Journal: Frontiers in immunology

Article Title: The significance of CD16+ monocytes in the occurrence and development of chronic thromboembolic pulmonary hypertension: insights from single-cell RNA sequencing.

doi: 10.3389/fimmu.2024.1446710

Figure Lengend Snippet: FIGURE 2 Changes in peripheral blood monocyte subsets in CTEPH patients. (A) UMAP plot of Mononuclear Phagocyte System (MPS) was divided into three cell types: monocytes, macrophages, and conventional dendritic cells (cDCs). Subsequently, monocytes were further divided into CD14+ monocytes and CD16+ monocytes. (B) A dot plot was used to annotate marker genes for CD14+ monocytes and CD16+ monocytes, along with a heatmap displaying the top 10 differentially expressed genes between the two cell types. (C) Ro/e (ratio of observed cell number to expected cell number) revealed the proportion of CD16+ monocytes in the CTEPH-N, CTEPH-I and HC groups. (D) The heat map showed the degree of correlation, and the numbers in the graph represented the correlation coefficient. (6MWD: 6-minute walk distance; PVR: pulmonary vascular resistance) (E) Schematic diagram of the gating strategy for distinguishing CD14+ monocytes from CD16+ monocytes by flow cytometry. (F) Differences in the proportion of CD16+ monocyte subsets in total monocytes between CTEPD patients (n = 15) and matched healthy controls (n = 15) were assessed by flow cytometry. All data were presented as means ± SEM, ***P < 0.001.

Article Snippet: Then the cells were resuspend in the flow cytometer wash buffer (2% FBS in PBS) and stained with the following antibodies according to the standard protocol: CD14-PE (Elabscience, E-ABF1209D) and CD16-FITC (Elabscience, E-AB-F1236C) for monocyte labeling.

Techniques: Marker, Cytometry

Journal: Frontiers in immunology

Article Title: The significance of CD16+ monocytes in the occurrence and development of chronic thromboembolic pulmonary hypertension: insights from single-cell RNA sequencing.

doi: 10.3389/fimmu.2024.1446710

Figure Lengend Snippet: FIGURE 3 The functional characteristics of peripheral blood CD16+ monocytes in CTEPH patients. (A) GO analysis (biological process) of upregulated genes in CD16+ monocytes compared with CD14+ monocytes in CTEPH patients. (B) KEGG analysis of upregulated genes in CD16+ monocytes compared with CD14+ monocytes in CTEPH patients. (C) GSEA bar plot of CD16+ monocytes versus CD14+ monocytes in patients with CTEPH. (D) GO analysis (biological process) of upregulated genes in CD16+ monocytes between the CTEPH and healthy control samples. (E) KEGG analysis of upregulated genes in CD16+ monocytes between CTEPH patients and healthy controls. (F) GSVA heatmap of CD16+ monocytes between CTEPH patients and healthy controls. (G) Heat map of transcription factors upregulated in CD16+ monocytes between CTEPH patients and healthy controls.

Article Snippet: Then the cells were resuspend in the flow cytometer wash buffer (2% FBS in PBS) and stained with the following antibodies according to the standard protocol: CD14-PE (Elabscience, E-ABF1209D) and CD16-FITC (Elabscience, E-AB-F1236C) for monocyte labeling.

Techniques: Functional Assay, Control

Journal: Frontiers in Immunology

Article Title: High Expression of IL-36γ in Influenza Patients Regulates Interferon Signaling Pathway and Causes Programmed Cell Death During Influenza Virus Infection

doi: 10.3389/fimmu.2020.552606

Figure Lengend Snippet: Elevated expression profile of IL-36 receptors in PBMCs of influenza-induced ARDS patients. (A) The relative mRNA expression levels of IL-36R and IL-1RAcp in PBMCs of influenza-induced ARDS patients ( n = 18) and healthy individuals ( n = 12) were detected by RT-PCR. (B) Flow cytometry to reveal IL-36R expression on the surface of T cells (CD4 + and CD8 + ), B cells (CD19 + ), monocytes (CD14 + ), and myeloid dendritic cells (mDCs, CD11c + HLA-DR + ) in healthy individuals and histograms are representative of healthy donors. Filled histogram: anti-IL-36R; Dotted histogram: isotype control Ab. The proportion of (C) IL-36R + B cells in total B cells, (D) IL-36R + monocytes in total monocytes, and (E) IL-36R + mDCs in total mDCs from patients ( n = 18) and healthy individuals ( n = 12) were detected using flow cytometry. (C–E) The expression level of IL-36R on B cells, monocytes, and mDCs from patients ( n = 18) and healthy individuals ( n = 12) were detected using flow cytometry. Results are presented as scatter plots with median of the proportion or mean fluorescence intensity (MFI) subtracting corresponding isotypic controls. Statistical significance is determined by Student’s t test. *P < 0.05, ***P < 0.001, ns, not significant.

Article Snippet: Then cells were washed and incubated with APC streptavidin antibody (TONBO biosciences), APC-H7 mouse anti-human CD3 Ab (BD PharmingenTM), BV605 mouse anti-human CD4 Ab, BV510 mouse anti-human CD8 Ab, BV650 mouse anti-human CD19 Ab (BD HorizonTM), RedFluorTM 710 anti-human CD14 Ab, VioletFluorTM 450 anti-human CD11c Ab, FITC anti-human HLA-DR Ab (TONBO biosciences), or parallel isotype antibodies at 4°C for 30 min in the dark.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Flow Cytometry, Control, Fluorescence

Journal: Cell reports

Article Title: High-dimensional profiling clusters asthma severity by lymphoid and non-lymphoid status

doi: 10.1016/j.celrep.2021.108974

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Anti-Human CD14 (M5E2)-175Lu , Fluidigm , Cat# 3175015B.

Techniques: Purification, Control, Antibody Labeling, Flow Cytometry, Software, Staining, Blocking Assay

Journal: PLoS ONE

Article Title: Glatiramer Acetate Increases Phagocytic Activity of Human Monocytes In Vitro and in Multiple Sclerosis Patients

doi: 10.1371/journal.pone.0051867

Figure Lengend Snippet: A–D. Immunocytochemistry showed distinct phagocytosis of polystyrene beads (green) by monocytes treated with 31.25 µg/ml glatiramer acetate (GA) for 24 h (red, labelled with anti-CD14 antibody) as compared to medium, human serum albumin (HA), and mannitol (Mt) vehicle control. Scale bar: 50 µm. E. Representative z-stack series of confocal microscopic images demonstrate that the particles were completely internalised and not merely attached to the outer membrane. Scale bar: 10 µm.

Article Snippet: After 30 minutes incubation with fluorescent latex beads (FluoresbriteTM YG carboxylate microspheres; 1 μm diameter; Polysciences, Eppelheim, Germany) at 37°C, cover slips were washed three times and cells were stained with the CD14 antibody (primary: rabbit anti-human IgG, Abcam, Cambridge, UK; secondary: goat anti-rabbit IgG, AbD Serotec, Düsseldorf, Germany) at 1∶100 dilution in DMEM+.

Techniques: Immunocytochemistry, Control, Membrane

Journal: PLoS ONE

Article Title: Glatiramer Acetate Increases Phagocytic Activity of Human Monocytes In Vitro and in Multiple Sclerosis Patients

doi: 10.1371/journal.pone.0051867

Figure Lengend Snippet: A–B. Gated monocytes in a forward vs. side scatter dot-plot analysis of peripheral blood mononuclear cells. Monocytes were then gated according to their surface expression of CD14 and CD16. Flow cytometric analysis of phagocytosis revealed that CD14 ++ CD16 + monocytes (gate III) engulfed polystyrene beads more effectively than the other subsets (gate I and II). CI–III. The mean fluorescence intensities represent the amount of incorporated fluorescent latex particles phagocytosed by 3×10 5 cells. D. Increase in the percentage of CD14 + CD16 + monocytes after 4 h and 8 h of treatment with 31.25 µg/ml glatiramer acetate (GA) in MACS isolated monocytes. Data are expressed as mean percentages ± SEM of three independent experiments. Significant effects vs. controls are indicated by asterisks (*p<0.05, **p<0.01, and ***p<0.001 using Bonferroni's Multiple Comparison Test) as determined by one-way ANOVA. E. Slight but not significant increase of CD16 expression after GA treatment.

Article Snippet: After 30 minutes incubation with fluorescent latex beads (FluoresbriteTM YG carboxylate microspheres; 1 μm diameter; Polysciences, Eppelheim, Germany) at 37°C, cover slips were washed three times and cells were stained with the CD14 antibody (primary: rabbit anti-human IgG, Abcam, Cambridge, UK; secondary: goat anti-rabbit IgG, AbD Serotec, Düsseldorf, Germany) at 1∶100 dilution in DMEM+.

Techniques: Expressing, Fluorescence, Isolation, Comparison

Journal: PLoS ONE

Article Title: Glatiramer Acetate Increases Phagocytic Activity of Human Monocytes In Vitro and in Multiple Sclerosis Patients

doi: 10.1371/journal.pone.0051867

Figure Lengend Snippet: Mean fluorescence intensities of CD36, CD210, and TIM-3 (± SEM) of three independent experiments after a treatment period of 24 h according to the different monocyte subsets, i.e. CD14 + CD16 − and CD14 + CD16 + monocytes.

Article Snippet: After 30 minutes incubation with fluorescent latex beads (FluoresbriteTM YG carboxylate microspheres; 1 μm diameter; Polysciences, Eppelheim, Germany) at 37°C, cover slips were washed three times and cells were stained with the CD14 antibody (primary: rabbit anti-human IgG, Abcam, Cambridge, UK; secondary: goat anti-rabbit IgG, AbD Serotec, Düsseldorf, Germany) at 1∶100 dilution in DMEM+.

Techniques: Fluorescence

Journal: PLoS ONE

Article Title: Glatiramer Acetate Increases Phagocytic Activity of Human Monocytes In Vitro and in Multiple Sclerosis Patients

doi: 10.1371/journal.pone.0051867

Figure Lengend Snippet: The MFI displays the amount of incorporated beads phagocytosed by 3×10 5 cells. Data are expressed as means of the mean fluorescence intensities (MFI) ± SEM of three independent experiments. Pre-treatment of monocytes with anti-CD14, anti-CD16, anti-CD32, anti-CD210, and anti-TIM3 antibodies reduced glatiramer acetate (GA) induced phagocytosis of polystyrene beads. The anti-CD36 and, at the highest concentration, the anti-IL10 antibody most effectively suppressed phagocytosis. Significant effects vs. GA 31.25 µg/ml are indicated by asterisks (*p<0.05, **p<0.01, and ***p<0.001 using Bonferroni's Multiple Comparison Test) as determined by one-way ANOVA. B–C. The anti-CD36 antibodies of two different clones show a diverse impact on the phagocytosis of polystyrene beads by untreated cells, whereas both antibodies effectively decreased the GA induced phagocytosis.

Article Snippet: After 30 minutes incubation with fluorescent latex beads (FluoresbriteTM YG carboxylate microspheres; 1 μm diameter; Polysciences, Eppelheim, Germany) at 37°C, cover slips were washed three times and cells were stained with the CD14 antibody (primary: rabbit anti-human IgG, Abcam, Cambridge, UK; secondary: goat anti-rabbit IgG, AbD Serotec, Düsseldorf, Germany) at 1∶100 dilution in DMEM+.

Techniques: Fluorescence, Concentration Assay, Comparison, Clone Assay

Journal: Journal of Clinical Laboratory Analysis

Article Title: High‐dimensional single‐cell proteomics analysis reveals the landscape of immune cells and stem‐like cells in renal tumors

doi: 10.1002/jcla.23155

Figure Lengend Snippet: Purified antibodies about the immune cell‐centric panel

Article Snippet: CD14 , 148Nd , RMO52 , Fluidigm , 3148010B.

Techniques: Purification