Human peripheral blood samples were obtained from healthy volunteer blood donors. All individuals provided their informed consent. Human peripheral blood mononuclear cells (PBMCs) were separated from heparinized peripheral blood by Ficoll–Hypaque density gradient centrifugation, and then washed twice with PBS. Next, PBMCs were re-suspended at 1 × 106 cells/mL in RPMI 1640 (Corning) containing 10% FBS, and cultured in the presence of anti-CD3 antibody (500 ng/mL, Miltenyi, Germany), IFN-γ (100 U/mL, Shanghai Kaimao, China), Polyhydroxyalkanoates (PHA)(10 μg/mL, Huizhou Hongyu, China) and recombinant human interleukin-2 (IL-2)(1000 U/mL, Beijing Sihuan, China) in culture flask for two days. Following this, Fresh medium containing IL-2 and anti-CD3 antibody was replenished every two or three days during culture. Cells were expanded over 3 weeks of time period. Phenotypes of CIK cells were weekly analyzed with a FACSCalibur flow cytometry (BD Biosciences). The following monoclonal antibodies (mAb) were used: CD3-FITC, CD4-FITC, CD8-PE, CD56-APC and CD314-APC (anti-NKG2D) (Miltenyi, Germany).
Cd314 apc anti nkg2d
Manufactured by Miltenyi Biotec
Sourced in Germany
CD314-APC (anti-NKG2D) is a flow cytometry reagent that detects the expression of the NKG2D receptor on cell surfaces. NKG2D is a type II transmembrane protein that functions as an activating receptor expressed on natural killer (NK) cells, CD8+ T cells, and other immune cell types. The CD314-APC conjugate enables the identification and analysis of NKG2D-expressing cells using flow cytometry.
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Lab products found in correlation
2 protocols using cd314 apc anti nkg2d
1
Expansion of Cytokine-Induced Killer Cells
2
Expansion and Characterization of CIK Cells
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Human peripheral blood samples were obtained from healthy donors. All individuals provided their informed consent. PBMCs were separated from heparinized peripheral blood samples by Ficoll-Hypaque density gradient centrifugation and subsequently washed twice with PBS. Next, PBMCs were cultured overnight in cell culture flasks at a cell density of 2 × 106 cells/mL in RPMI 1640 (Corning) supplemented with 10% FBS in 100 U/mL IFNγ (Shanghai Kaimao, China) and 10 μg/mL Polyhydroxyalkanoates (PHA) (Huizhou Hongyu, China). After 24 h in cultured at 37°C and 5% CO2, 50 ng/mL anti-CD3 antibody (Miltenyi, Germany) and 1000 U/mL recombinant human interleukin-2 (IL-2) (Beijing Sihuan, China) were added. Fresh medium with IL-2 and anti-CD3 antibody was added as needed. Cells were expanded over 3 weeks of time period. Cells were obtained from CIK cell cultures for phenotype analysis with the appropriated monoclonal antibodies (mAb), including CD3-FITC, CD4-FITC, CD8-PE, CD56-APC, and CD314–APC (anti-NKG2D) (Miltenyi, Germany) by standard flow cytometric assays (BD Biosciences).
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