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. 2022 May 23;23(10):5857. doi: 10.3390/ijms23105857

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© 2022 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

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Phenotypic analysis of differentially expanded NK cells and fresh NK cells. Depicted is a representative flow cytometry analysis from expanded and fresh NK cells from one donor. As described in the material and methods section, NK cells were stained at day 14 after the start of expansion with an NKG2C-specific antibody and were simultaneously stained with indicated antibodies (a) specific for inhibitory receptors NKG2A, KIR2DL1/S1/S4, KIR2DL2/L3/S2 and KIR3DL1, (b) specific for activating receptors CD16, NKG2D and for the activation/proliferation marker CD25 as well as (c) specific for surface markers related to exhaustion/immune checkpoint and maturation (PD-1, LAG-3, CD57). (d) Quantitative analysis of marker expression in differentially expanded total NK cells and total fresh NK cells. For analysis of KIRs, CD16, NKG2D, CD25 and PD-1 six expansion experiments from six donors are included. For analysis of CD57 and LAG-3 four expansion experiments from three donors are included. Dots and the horizontal lines inside the Whisker boxes indicate the mean and the median. * p < 0.05, ** p < 0.01, *** p < 0.001.

Phenotypic analysis of differentially expanded NK cells and fresh NK cells. Depicted is a representative flow cytometry analysis from expanded and fresh NK cells from one donor. As described in the material and methods section, NK cells were stained at day 14 after the start of expansion with an NKG2C-specific antibody and were simultaneously stained with indicated antibodies (a) specific for inhibitory receptors NKG2A, KIR2DL1/S1/S4, KIR2DL2/L3/S2 and KIR3DL1, (b) specific for activating receptors CD16, NKG2D and for the activation/proliferation marker CD25 as well as (c) specific for surface markers related to exhaustion/immune checkpoint and maturation (PD-1, LAG-3, CD57). (d) Quantitative analysis of marker expression in differentially expanded total NK cells and total fresh NK cells. For analysis of KIRs, CD16, NKG2D, CD25 and PD-1 six expansion experiments from six donors are included. For analysis of CD57 and LAG-3 four expansion experiments from three donors are included. Dots and the horizontal lines inside the Whisker boxes indicate the mean and the median. * p < 0.05, ** p < 0.01, *** p < 0.001.