Hepatocytes and hepatic non-parenchymal cells were separated by centrifugation. After washing, live vs. dead non-parenchymal cells were assessed using Zombie Aqua fluorescent dye with Zombie Aqua™ Fixable Viability Kit (Biolegend, San Diego, CA). Cells were washed and blocked with CD16/CD32 antibody (Biolegend, San Diego, CA), followed by incubation with specific antibodies targeting surface proteins for 30 min. After washing, cells were fixed and permeabilized with BD Cytofix/Cytoperm™ Fixation/Permeabilization kit (BD Biosciences, San Jose, CA) according to the manufacturer’s introductions. Fixed cells were further incubated with a specific antibody targeting intracellular protein. After washing, resuspended labeled single cells were analyzed by flow cytometry in a BD LSRFortessa-X20™ (BD Biosciences, San Jose, CA). Detailed information for antibodies used for flow cytometry analysis is provided in Supplementary Table 1 (Table S1 ).
Cd16 cd32 antibody
Manufactured by BioLegend
Sourced in United States
The CD16/CD32 antibody is a laboratory tool used for the detection and analysis of immune cells expressing the CD16 and CD32 receptors. This antibody can bind to these receptors, allowing researchers to identify and study the presence and functionality of these cells in various biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Lsrfortessa x 20, by BD (1 mentions)
Zombie aqua fixable viability kit, by BioLegend (1 mentions)
Cytofix cytoperm fixation permeabilization kit, by BD (1 mentions)
Facscanto 2 cell analyzer, by BD (1 mentions)
Foxp3 transcription factor staining buffer set, by Thermo Fisher Scientific (1 mentions)
Facsaria fusion cell sorter, by BD (1 mentions)
Cd80 fitc, by BioLegend (1 mentions)
Cd11c apc, by BioLegend (1 mentions)
Expo32 adc software, by Beckman Coulter (1 mentions)
Epics xl adc, by Beckman Coulter (1 mentions)
4 protocols using cd16 cd32 antibody
1
Multiparametric Hepatic Cell Analysis
2
Comprehensive Immune Cell Analysis of White Adipose Tissue
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Flow cytometry analysis was performed using BD FACSCanto™ II Cell Analyzer (BD Biosciences), and ATMs were sort-purified using BD FACSAria™ Fusion Cell Sorter (BD Biosciences). After blocking the Fc receptors with CD16/CD32 antibody (Bio-Legend), cell suspensions of WAT were incubated on ice with fluorochrome-conjugated antibodies in FACS buffer. The following antibodies were used to stain cells from mouse tissues: CD45 (30-F11), CD25 (PC61), CD90.2 (30-H12), IL-33Rα/ST2 (DIH9), CD5 (53-7.3), CD19 (1D3/CD19), CD11b (M1/70), CD11c (N418), FcϵRI (MAR-1), NK1.1 (PK136), CD3ϵ (145-2C11), CD49b (DX5), SiglecF (E50-2440), CD31 (390), F4/80 (BM8), CD4 (RM4-5), CD8 (53-6.7), CD25 (PC61). Foxp3 (FJK-16S) was stained using the Foxp3 transcription factor staining buffer set (eBioscience). All antibodies used in flow cytometry were purchased from eBioscience, BioLegend, or BD Biosciences if not indicated otherwise above. Data were analyzed with FlowJo v10.4.1 software (TreeStar).
3
Investigating NP/mDs Modulation of mDCs
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CT26 cells were seeded in 6-well plates (2 × 105 cells/well) and incubated with 1.0 mL of complete medium for 24 h. The cells were then treated with NP/mDs and each component for another 24 h and washed thrice with PBS. Afterward, the imDCs were cultured with the above pre-treated CT26 cells for 24 h before being incubated with CD16/CD32 antibody (Biolegend, USA) to reduce nonspecific binding to FcRs, and then APC-CD11c, FITC-CD80, and PE-CD86 antibodies (Biolegend, USA). The frequency of mDCs was analyzed using flow cytometry.
4
Splenic T Cell Subpopulation Analysis
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Splenocytes were pre-incubated with anti-cluster of differentiation (CD)16/CD32 antibody (10139, Clone: 93, Biolegend) to block the Fc receptor and then were stained with monoclonal antibodies according to each manufacturer's instructions. Splenocytes were stained with fluorescein isothiocyanate-labelled anti-CD3 antibody (100306, Clone: 145-2C11; Biolegend), phycoerythrin-labelled anti-CD4 antibody (12-0042-82, Clone: RM4-5; eBioscience) and peridinin chlorophyll protein complex-labelled anti-CD8 antibody (100732, Clone: 53-6.7; Biolegend). CD3 þ CD4 þ cells were defined as helper T cells and CD3 þ CD8 þ cells were defined as cytotoxic T cells. Splenocyte subpopulations were analysed by flow cytometry (EPICS XL ADC using EXPO 32 ADC software; Beckman Coulter), and the absolute count of each subpopulation was calculated as the total splenocyte count multiplied by the percentage of each cell type.
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