Journal: bioRxiv

Article Title: Adjuvant Discovery via a High Throughput Screen using Human Primary Mononuclear Cells

doi: 10.1101/2022.06.17.496630

Figure Lengend Snippet: Key resource table

Article Snippet: CD86 Antibody, anti-human, PE, REAfinityTM (Clone- REA968) , Miltenyi Biotec , Order Number: 130-116-160.

Techniques: Recombinant, Sterility, Injection, Modification, Staining, Saline, Enzyme-linked Immunosorbent Assay, Multiplex Assay, Software

Journal: Central European Journal of Immunology

Article Title: Macrophage M2 polarization induced by ANKRD22 in lung adenocarcinoma facilitates tumor angiogenesis

doi: 10.5114/ceji.2025.149372

Figure Lengend Snippet: Fig. 2. Cont. F-H) Flow cytometry was used to detect the effects of ANKRD22 overexpression or knockdown on the proportions of CD68+CD86+ and CD68+CD206+ macrophages. All experiments were conducted with 3 independent replicates. The data are presented as mean ± standard deviation. *p < 0.05

Article Snippet: The cells were incubated with PerCP Anti-Human CD68 Antibody (1 : 100; 333813, BioLegend, USA), PE Anti-Human CD86 Antibody (1 : 100; E-AB-F1012D, Elabscience, China), and/or FITC AntiHuman CD206/MMR Antibody (1 : 100; E-AB-F1161C, Elabscience, China) for 15 min. After washing with PBS, the cells were resuspended, and the flow cytometry data were collected and analyzed using Agilent’s flow analysis software.

Techniques: Flow Cytometry, Over Expression, Knockdown, Standard Deviation

Journal: Central European Journal of Immunology

Article Title: Macrophage M2 polarization induced by ANKRD22 in lung adenocarcinoma facilitates tumor angiogenesis

doi: 10.5114/ceji.2025.149372

Figure Lengend Snippet: Fig. 2. Cont. I) Flow cytometry was used to detect the ef- fects of ANKRD22 overexpression or knockdown on the pro- portions of CD68+CD86+ and CD68+CD206+ macrophages. J) WB was used to detect the effects of ANKRD22 over- expression or knockdown on the protein expression levels of CD80 and CD163. All experiments were conducted with 3 independent replicates. The data are presented as mean ± standard deviation. *p < 0.05

Article Snippet: The cells were incubated with PerCP Anti-Human CD68 Antibody (1 : 100; 333813, BioLegend, USA), PE Anti-Human CD86 Antibody (1 : 100; E-AB-F1012D, Elabscience, China), and/or FITC AntiHuman CD206/MMR Antibody (1 : 100; E-AB-F1161C, Elabscience, China) for 15 min. After washing with PBS, the cells were resuspended, and the flow cytometry data were collected and analyzed using Agilent’s flow analysis software.

Techniques: Flow Cytometry, Over Expression, Knockdown, Expressing, Standard Deviation

Journal: PLoS ONE

Article Title: Viscum album neutralizes tumor-induced immunosuppression in a human in vitro cell model

doi: 10.1371/journal.pone.0181553

Figure Lengend Snippet: CD14 + monocytes were matured to immature DC and incubated with medium (DC), maturation cocktail (DC Stim; 500 ng/mL LPS; 50 ng/mL TNF-alpha and 10 ng/mL IL-1beta), or VAEI (Iscador® Qu Spez; 0.5 μg/ml). After cultivation flow cytometric analysis of CD83 (A) , CD86 (B) and HLA-DR (C) expression was carried out. High CD83, CD86 and HLA-DR levels indicate DC maturation. MFI = Mean fluorescence intensity. Data and mean of 16 (DC Stim, CD83 and CD86), 13 (DC + VAEI, CD83 and CD86) or 5 (HLA-DR) individual experiments are presented in relation to untreated cells (DC = 100%). (D) The mediators IL-12p70, IL-6, IL-8 and TNFα were detected in the supernatants of cultured cells of 6 independent experiments and analyzed using cytokine bead array assay. Asterisks indicate significant differences between the groups (*P < 0.05, **P < 0.01, ***P < 0.001).

Article Snippet: The surface marker expression of dendritic cells was analyzed by microscopy after staining with anti-human HLA-DR Alexa 488 and anti-human CD86 Alexa 647 (both from Biolegend, London, UK) and putting the cells in a 96-well black clear bottom plate (BD Bioscience, Heidelberg, Germany).

Techniques: Incubation, Expressing, Fluorescence, Cell Culture

Journal: PLoS ONE

Article Title: Viscum album neutralizes tumor-induced immunosuppression in a human in vitro cell model

doi: 10.1371/journal.pone.0181553

Figure Lengend Snippet: CD14 + monocytes were matured to immature DC and incubated with medium (DC), maturation cocktail (DC Stim; 500 ng/mL LPS; 50 ng/mL TNF-alpha and 10 ng/mL IL-1beta), or VAEA (DC + VAEA; abnobaVISCUM® Fraxini; 0.06 μg/ml). After cultivation flow cytometric analysis of CD83 (A), CD86 (B) and HLA-DR (C) expression was carried out. High CD83, CD86 and HLA-DR levels indicate DC maturation. MFI = Mean fluorescence intensity. Data and mean of 16 (DC Stim, CD83 and CD86) and 6 (DC + VAEA, CD83 and CD86) or 5 (HLA-DR) individual experiments are presented in relation to untreated cells (DC = 100%). (D) The mediators IL-12p70, IL-6, IL-8 and TNFα were detected in the supernatants of cultured cells of 6 independent experiments and analyzed using cytokine bead array assay. Asterisks indicate significant differences between the groups (*P < 0.05, **P < 0.01, ***P < 0.001).

Article Snippet: The surface marker expression of dendritic cells was analyzed by microscopy after staining with anti-human HLA-DR Alexa 488 and anti-human CD86 Alexa 647 (both from Biolegend, London, UK) and putting the cells in a 96-well black clear bottom plate (BD Bioscience, Heidelberg, Germany).

Techniques: Incubation, Expressing, Fluorescence, Cell Culture

Journal: PLoS ONE

Article Title: Viscum album neutralizes tumor-induced immunosuppression in a human in vitro cell model

doi: 10.1371/journal.pone.0181553

Figure Lengend Snippet: CD14 + monocytes were matured to immature DC and incubated with medium (DC), maturation cocktail (DC Stim; 500 ng/mL LPS; 50 ng/mL TNF-alpha and 10 ng/mL IL-1beta), VAEI (DC + VAEI; Iscador® Qu Spez; 0.5 μg/ml) or VAEA (DC + VAEA; abnobaVISCUM® Fraxini; 0.06 μg/ml). After cultivation automated fluorescence microscopy and analysis of HLA-DR (A) and CD86 (B) expression was carried out. High HLA-DR and CD86 levels indicate DC maturation. MFI = Mean fluorescence intensity. Data and mean of 4 individual experiments are presented in relation to untreated cells (DC = 100%). Asterisks indicate significant differences between the groups (*P < 0.05, **P < 0.01, ***P < 0.001).

Article Snippet: The surface marker expression of dendritic cells was analyzed by microscopy after staining with anti-human HLA-DR Alexa 488 and anti-human CD86 Alexa 647 (both from Biolegend, London, UK) and putting the cells in a 96-well black clear bottom plate (BD Bioscience, Heidelberg, Germany).

Techniques: Incubation, Fluorescence, Microscopy, Expressing

Journal: PLoS ONE

Article Title: Viscum album neutralizes tumor-induced immunosuppression in a human in vitro cell model

doi: 10.1371/journal.pone.0181553

Figure Lengend Snippet: Dendritic cells derived from CD14 + monocytes were cultured in medium alone (DC) or supplemented with VAEI (DC + VAEI; Iscador® Qu Spez; 0.5 μg/ml), ML-depleted VAEI (DC + VAEI-ML; Iscador®; 0.66 μg/ml) or VAEI and anti-ML antibody (DC + VAEI + aML-Ab). DC maturation was assessed by flow cytometric analysis of CD83 (A) , CD86 (B) and HLA-DR (C) expression. MFI = Mean fluorescence intensity. Data and mean of 13 (DC + VAEI; CD83 and CD86), 3 (DC + VAEI-ML and DC + VAEI + aML-Ab; CD83 and CD86) or 5 (HLA-DR) individual experiments are represented in relation to untreated cells (DC = 100%) or VAEI-treated DC (DC + VAEI = 100%). Asterisks indicate significant differences between the groups (*P < 0.05, **P < 0.01, ***P < 0.001).

Article Snippet: The surface marker expression of dendritic cells was analyzed by microscopy after staining with anti-human HLA-DR Alexa 488 and anti-human CD86 Alexa 647 (both from Biolegend, London, UK) and putting the cells in a 96-well black clear bottom plate (BD Bioscience, Heidelberg, Germany).

Techniques: Derivative Assay, Cell Culture, Expressing, Fluorescence

Journal: PLoS ONE

Article Title: Viscum album neutralizes tumor-induced immunosuppression in a human in vitro cell model

doi: 10.1371/journal.pone.0181553

Figure Lengend Snippet: Dendritic cells derived from CD14 + monocytes were stimulated with maturation cocktail (DC Stim; 500 ng/mL LPS; 50 ng/mL TNF-alpha and 10 ng/mL IL-1beta) and incubated with IL-6 as an inhibition positive control (DC Stim + IL-6; 12.5 ng/ml). Unstimulated DC were further treated with 10% RPMI 1640 medium (DC + Medium), 10% tumor supernatant of an RCC line (DC + TS) or 10% tumor supernatant and VAEI (DC + TS + VAEI; Iscador® Qu Spez; 0.5 μg/ml) or VAEA (DC + TS + VAEA; abnobaVISCUM® Fraxini; 0.06 μg/ml). DC maturation was assessed by flow cytometric analysis of CD83 (A) and CD86 (B) surface marker expression. MFI = Mean fluorescence intensity. Data and mean of 7 (IL-6 and DC + TS + VAEA), 16 (DC + TS) or 9 (DC + TS + VAEI) individual experiments are presented in relation to stimulated cells (DC Stim = 100%), stimulated DC cultured in medium alone (DC + Medium = 100%) or stimulated cells cultured with tumor supernatant (DC + TS = 100%). Asterisks indicate significant differences between the groups (*P < 0.05, **P < 0.01, ***P < 0.001).

Article Snippet: The surface marker expression of dendritic cells was analyzed by microscopy after staining with anti-human HLA-DR Alexa 488 and anti-human CD86 Alexa 647 (both from Biolegend, London, UK) and putting the cells in a 96-well black clear bottom plate (BD Bioscience, Heidelberg, Germany).

Techniques: Derivative Assay, Incubation, Inhibition, Positive Control, Marker, Expressing, Fluorescence, Cell Culture

Journal: PLoS ONE

Article Title: Viscum album neutralizes tumor-induced immunosuppression in a human in vitro cell model

doi: 10.1371/journal.pone.0181553

Figure Lengend Snippet: Dendritic cells derived from CD14 + monocytes were incubated with 10% tumor supernatant (DC + TS). Further treatment of DC + TS was carried out with VAEI (DC + TS + VAEI; Iscador® Qu Spez; 0.5 μg/ml), ML-depleted VAEI (DC + TS + VAEI-ML; Iscador®; 0.66 μg/ml), or VAEI and anti-ML antibody (DC + TS + VAEI + aML-Ab; 2.5 μg/ml). DC maturation was assessed by flow cytometric analysis of CD83 (A), CD86 (B) and HLA-DR (C) expression. MFI = Mean fluorescence intensity. Data and mean of 12 (DC + TS + VAEI), 3 (DC + TS + VAEI-ML and DC + TS + VAEI + aML-Ab) or 5 (HLA-DR) individual experiments are presented in relation to stimulated and TS-treated cells (DC + TS = 100%), or TS and VAEI-treated DC (DC + TS + VAEI = 100%). Asterisks indicate significant differences between the groups (*P < 0.05, **P < 0.01, ***P < 0.001).

Article Snippet: The surface marker expression of dendritic cells was analyzed by microscopy after staining with anti-human HLA-DR Alexa 488 and anti-human CD86 Alexa 647 (both from Biolegend, London, UK) and putting the cells in a 96-well black clear bottom plate (BD Bioscience, Heidelberg, Germany).

Techniques: Derivative Assay, Incubation, Expressing, Fluorescence

Journal: PLoS ONE

Article Title: Viscum album neutralizes tumor-induced immunosuppression in a human in vitro cell model

doi: 10.1371/journal.pone.0181553

Figure Lengend Snippet: Unstimulated CD14 + -derived DC were treated with 10% RPMI 1640 medium (DC + Medium), 10% tumor supernatant of an RCC line (DC + TS) or 10% tumor supernatant and VAEI (DC + TS + VAEI; Iscador® Qu Spez; 0.5 μg/ml), ML-depleted VAEI (DC + TS + VAEI-ML; Iscador®; 0.66 μg/ml), or VAEI and anti-ML antibody (DC + TS + VAEI + aML-Ab; 2.5 μg/ml). After cultivation automated fluorescence microscopy and quantitative analysis of HLA-DR (A) and CD86 (B) expression was carried out. MFI = Mean fluorescence intensity. Data and mean of 4 individual experiments are presented in relation to untreated cells (DC = 100%). Asterisks indicate significant differences between the groups (*P < 0.05, **P < 0.01, ***P < 0.001).

Article Snippet: The surface marker expression of dendritic cells was analyzed by microscopy after staining with anti-human HLA-DR Alexa 488 and anti-human CD86 Alexa 647 (both from Biolegend, London, UK) and putting the cells in a 96-well black clear bottom plate (BD Bioscience, Heidelberg, Germany).

Techniques: Derivative Assay, Fluorescence, Microscopy, Expressing