Journal: iScience

Article Title: Hypoxic-immune axis orchestrates metastatic dissemination via HIF isoform imbalance in pancreatic neuroendocrine tumors

doi: 10.1016/j.isci.2025.114340

Figure Lengend Snippet: MDSC subsets in KRAS-mutated PNETs (A) Representative flow cytometry plots and histogram overlays demonstrate elevated CD11b + cell proportions in KRAS G12C patient blood samples compared to wild-type KRAS tumors and healthy controls. (B) Quantification shows a slight elevation in CD14 + cell frequency compared to wild-type KRAS, alongside a marked enrichment relative to healthy controls. (C) A moderate rise in CD15 + cell frequency in KRAS G12C samples versus wild-type KRAS, with a pronounced elevation compared to normal baseline levels. (D) Statistical comparisons confirm a significant increase in CD33 + cell populations relative to both wild-type KRAS and healthy controls.

Article Snippet: PerCP/Cyanine5.5 Anti-Human CD14 Antibody [M5E2] , Elabscience , E-AB-F1209J.

Techniques: Flow Cytometry

Journal: Mucosal immunology

Article Title: Th22 cells are efficiently recruited in the gut by CCL28 as an alternative to CCL20 but do not compensate for the loss of Th17 cells in treated HIV-1-infected individuals.

doi: 10.1038/s41385-020-0286-6

Figure Lengend Snippet: Fig. 3 Impact of the interactions between IL-17A/Th17 cells and IL-22/Th22 cells on CCL20 and CCL28 expression by enterocytes. Effect of IL-17A (white bars) and IL-22 (gray bars) on a CCL20 and b CCL28 mRNA expression by enterocytes. Monolayers of differentiated human primary enterocytes on transwell inserts were stimulated by 0.5, 5, and 50 ng/mL of cytokine. CCL20 and CCL28 mRNA was quantified in the enterocytes by qRT-PCR. CCL20 and CCL28 expression following cytokine stimulation was normalized relatively to the expression in unstimulated epithelial cells (set to 1) and expressed as fold change (log2 scale). Presented data were obtained from at least eight independent experiments performed with different donors. Cuzick’s test for trend was used to compare chemokine expression across the increasing concentrations of cytokines (the corresponding P value is shown on the line above the IL17-A and IL-22 bars); paired Wilcoxon’s test was used to compare chemokine expression upon stimulation vs. unstimulated condition (intradonor pairing); P value is shown above each bar; *P < 0.05; **P < 0.01. Means and SEM are shown. c Effect of Th22:IEC coculture on CCL20 and CCL28 mRNA expression by enterocytes. Enterocytes were cocultured with FACS-sorted Th22 or Th17 cells, added in the bottom chamber for 15 h. CCL20 and CCL28 mRNA were quantified in the enterocytes by qRT-PCR. CCL20 and CCL28 expression in Th22:IEC coculture was normalized relatively to the expression in Th17:IEC coculture (set to 1) and expressed as fold change (log2 scale). Enterocytes production of d CCL20 and e CCL28 proteins in Th22:IEC and Th17:IEC cocultures. CCL20 and CCL28 were quantified in the bottom chamber by ELISA. Presented data were obtained from ten independent experiments performed with different donors. Paired Wilcoxon’s test was used to compare CCL20 and CCL28 in Th22:IEC vs. Th17:IEC cocultures (intradonor pairing); *, P < 0.05. IEC, intestine epithelial cells. Means and SEM are shown.

Article Snippet: Quantification of sCD14 and sCD163 Soluble CD14 (sCD14) and sCD163 were quantified in plasma by ELISA (DC140, R&D systems; IQP-383, Eurobio, respectively).

Techniques: Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

Journal: Cell Death & Disease

Article Title: SMAC mimetic drives microglia phenotype and glioblastoma immune microenvironment

doi: 10.1038/s41419-024-07056-z

Figure Lengend Snippet: A Representative experiment of 3 independent experiments of IAP expression levels analyzed by Western blotting after 72 h of vehicle or SMg treatment in C8B4 microglia cell line. B Quantification of GL261-DsRed spheroids area upon vehicle ( n = 10) and SMg treatment ( n = 9) and in the presence or not of the C8B4 cells ( n = 10). Statistical analyses were performed by using ANOVA test and ANOVA post-hoc Tukey test; alpha = 0.05, bilateral p -value: *** p < 0.0005. C Concentrations of CD14 and CCL17/TARC quantified in the supernatant of C8B4 and GL261-DsRed co-cultures by ELISA assay after 72 h of vehicle or SMg treatments ( n = 6 independent experiments). Statistical analyses were performed by using Mann–Whitney test; alpha = 0.05, bilateral p-value: ** p < 0.005. D Quantification of GL261-DsRed spheroids area in the presence of the C8B4 cells expressing ML-IAP (siCTRL) or down-expressing ML-IAP (siML-IAP) ( n = 10) after 72 h of treatment vehicle ( n = 10) and SMg treatment ( n = 9). Data from three independent experiments. Statistical analyses were performed by using ANOVA test and ANOVA post-hoc Tukey test; alpha = 0.05, bilateral p -value: * p < 0.05 ; ** p < 0.005 ; **** p < 0.0001. E Quantification of GL261-DsRed spheroid area in the presence of C8B4 cells treated with vehicle and SMg for 72 h. C8B4 cells were pre-treated for 24 h with ZVAD (vehicle n = 23, SMg n = 23) and TNFαi (vehicle n = 27, SMg n = 23) or without pre-treatment (vehicle n = 28, SMg n = 25). Statistical analyses were performed by using ANOVA test and ANOVA post-hoc Tukey test; alpha = 0.05, bilateral p -value: *** p < 0.0005. F Representative experiment ( n = 2) of IAP expression levels analyzed by western blotting after 72 h of vehicle or SMg treatment in C8B4 microglia cell line. C8B4 cells were pre-treated for 24 h with ZVAD and TNFαi. G Representative experiment ( n = 2) of expression levels of p65, phospho-p65, IκBα, phospho IκB, caspase-3, cleaved caspase-3 analyzed by Western blotting in C8B4 microglia cell line after 24 h of vehicle or SMg treatment. H Representative experiment out of 3 experiments of expression levels of CD206 and iNOS analyzed by Western blotting after 24 h of vehicle or SMg treatment. C8B4 cells were pre-treated for 24 h with ZVAD and TNFαi. Expression level of actin β served as loading control. I iNOS/CD206 ratio in C8B4 microglia cell line after 24 h of vehicle or SMg treatment. C8B4 cells were pre-treated for 24 h with ZVAD and TNFαi. Quantification was performed from 3 independent experiments using ImageJ software and data presented were normalized to actin β expression. iNOS/CD206 ratio fold changes were normalized on vehicle condition. B – E , I Bar graphs represent mean ± s.e.m.

Article Snippet: Supernatants were collected after 72 h of treatment and processed using the proteome profiler human XL cytokine array kit (R&D systems, #ARY022B) or the ELISA kits Quantikine ELISA Mouse CCL17/TARC Immunoassay (R&D systems, MCC170) and Mouse CD14 Immunoassay (R&D systems, MC140) accordingly to the manufacturer protocol (Data represented were normalized to reference spot expression in array membrane).

Techniques: Expressing, Western Blot, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY, Control, Software

Journal: bioRxiv

Article Title: IFI16/AIM2 inflammasomes control Gal-9 and PVR in myeloid cells from PWH and their targeting improves immunotherapy against HIV-1

doi: 10.64898/2026.01.26.701473

Figure Lengend Snippet: (A): ELISA analysis of plasma concentration of IL-1β, comparing PWH with normal CD4/CD8 T cell ratio (defined as ≥1) and low CD4/CD8 T cell ratio (defined as < 0.8). Statistical significance was calculated using two-tailed Wilcoxon test. (B): Spearman correlation between plasma concentrations of soluble CD14 (ssCD14) and IL-1β. P and R values are shown (C): Expression of Gal-9 (Gal9; higher plot) or PVRhi (lower plot) in activated CD40hi in Mo from HIV- 1-negative controls, and PWH with low CD4/CD8 T cell ratio or normal CD4/CD8 T cell ratio after 16-hour stimulation with PAMPs mimicking bacterial translocation (LPS, red; Flagellin, Flag, purple) or viral replication (CL097, ssRNA, orange; Poly I:C, dsRNA, blue). Statistical significance was calculated using Kruskal-Wallis test for comparison of stimulation with the same PAMP between groups (*p<0.05; **p<0.01), and One-Way ANOVA Friedman test with Dunn’s multiple comparison test for comparison within the same group (*p<0.05; **p<0.01; ***p<0.001; ****p<0.0001).

Article Snippet: Plasma levels of soluble CD14, IL-6 and IL-1β were determined using the Human CD14 DuoSet ELISA, the Human IL-6 DuoSet ELISA kits (both from R&D systems) or the Quantikine ELISA Human IL- 1β/IL-1F2 kit (R&D systems), respectively.

Techniques: Enzyme-linked Immunosorbent Assay, Clinical Proteomics, Concentration Assay, Two Tailed Test, Expressing, Translocation Assay, Comparison

Journal: bioRxiv

Article Title: IFI16/AIM2 inflammasomes control Gal-9 and PVR in myeloid cells from PWH and their targeting improves immunotherapy against HIV-1

doi: 10.64898/2026.01.26.701473

Figure Lengend Snippet: (A-B): Histological analysis of IFI-16 (A) or AIM2 (B) (green) and Caspase-1 (red) expression and DAPI (blue) in the spleen from a viremic HIV-1 infected humanized BLT mouse. Quantifications of cells positive cells for each inflammasome sensor alone (top plots), co-expressing Caspase 1 (CASP1) (middle plots) or both Caspase 1 and CD14 (bottom plots) are shown on the right. (C): Histological analysis of Gal-9 (Gal9; green), CD14 (white) and Caspase-1 (Casp1, red) expression and DAPI (blue) in the spleen from a viremic HIV-1 infected humanized BLT mouse. Zoomed area demonstrating co-expression between Gal9, CD14 and Caspase-1 are shown on the right. Arrows highlight cells co-expressing the mentioned markers

Article Snippet: Plasma levels of soluble CD14, IL-6 and IL-1β were determined using the Human CD14 DuoSet ELISA, the Human IL-6 DuoSet ELISA kits (both from R&D systems) or the Quantikine ELISA Human IL- 1β/IL-1F2 kit (R&D systems), respectively.

Techniques: Expressing, Infection

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Evidence for the presence of toll-like receptor 4 system in the human endometrium.

doi: 10.1210/jc.2004-0241

Figure Lengend Snippet: FIG. 1. Expression of TLR4, CD14, MD2, and MyD88 mRNA in EECs and ESCs. TLR4, CD14, MD2, and MyD88 mRNA was detected by RT-PCR. Total RNA was extracted from cultured EECs and ESCs of six patients in proliferative or secretory phase (n 3 in each phase). RNA without RT was used for negative controls. Lane E, EECs; lane S, ESCs.

Article Snippet: Human recombinant CD14 and IFN- were obtained from R&D systems (Minneapolis, MN).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Cell Culture

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Evidence for the presence of toll-like receptor 4 system in the human endometrium.

doi: 10.1210/jc.2004-0241

Figure Lengend Snippet: FIG. 6. Cell-surface expression of CD14 determined by flow cytom- etry. Cultured EECs and ESCs were collected and stained with anti- CD14 monoclonal antibody (solid line) or isotype control mouse IgG2b (dotted line). The result is representative of five separate experiments using samples from different patients.

Article Snippet: Human recombinant CD14 and IFN- were obtained from R&D systems (Minneapolis, MN).

Techniques: Expressing, Cell Culture, Staining, Control

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Evidence for the presence of toll-like receptor 4 system in the human endometrium.

doi: 10.1210/jc.2004-0241

Figure Lengend Snippet: FIG. 7. Immunohistochemistry of CD14 in the human endometrium. Sections were immunostained with antihuman CD14 antibody (A and C) or mouse IgG2a (B and D). Insets show luminal epithelial cells stained with antihuman CD14 (C) or mouse IgG2a (D). Magnification (A, B), 100, (C, D) 200

Article Snippet: Human recombinant CD14 and IFN- were obtained from R&D systems (Minneapolis, MN).

Techniques: Immunohistochemistry, Staining

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Evidence for the presence of toll-like receptor 4 system in the human endometrium.

doi: 10.1210/jc.2004-0241

Figure Lengend Snippet: FIG. 8. Effects of anti-TLR4 antibody (aTLR4) and anti-CD14 (aCD14) antibody on LPS-induced IL-8 secretion by ESCs. ESCs were preincubated in 1% FBS medium with or without mouse IgG2a (20 g/ml), anti-TLR4 antibody (20 g/ml), sheep IgG (10 g/ml), or anti- CD14 antibody (10 g/ml) for 30 min and stimulated with or without LPS (100 ng/ml) for 24 h. Concentrations of IL-8 in the conditioned media were measured by ELISA. All values are expressed as the mean SEM of quadruplicate cultures. *, P 0.001 vs. LPS; **, P 0.0001 vs. LPS. The result is representative of five separate experi- ments using samples from different patients.

Article Snippet: Human recombinant CD14 and IFN- were obtained from R&D systems (Minneapolis, MN).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Evidence for the presence of toll-like receptor 4 system in the human endometrium.

doi: 10.1210/jc.2004-0241

Figure Lengend Snippet: FIG. 9. Effects of anti-TLR4 antibody and anti-CD14 antibody on LPS-induced NF-B (p65) nuclear translocation in ESCs. ESCs were stimulated with (B) or without (A) LPS (100 ng/ml) in 1% FBS medium for 24 h. ESCs were preincubated in 1% FBS medium with mouse IgG2a (20 g/ml) (C), anti-TLR4 antibody (20 g/ml) (D), sheep IgG (10 g/ml) (E), anti-CD14 antibody (10 g/ml) (F), or polymyxin B (G) for 30 min and stimulated with LPS (100 ng/ml) for 24 h.

Article Snippet: Human recombinant CD14 and IFN- were obtained from R&D systems (Minneapolis, MN).

Techniques: Translocation Assay

Journal: Veterinary immunology and immunopathology

Article Title: Flow cytometry analysis of CD11c-positive peripheral blood mononuclear cells in horses.

doi: 10.1016/j.vetimm.2022.110504

Figure Lengend Snippet: Fig. 2. Cell immunophenotyping of horse PBMCs using a panel with CD11c, MHC-II, CD14 and TLR4 and cross-reactivity assays. (a) Gate strategy to remove doublets in the forward and side scatter with height to area and cell population identification of population 1 (P1), population 2 (P2) and population 3 (P3). (b) Cell expression dot plot of CD11c+CD14+, CD11c+MHCII+ and CD11c+TLR4+ P1, P2 and P3. (c) Cross-reactivity assay of antibodies for their recognition capacity in equine PBMCs. Black histograms represent the negative control, and colored histograms are representative of P1, P2 and P3. Data representative from one horse. Samples were analyzed by flow cytometry; 50,000 events were obtained. The analysis was performed using FlowJo V10 software.

Article Snippet: The cells were incubated with other surface markers using the following antibodies: anti-horse MHCII-FITC (Batch No. 0515, Bio-Rad), CD14-AF700 (FAB4597N, Novus Biologicals), and TLR4-PE (clone: HTA125, Cat: 312805, Bio-Legend).

Techniques: Expressing, Negative Control, Flow Cytometry, Software