NK cells isolated from PBMCs (AllCells) were cocultured with K562 cells for 4 h in the presence of different doses of BMK, IBI352g4a or hIgG control antibody at a 5:1 ratio and analyzed via a CytoTox 96® Non-Radioactive Cytotoxicity Assay (Sigma‒Aldrich).
Pbmcs
Manufactured by Merck Group
PBMCs are a type of lab equipment used for the isolation and purification of peripheral blood mononuclear cells (PBMCs) from whole blood samples. PBMCs include lymphocytes (T cells, B cells, and NK cells) and monocytes.
Automatically generated - may contain errors
Lab products found in correlation
Deoxyribonuclease 1, by Merck Group (1 mentions)
Easysep human nk cell enrichment kit, by STEMCELL (1 mentions)
Monensin, by BioLegend (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Pbmcs, by Thermo Fisher Scientific (1 mentions)
Dmem medium, by Thermo Fisher Scientific (1 mentions)
3 protocols using pbmcs
1
NK Cell-Mediated Cytotoxicity Assay
2
NK Cell Activation and Cytotoxicity Assay
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PBMCs (AllCells) were thawed and treated with deoxyribonuclease I (Sigma) for 15 min at 37 °C. NK cells were isolated from PBMCs by using an EasySep™ Human NK Cell Enrichment Kit (Stemcell, Vancouver, BC, Canada) and cultured for 16 h. After overnight incubation at 37 °C, the NK cells were washed and cocultured with K562 cells at a 5:1 ratio for 4 h in the presence of 10 μg/mL BMK, 10 μg/mL IBI352g4a or 10 μg/mL control hIgG antibody. PE-conjugated anti-human 107a antibody (BioLegend, cat. # 328,608) and monensin (BioLegend, cat. # 420,701) were added to each well. After culture, the cells were stained with BV421 anti-human CD3 (BioLegend, cat. # 300,434) and AF488 anti-human CD335 (NKp46) (BioLegend, cat. # 331,938) and analyzed by flow cytometry.
3
PBMC Stimulation and Anti-HBs Detection
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The optimal conditions for lymphocyte stimulation are presented in Livramento et al. (2013) . The PBMCs (Sigma-Aldrich) were isolated and purified using lymphocyte separation medium and heparinized venous blood. PBMCs were resuspended with DMEM medium (Gibco, Grand Island, NY, USA), supplemented with 10% fetal bovine serum (Gibco), 200 mM L-glutamine, 10 mM HEPES, 110 mg/mL sodium pyruvate, 50 U/mL penicillin, and 50 mg/mL streptomycin. PBMCs were then distributed into the wells of a 96-well cell culture plate at 4 x 10 6 cells/mL. The sample was stimulated with 50 ng/mL HBsAg under culture conditions at 37°C and 5% CO 2 according to a previously described method (Wang et al., 2004) . Cells were stimulated for 3 days and anti-HBs were detected by an ELISA quantitative detection kit (Sigma-Aldrich) in the media of the culture solution.
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