anti human cd14, by BioLegend (1 mentions)
5 protocols using anti human cd11b
1
Phenotyping Peripheral Monocytes in TBI
2
Flow Cytometric Analysis of HDPSCs
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Flow cytometric analysis was carried out by entrusting a biotechnology company
(Jiamai Biotechnology Co., Ltd, Guangzhou, China). HDPSCs were washed and
collected into the flow tube. Then, PBS containing primary antibodies were added
into the sample for incubation for 2 h at room temperature in the dark:
fluorescein isothiocyanate (FITC)-conjugated anti-human Stro-1 (Proteintech,
China, FITC-65184), allophycocyanin (APC)-conjugated anti-human CD45 (Biolegend,
USA, 304011), or anti-human CD90 (Biolegend, USA, 328113) and phycoerythrin
(PE)-conjugated anti-human CD146 (Proteintech, China, PE-65181), anti-human CD29
(Biolegend, USA, 303003), anti-human CD11b (Biolegend, USA, 393111), or
anti-human CD34 (Biolegend, USA, 343505). No primary antibody incubated sample
was used as a negative control. The cell suspensions were washed twice,
resuspended in PBS, and analyzed with a Novocyte D2060R Flow Cytometer (Agilent,
USA). Novoexpress 1.5.6 (Novoexpress Software, USA) was employed for data
analysis and graph plotting.
(Jiamai Biotechnology Co., Ltd, Guangzhou, China). HDPSCs were washed and
collected into the flow tube. Then, PBS containing primary antibodies were added
into the sample for incubation for 2 h at room temperature in the dark:
fluorescein isothiocyanate (FITC)-conjugated anti-human Stro-1 (Proteintech,
China, FITC-65184), allophycocyanin (APC)-conjugated anti-human CD45 (Biolegend,
USA, 304011), or anti-human CD90 (Biolegend, USA, 328113) and phycoerythrin
(PE)-conjugated anti-human CD146 (Proteintech, China, PE-65181), anti-human CD29
(Biolegend, USA, 303003), anti-human CD11b (Biolegend, USA, 393111), or
anti-human CD34 (Biolegend, USA, 343505). No primary antibody incubated sample
was used as a negative control. The cell suspensions were washed twice,
resuspended in PBS, and analyzed with a Novocyte D2060R Flow Cytometer (Agilent,
USA). Novoexpress 1.5.6 (Novoexpress Software, USA) was employed for data
analysis and graph plotting.
3
Tumor Immune Profiling in EC Mouse Model
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Tumor tissues from EC cell‐bearing mice were removed, minced, and digested into single cells. The cells were resuspended in 100 μl PBS and incubated with the indicated Abs at 4°C for 40 min. Anti‐mouse MHC class II (BioLegend), CD11b (101263; BioLegend), CD86 (105032; BioLegend), CD206 (141708; BioLegend), CD3 (100235; BioLegend), CD8a (100713; BioLegend), γ‐interferon (IFN‐γ) (505808; BioLegend), anti‐human CD11b (101263; BioLegend), CD86 (305412; BioLegend), CD206 (321119; BioLegend), CD3 (344747; BioLegend), CD8 (344747; BioLegend), IFN‐γ (502528; BioLegend), p‐P65 (Cell Signaling Technology), and Ki‐67 (130–100–340; Miltenyi Biotec) were used. Cells were incubated with PBS twice before analysis using a flow cytometer (CytoFLEX; BD Biosciences). CytExpert analysis software (BD Biosciences) was used for the analysis.
4
Annexin V-FITC Apoptosis, Cell Cycle, and Differentiation Analysis
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The Annexin V-FITC Apoptosis Detection kit (Dojindo) was used for apoptosis assays. Cells were harvested and washed twice with cold PBS. After being suspended in 1 × Annexin V binding buffer, cells were stained with Annexin V-FITC and propidium iodide (PI) for 15 min on ice. For cell cycle assays, a DNA labeling solution (Cytognos) was used. Briefly, after collection and washing with cold PBS, cells were stained with DNA labeling solution for 15 min at 25 °C. For cell differentiation assays, human AML cells were harvested and washed with cold PBS; peripheral blood (PB), BM, and spleen cells were harvested from AML1-ETO9a AML mice, and erythrocytes were lysed using RBC Lysis Buffer (00-4333-57, Invitrogen). Human AML cells were stained with anti-human CD11b (301310, Biolegend) and isolated mice cells were stained with anti-CD11b-BV421 (101251, Biolegend) for 30 min on ice. Stained cells were analyzed using a CytoFLEX S Flow Cytometer (Beckman Coulter).
5
Xenograft Mouse Model for Rituximab
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All chemicals and proteins were purchased from Sigma-Aldrich (St. Louis, Missouri, USA) unless otherwise noted. All cell culture reagents were purchased from ThermoFisher Scientific (Waltham, Massachusetts, USA) unless otherwise noted. Hydrolysable crosslinker poly(DL-lactide)-b-poly(ethylene glycol)-b-poly(DL-lactide)-diacrylate triblock (PLA–PEG–PLA) was purchased from PolySciTech Akina (West Lafeyette, Indiana, USA). Capture antibody for ELISA against RTX was purchased from Bio-Rad Laboratories (MCA2260, Hercules, California, USA). HRP-conjugated goat antihuman IgG Fc for ELISA assay was purchased from ThermoFisher Scientific. antihuman CD45, antihuman CD3, antihuman CD56, antihuman CD11b, antihuman CD14, antihuman CD4, and antihuman CD8 were purchased from BioLegend (San Diego, California, USA). RTX (RITUXANTM: Genentech, San Francisco, California, USA) and HER (Herceptin: Genentech) were obtained at the UCLA hospital pharmacy. All NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) and NOD.Cg−Hc1Prkdcscid Il2rgtm1Wjl/SzJ (NSG−Hc1) mice were purchased from The Jackson Laboratory and housed in specific pathogen-free vivarium.
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