1
Supporting Information to the „In vitro study on the immunomodulatory effects of differently functionalized silver nanoparticles on human
peripheral blood mononuclear cells“
Figure S1. Gating strategy for analysis of B- and NK-cell after activation with PLL-AgNPs (2 mg Ag/L, 6 h). First, all lymphocytes are gated in FSC-A / SSC-A dot-plot according to their size and inner complexity (a), and then all doublets (b) and dead cells (c) are excluded. After that, the gate is set on all B-cells (CD19+) (d). Activated B-cells (CD19+/CD69+) are gated in dot plot quadrant Q2 (e) and presented in histogram (f). Again, the gate is set on live CD45+
lymphocytes (g). All NK-cells (CD16+56+/CD3-) are gated in dot plot quadrant Q1 (h), and then activated NK-cells (CD16+56+/CD69+) are presented in histogram (i).
2 Figure S2. Gating strategy example for B- and NK-cell analysis after PMA/ionomicin
stimulation. First, all lymphocytes are gated in FSC-A / SSC-A dot-plot according to their size and inner complexity (a), and then all doublets (b) and dead cells (c) are excluded. After that, the gate is set on all B-cells (CD19+) (d). Activated B-cells (CD19+/CD69+) are gated in dot plot quadrant Q2 (e) and presented in histogram (f). Again, the gate is set on live CD45+
lymphocytes (g). All NK-cells (CD16+56+/CD3-) are gated in dot plot quadrant Q1 (h), and then activated NK-cells (CD16+56+/CD69+) are presented in histogram (i).
3 Figure S3. Gating strategy example for T-cell subset analysis after activation with PLL- AgNPs (1 mg Ag/L, 24 h). First, all lymphocytes are gated in FSC-A / SSC-A dot-plot according to their size and inner complexity (a), and then all doublets (b) and dead cells (c) are excluded. After that, the gate is set on all T-cells (CD3+) (d). In that gate, activated T-cells (CD3+/CD69+) are presented in histogram (i) and T-helper cells (CD3+/CD4+) and T-
cytotoxic cells (CD3+/CD8+) are separated by dot plot quadrants Q3 and Q1, respectively (f).
Activated CD69+ T-helper cells (g) and activated T-cytotoxic cells (h) are presented in histograms.
4 Figure S4. Gating strategy example for T-cell subset analysis after PMA/ionomicin
stimulation. First, all lymphocytes are gated in FSC-A / SSC-A dot-plot according to their size and inner complexity (a), and then all doublets (b) and dead cells (c) are excluded. After that, the gate is set on all T-cells (CD3+) (d). In that gate, activated T-cells (CD3+/CD69+) are presented in histogram (i) and T-helper cells (CD3+/CD4+) and T-cytotoxic cells (CD3+/CD8+) are separated by dot plot quadrants Q3 and Q1, respectively (f). Activated CD69+ T-helper cells (g) and activated T-cytotoxic cells (h) are presented in histograms.
5 Figure S5. Gating strategy example for monocyte cell subset analysis after activation with BSA-AgNPs (1 mg Ag/L, 24 h). First, all monocytes are gated in FSC-A / SSC-A dot-plot according to their size and inner complexity (a), and then all doublets (b) are excluded. The gate is set on all CD14+ monocytes (c). Activated monocytes (CD14+/CD25+/HLA-DR+) are separated in dot plota quadrant Q2. The same gating strategy (a-d) is applied on untreated cells as negative control (e-h).
