A culture method for inducing NK cells in vitro after cryopreserved human pbmc recovery
A technology of NK cells and culture methods, which is applied in the field of in vitro induction of NK cells after recovery from cryopreserved human PBMCs. It can solve the problems of poor viability of NK cells, unstable induction methods, and difficulties in NK cells, and reduce the cost of cell culture. , non-toxic and side effects, and the effect of increasing the amplification factor
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Embodiment 1
[0061] Example 1 Cell Viability Measurement and Control Experiment
[0062] Collect human peripheral blood from healthy volunteers, separate peripheral blood mononuclear cells, and take out cell samples at the same time point, use the cell recovery method in the culture method for inducing NK cells in vitro after recovery of frozen human PBMCs of the present invention, perform cell recovery, and calculate the number of cells survival rate.
[0063] 1. Preparation of PBMCs
[0064] Carefully pour 20-30ml of blood on 15ml of lymphocyte separation medium; at room temperature, centrifuge at 800xg for 20min, slowly rise and fall, if the blood is stored for more than 2 hours, please increase the centrifugation time to 30min; after centrifugation, the blood is separated 4 layers consisting of plasma (upper layer), mononuclear cell layer between plasma and separation fluid (layer 2), separation fluid (layer 3) and erythrocyte layer (bottom layer); the upper layer of plasma is collect...
Embodiment 2
[0075] Collect 40ml of peripheral blood from healthy volunteers, separate peripheral blood mononuclear cells, and induce NK cells (abbreviated as the experimental group) by using the culture method for inducing NK cells in vitro after recovery of frozen human PBMCs of the present invention, observe the cell morphology at different time points, and take out Count the cell samples, detect the cell phenotype, and observe whether the induced NK cells can meet the requirements of clinical treatment.
[0076] 1. Preparation of PBMCs
[0077] Carefully pour 20-30ml of blood on 15ml of lymphocyte separation medium; at room temperature, centrifuge at 800xg for 20min, slowly rise and fall, if the blood is stored for more than 2 hours, please increase the centrifugation time to 30min; after centrifugation, the blood is separated It consists of 4 layers, consisting of plasma (upper layer), mononuclear cells between plasma and separation fluid (second layer), separation liquid (third layer...
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