mouse dkk1 quantikine elisa kit  (R&D Systems)

Journal: Frontiers in Immunology

Article Title: Leishmania major surface components and DKK1 signalling via LRP6 promote migration and longevity of neutrophils in the infection site

doi: 10.3389/fimmu.2024.1473133

Figure Lengend Snippet: Decreased CD11b and MHC class II positive neutrophils derived from LRP6 NKO DKK1 PKO and LRP6 NKO infected mice. The WT-BALB/c, LRP6 NKO and LRP6 NKO DKK1 PKO mice were challenged with infective metacyclic promastigote (2 x 10 6 parasites, n = 5) of L. major via the footpad. Non-infected BALB/c mice (n = 10/2 feet per mouse) were given 0.9% NaCl saline. Neutrophils were isolated from the footpads of all infected and non-infected mice at day 3 PI. Isolated neutrophil samples were analyzed by flow cytometry for CD11b and MHC class II positive neutrophils. Each dot indicates MHC class II (A) and CD11b (B) positive cells. Representative flow cytometry dot plots showing the analyses of CD11b and MHC class II + neutrophils performed on day 3 PI, as well as the concatenated dot plot of each sample in all the experimental groups are presented in Supplementary Figure S3 . In all the experiments, WT-infected and non-infected mice served as positive and negative controls, respectively. Results are presented as mean (± SEM). One-way ANOVA with Bonferroni’s post hoc test was performed to analyze the data **p < 0.01, ***p < 0.001(p > 0.05).

Article Snippet: The concentration of DKK1 in plasma was determined by Enzyme-linked immunosorbent assays using a mouse DKK1 ELISA kit (Thermo Fisher Scientific) according to the manufacturer’s protocol.

Techniques: Derivative Assay, Infection, Saline, Isolation, Flow Cytometry

Journal: Frontiers in Immunology

Article Title: Leishmania major surface components and DKK1 signalling via LRP6 promote migration and longevity of neutrophils in the infection site

doi: 10.3389/fimmu.2024.1473133

Figure Lengend Snippet: Decreased myeloperoxidase positive neutrophils derived from LRP6 NKO DKK1 PKO and LRP6 NKO infected mice on day 3 PI. The WT-BALB/c, LRP6 NKO and LRP6 NKO DKK1 PKO mice were challenged with infective metacyclic promastigote (2 x 10 6 parasites, n = 5) of L. major via the footpad. Non-infected BALB/c mice (n = 10/2 feet per mouse) were given 0.9% NaCl saline. Neutrophils were isolated from the footpads of all infected and non-infected mice at day 3 PI. Isolated neutrophil samples were analyzed by flow cytometry for myeloperoxidase + cells. Each dot indicates myeloperoxidase + neutrophils, and the percentage of myeloperoxidase+ cells in the different experimental groups is shown in column graphs. Representative flow cytometry dot plots showing the analyses of myeloperoxidase + neutrophils and IgG1 isotype control for non-specific antibody staining, as well as a dot plot of each sample in all the experimental groups are presented in Supplementary Figure S4 . In all the experiments, WT-infected and non-infected mice served as positive and negative controls, respectively. Results are presented as mean (± SEM). One-way ANOVA with Bonferroni’s post hoc test was performed to analyze the data **p < 0.01; ***p < 0.001; ns, non-significant (p > 0.05).

Article Snippet: The concentration of DKK1 in plasma was determined by Enzyme-linked immunosorbent assays using a mouse DKK1 ELISA kit (Thermo Fisher Scientific) according to the manufacturer’s protocol.

Techniques: Derivative Assay, Infection, Saline, Isolation, Flow Cytometry, Control, Staining

Journal: Frontiers in Immunology

Article Title: Leishmania major surface components and DKK1 signalling via LRP6 promote migration and longevity of neutrophils in the infection site

doi: 10.3389/fimmu.2024.1473133

Figure Lengend Snippet: Infected LRP6 NKO and LRP6 NKO DKK1 PKO mice show elevated neutrophil apoptosis. The WT-BALB/c, LRP6 NKO and LRP6 NKO DKK1 PKO mice were challenged with infective metacyclic promastigote (2 x 10 6 parasites, n = 5) of L. major via the footpad. Non-infected BALB/c mice (n = 10/2 feet per mouse) were given 0.9% NaCl saline. Neutrophils were isolated from the footpads of all infected and non-infected mice at day 3 PI. Apoptotic neutrophils were analyzed by flow cytometry and identified by the increase in fluorescence intensity of annexin V-FITC. Each dot indicates either viable or apoptotic Ly6G+ cells. Percentage of apoptotic cells (early apoptosis: Annexin + , late apoptosis: Annexin + PI + ) in the infected and non-infected mice are shown in column graph (A) & neutrophil viability is shown in column graph (B) . Representative flow cytometry dot plots showing apoptotic and viable neutrophils after double staining with Annexin V-FITC and propidium iodide performed on day 3 PI. The first quadrant (Q1) represents necrotic neutrophils, the second quadrant (Q2) represents later apoptotic neutrophils, the third quadrant (Q3) represents early apoptotic neutrophils, and the fourth quadrant (Q4) represents normal neutrophils Supplementary Figure S5 . A concatenated dot plot of each sample in all the experimental groups is presented in Supplementary Figure S5 . In all the experiments, WT-infected and non-infected mice served as positive and negative controls, respectively. Results are presented as mean (± SEM). One-way ANOVA with Bonferroni’s post hoc test was performed to analyze the data *p < 0.05; **p < 0.01, ***p < 0.001.

Article Snippet: The concentration of DKK1 in plasma was determined by Enzyme-linked immunosorbent assays using a mouse DKK1 ELISA kit (Thermo Fisher Scientific) according to the manufacturer’s protocol.

Techniques: Infection, Saline, Isolation, Flow Cytometry, Fluorescence, Double Staining

Journal: Frontiers in Immunology

Article Title: Leishmania major surface components and DKK1 signalling via LRP6 promote migration and longevity of neutrophils in the infection site

doi: 10.3389/fimmu.2024.1473133

Figure Lengend Snippet: Recombinant DKK1 delayed neutrophil apoptosis in a dose-dependent manner. Neutrophils isolated from naïve mice were incubated with various concentrations of rDKK1. Neutrophil samples harvested at 0-, 4-, and 24 hrs post incubation were used to determine neutrophil apoptosis by flow cytometry. The percentage of apoptotic cells in the different experimental conditions is shown in the bar graph. Representative flow cytometry contour plots generated 24 hrs post incubation showed total apoptotic neutrophils (Q3 & Q2) after double staining with Annexin V-FITC and propidium iodide, as presented in Supplementary Figure S6 . Also, a contour plot of each sample in all the experimental conditions is presented in Supplementary Figure S6 . In all the experiments, non-treated neutrophils served as controls. Results are presented as mean (± SEM). One-way ANOVA with Bonferroni’s post hoc test was performed to analyze the data (*p < 0.05, **p < 0.01). ns, non-significant.

Article Snippet: The concentration of DKK1 in plasma was determined by Enzyme-linked immunosorbent assays using a mouse DKK1 ELISA kit (Thermo Fisher Scientific) according to the manufacturer’s protocol.

Techniques: Recombinant, Isolation, Incubation, Flow Cytometry, Generated, Double Staining

Journal: Frontiers in Immunology

Article Title: Leishmania major surface components and DKK1 signalling via LRP6 promote migration and longevity of neutrophils in the infection site

doi: 10.3389/fimmu.2024.1473133

Figure Lengend Snippet: Parasite load in the infected LRP6 NKO and LRP6 NKO DKK1 PKO mice were significantly reduced, while plasma DKK1 production in infected BALB/c and LRP6 NKO mice were comparable. BALB/c, LRP6 NKO and LRP6 NKO DKK1 PKO mice were challenged with infective metacyclic promastigote (2 x 10 6 parasites, n = 5) of WT L. major strain via the footpad. Control mice (n = 10/2 feet per mouse) were given 0.9% NaCl Saline. The infected foot from each mouse was used to determine parasite load using a limiting dilution assay on day 14 PI. Parasite load in the infected BALB/c group was compared with the infected LRP6 NKO and LRP6 NKO DKK1 PKO mice. To determine DKK1 production, blood was collected via the maxillary vein at day 3 PI. Plasma samples were analyzed by ELISA. In all experiments, Infected and non-infected BALB/c mice served as positive and negative controls, respectively. Results of parasite load are presented as mean +/- SEM, and data analysis was done using one-way ANOVA with Bonferroni’s post hoc test *p < 0.05, **p < 0.01, (p > 0.05). For DKK1 concentration, results are presented as mean +/- SEM of replicate wells. One-way ANOVA with Bonferroni’s post hoc test was performed to analyze the data **p < 0.01, ‘ns’ indicates not significant (p > 0.05).

Article Snippet: The concentration of DKK1 in plasma was determined by Enzyme-linked immunosorbent assays using a mouse DKK1 ELISA kit (Thermo Fisher Scientific) according to the manufacturer’s protocol.

Techniques: Infection, Clinical Proteomics, Control, Saline, Limiting Dilution Assay, Enzyme-linked Immunosorbent Assay, Concentration Assay

Journal: Nature communications

Article Title: Stroma-derived Dickkopf-1 contributes to the suppression of NK cell cytotoxicity in breast cancer.

doi: 10.1038/s41467-025-56420-w

Figure Lengend Snippet: Fig. 8 | DKK1 levels correlate with reduced therapeutic responses and cytotoxic NK cells. A Schematic representation of NK cell isolation from healthy donor human PBMCs and incubation with target cells in the presence of PBS or rhDKK1 for 4 h. B–D Analysis of percent specific killing measured by 7-AAD+ MDA-MB-231 (B, n = 7 NK donors), T47D (C, n = 3 NK donors), and K562 (D, n = 3 NK donors) target tumor cells after 4 h incubation with human NK cells in the presence of rhDKK1 or PBS as a control. E FACS analysis of NK cell activating receptors on human NK cells from different donors (n = 11) stimulated with rhDKK1 for 24 h or PBS. F Schematic representation of blood sample collection of advanced breast

Article Snippet: Serum or plasma levels of DKK1 frommice or patients were quantified using ELISA kits specific for DKK1 (Mouse Dkk1 Quantikine ELISA kit or Human Dkk1 Quantikine ELISA kit, R&D systems) as per the manufacturer’s protocols.

Techniques: Cell Isolation, Incubation, Control