anti mouse ly6g  (Bio X Cell)

Journal: The FASEB Journal

Article Title: Lymphocyte Antigen 6G Mediates Vagotomy‐Associated Reduction in Body Weight

doi: 10.1096/fj.202600151RR

Figure Lengend Snippet: Vagotomy promoted Ly6G + cell infiltration into eWAT. Wild‐type mice were subjected to left cervical vagotomy (VX) or sham surgery and tissues were collected at 7 days following surgery. (A) The percentage of non‐adipocyte nuclei of total nuclei per section was quantified using ImageJ ( n = 4) and right panels show representative images of paraffin sections of eWAT stained with H&E in sham and VX animals. (B) CCL2 release from eWAT was analyzed by ELISA. The bar shows the CCL2 levels from sham ( n = 4) or VX ( n = 4) mice normalized to eWAT weight: ng/mL per g ± SEM (unpaired Student's t test). (C) eWAT was collected at 1 ( n = 3), 4 ( n = 4 sham, n = 5 VX), and 7 ( n = 15) days following VX or sham surgery and the eWAT SVCs were analyzed by flow cytometry. The bar shows the % ± SEM of CD11b + Ly6G + cells from CD45 + (one‐way ANOVA, Uncorrected Fisher's LSD). (D) Graphs show representative gating for CD11b + Ly6G + cells in sham and VX eWAT at 7 days (concatenated n = 5–6). (E) Representative immunostaining of Ly6G (red) and Perilipin1 (green) in paraffin sections of eWAT. (F–H) Bone marrow neutrophils after sham ( n = 9) or VX ( n = 5) surgery were isolated using negative magnetic beads and analyzed using bulk RNAseq (DESeq2). Heatmap (F), volcano plot (G) of differentially expressed genes, and GO (Gene Ontology) (H) enrichment bar plot. ns = not significant, * p < 0.05. VX, Vagotomy; eWAT, epididymal white adipose tissue; H&E, hematoxylin–eosin; SVCs, stromal vascular cells.

Article Snippet: To ligate Ly6G surface epitopes, male C57BL/6 mice were intraperitoneally injected with InVivo Mab anti‐mouse Ly6G (100 ug/100 uL) (Bioxcell, #BE0075) or for control InVivoMAb rat IgG2a (anti‐trinitrophenol) isotype (100 ug/100 uL) (Bioxcell, #BE0089) antibodies 2 days before the sham or VX surgery.

Techniques: Staining, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Immunostaining, Isolation, Magnetic Beads, RNA sequencing

Journal: The FASEB Journal

Article Title: Lymphocyte Antigen 6G Mediates Vagotomy‐Associated Reduction in Body Weight

doi: 10.1096/fj.202600151RR

Figure Lengend Snippet: Neutrophil deficiency attenuated the VX‐associated reduction of eWAT weight. Ly6G cre Mcl1 fl/fl mice were subjected to VX or sham surgery and tissues were collected at 7 days following the surgery. (A) Flow cytometry analysis of CD11b + Ly6G + cells in eWAT ( n = 7–9), blood ( n = 3–6), and bone marrow ( n = 3–6) following VX or sham surgery in wild‐type (WT) and Ly6G cre Mcl1 fl/fl (KO) mice. Bars show the proportion of cells from CD45 + (one‐way ANOVA, uncorrected Fisher's LSD). (B) The graph shows the change in body weight of WT ( n = 3) and Ly6G cre Mcl1 fl/fl (KO) ( n = 3–6) mice shown as grams±SEM from body weight day 0 (two‐way ANOVA, Tukey's multiple comparisons test—Significant differences between experimental groups at each time point are indicated with a and b, and the detailed description can be found in Table ). (C) eWAT weight was recorded at 7 days following VX or sham surgery ( n = 8–10 sham, n = 9 VX). The bars depict the relative eWAT weight to sham eWAT weight in % ± SEM (one‐way ANOVA, Šídák's multiple comparisons test). (D) Flow cytometry analysis of CD11b + Ly6G − F4/80 + cells in eWAT ( n = 5–8), blood ( n = 3–6), and bone marrow ( n = 3) following VX or sham surgery in WT and Ly6G cre Mcl1 fl/fl (KO) mice. Bars show the proportion of cells from CD45 (one‐way ANOVA, Šídák's multiple comparisons test). eWAT, epididymal white adipose tissue; ns, not significant, * p < 0.05, *** p < 0.001, VX, Vagotomy.

Article Snippet: To ligate Ly6G surface epitopes, male C57BL/6 mice were intraperitoneally injected with InVivo Mab anti‐mouse Ly6G (100 ug/100 uL) (Bioxcell, #BE0075) or for control InVivoMAb rat IgG2a (anti‐trinitrophenol) isotype (100 ug/100 uL) (Bioxcell, #BE0089) antibodies 2 days before the sham or VX surgery.

Techniques: Flow Cytometry

Journal: The FASEB Journal

Article Title: Lymphocyte Antigen 6G Mediates Vagotomy‐Associated Reduction in Body Weight

doi: 10.1096/fj.202600151RR

Figure Lengend Snippet: Temporary extracellular Ly6G depletion effect on VX‐associated reduction of eWAT weight. (A) Flow cytometry analysis of the frequency of extracellular Ly6G and CD11b double positive cells following one intraperitoneal injection of either anti‐Ly6G or vehicle (PBS). Bars show the % ± SEM of CD11b + Ly6G + in vehicle ( n = 4–5) and at 1 ( n = 2), 2 ( n = 4), 5 ( n = 4), and 9 ( n = 3) days following injection in eWAT, blood, and bone marrow (one‐way ANOVA, Šídák's multiple comparisons test). (B) Schematic diagram depicting the experimental setup: Wild‐type mice were intraperitoneally injected once with anti‐Ly6G antibody or IgG2a antibody 2 days before sham or VX surgery, and tissue was collected 7 days following surgery. (C) Flow cytometry analysis of CD11b + Ly6G + cells in eWAT ( n = 3) following VX or sham. Bars show the proportion of cells from CD45 + (one‐way ANOVA, uncorrected Fisher's LSD). (D) Correlation between extracellular and intracellular expression of Ly6G in flow cytometry analysis. Circles represent each sample stained for both extracellular and intracellular Ly6G in separate fluorescent channels (Pearson r correlation). (E) The mice were weighed daily. The graph shows the difference in body weight (g) of the mice from day 0 (before surgery) of each experimental group ( n = 3) in g ± SEM (two‐way ANOVA, Tukey's multiple comparisons test—Significant differences between experimental groups at each time point are indicated with a, b, and c, and the detailed description can be found in Table ). (F) eWAT weight ( n = 3) was recorded at 7 days following VX or sham surgery. The bars show the relative eWAT weight to sham eWAT weight in % ± SEM (one‐way ANOVA, uncorrected Fisher's LSD). (G) Mice were kept in separate cages according to experimental groups: Sham+IgG2a, VX + IgG2a, sham+anti‐Ly6G, VX + anti‐Ly6G ( n = 3). The food for each cage was weighed at the same time point daily. The curve shows the grams of food consumed per day per cage in g. ns = not significant, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. VX, Vagotomy; eWAT, epididymal white adipose tissue.

Article Snippet: To ligate Ly6G surface epitopes, male C57BL/6 mice were intraperitoneally injected with InVivo Mab anti‐mouse Ly6G (100 ug/100 uL) (Bioxcell, #BE0075) or for control InVivoMAb rat IgG2a (anti‐trinitrophenol) isotype (100 ug/100 uL) (Bioxcell, #BE0089) antibodies 2 days before the sham or VX surgery.

Techniques: Flow Cytometry, Injection, Expressing, Staining

Journal: The FASEB Journal

Article Title: Lymphocyte Antigen 6G Mediates Vagotomy‐Associated Reduction in Body Weight

doi: 10.1096/fj.202600151RR

Figure Lengend Snippet: Extracellular Ly6G depletion attenuated VX‐mediated reduction of body weight. (A) Schematic of the experimental setup: Wild‐type mice were intraperitoneally injected with anti‐Ly6G antibody or IgG2a antibody 2 days before, as well as 3 and 5 days following sham or VX surgery, and tissue was collected 7 days following surgery. (B–C) Flow cytometry analysis of CD11b + Ly6G + cells in eWAT ( n = 6) and blood ( n = 6) following VX or sham surgery, with IgG2a or anti‐Ly6G treatment. Bars show the proportion of cells from CD45 + (One‐way Anova, Uncorrected Fisher's LSD). (D–E) Flow cytometry analysis of intracellular Ly6G + cells in eWAT ( n = 3) and blood ( n = 3) following VX or sham surgery, with IgG2a or anti‐Ly6G treatment. Bars show the proportion of cells from CD45 + (One‐way Anova, Uncorrected Fisher's LSD). (F–G) Flow cytometry analysis of CD11b + (IN)Ly6G − F4/80 + cells in eWAT ( n = 3) and blood ( n = 3) following VX or sham surgery, with IgG2a or anti‐Ly6G treatment. Bars show the proportion of cells from CD45 + (One‐way Anova, Uncorrected Fisher's LSD). (H) eWAT weight ( n = 6) was recorded at 7 days following VX or sham surgery. The bars depict the relative eWAT weight to sham eWAT weight in % ± SEM (one‐way ANOVA, uncorrected Fisher's LSD). (I) The mice were weighed daily. The graph shows the difference in body weight of the mice from day 0 (before initiation of surgery) of each experimental group ( n = 6) in % ± SEM (two‐way ANOVA, Tukey's multiple comparisons test—Significant differences between experimental groups at each time point are indicated with a, b, and c, and the detailed description can be found in Table ). ns = not significant, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. VX, Vagotomy; eWAT, epididymal white adipose tissue; BM, bone marrow; SVCs, stromal vascular cells; intracellular (IN).

Article Snippet: To ligate Ly6G surface epitopes, male C57BL/6 mice were intraperitoneally injected with InVivo Mab anti‐mouse Ly6G (100 ug/100 uL) (Bioxcell, #BE0075) or for control InVivoMAb rat IgG2a (anti‐trinitrophenol) isotype (100 ug/100 uL) (Bioxcell, #BE0089) antibodies 2 days before the sham or VX surgery.

Techniques: Injection, Flow Cytometry

Journal: European Journal of Immunology

Article Title: Depletion of Neonatal Neutrophilic Cells Worsens the Outcome of E. coli Sepsis in Newborn Mice

doi: 10.1002/eji.70171

Figure Lengend Snippet: Sepsis mortality after depletion of Ly6G‐positive NNC. (A) Schematic representation of the experimental procedure. (B–D) Newborn mice received either an intraperitoneal injection of an anti‐Ly6G antibody to deplete NNC or an isotype control antibody on the first postnatal day (P1). At P2 or P8 spleen cells were isolated and analyzed by flow cytometry. (B) Representative density plots showing the population of CD11b + /Ly6G + (upper right quadrant) NNC in newborn mice at P2 after administering either isotype control (left side, Isotype) or the depleting antibody (right side, anti Ly6G). Cells were pre‐gated for living CD45 + cells. (C, D) Bar graphs showing percentages of total NNC from CD45 + /CD11b + myeloid cells in spleens of newborn mice at P2 (C; 1 day after depletion; isotype: n = 7, anti Ly6G: n = 5) and P8 (D; 7 days after depletion; isotype: n = 5, anti Ly6G: n = 5). Bars show mean and standard deviation, ** p < 0.01, Mann–Whitney test. (E, F) Newborn mice received either an intraperitoneal injection of an anti‐Ly6G antibody to deplete NNC or an isotype control antibody at the first day of life (P1). At P2, mice were injected subcutaneously with either 10,000 CFU or 30,000 CFU E. coli . The pups were sacrificed when reaching the criteria for critical illness, or 48 h after sepsis induction. Probability of survival after subcutaneous injection of 10,000 CFU E. coli (E; anti Ly6G n = 8, isotype n = 8; * p < 0.05, log‐rank [Mantel–Cox] test) or 30,000 CFU E. coli (F; anti Ly6G n = 8, isotype n = 9; p:0,186: not significant, log‐rank [Mantel–Cox] test) in mice after NNC‐depletion (anti Ly6G) or injection of an isotype control (isotype). Each n is representing a single mouse. Mice for the experiments came from three litters (B–F).

Article Snippet: To study the effect of NNC depletion on the course of neonatal sepsis, newborn mice were administered either an intraperitoneal injection of 10 μg/mouse of anti‐Ly6G (1A8, anti‐mouse Ly6G, Bio X Cell Lebanon, USA) to deplete Ly6G‐expressing cells, or 10 μg/mouse of isotype control antibody (2A3, rat IgG2a isotype control, Bio X Cell Lebanon, USA) 1 day prior to sepsis induction.

Techniques: Injection, Control, Isolation, Flow Cytometry, Standard Deviation, MANN-WHITNEY

Journal: European Journal of Immunology

Article Title: Depletion of Neonatal Neutrophilic Cells Worsens the Outcome of E. coli Sepsis in Newborn Mice

doi: 10.1002/eji.70171

Figure Lengend Snippet: Bacterial load and serum concentrations of IL‐6 and MCP‐1 after depletion of Ly6G‐positive NNC. Newborn mice received either an intraperitoneal injection of an anti‐Ly6G antibody (anti Ly6G) to deplete NNC or an isotype control antibody (Iso) at the first day of life (P1). At P2, mice were injected subcutaneously with either 10,000 CFU or 30,000 CFU E. coli . Injection of PBS served as negative control. The pups were sacrificed when reaching the criteria for critical illness or 48 h after sepsis induction. Spleens, lungs, livers, blood, and peritoneal lavage were collected. Organs were homogenized and samples were diluted with PBS. Suspensions were incubated for 24 h on Columbia agar plates with 5% sheep blood and colony forming units (CFU) were counted to calculate CFU per gram (g) organ weight or per milliliter (mL) fluid (blood or peritoneal lavage). Scatter diagrams with bars showing CFUs per gram organ weight (A–C) or per ml (D, E) in spleens (A), livers (B), lungs (C), blood (D), and peritoneal fluid (E) of control mice (Iso, white bars) or NNC‐depleted mice (anti‐Ly6G, striped bars) without sepsis induction (control, left side) or with sepsis induction with 10,000 CFU (middle) or 30,000 CFU (right side). Bars show mean and standard deviation. n = 5–9, * p < 0.05, ns: not significant, Mann–Whitney test. (F, G) Newborn mice received either an intraperitoneal injection of an anti‐Ly6G antibody (anti Ly6G, checked bar) to deplete NNC or an isotype control (Iso, plain bar) on the first day of life (P1). On P2, newborn mice from a litter with a size of 5–6 pups were injected subcutaneously with either 10,000 CFU (middle, n = 5–8) or 30,000 CFU (right side, n = 5–8) or with PBS as a negative control (left side, n = 6). After termination of the experiment or 48 h after beginning of the experiment, if the pups survived IL‐6 (F) and MCP‐1 concentration in serum samples was measured using enzyme‐linked immunosorbent assay (ELISA) IL‐6 (F) and MCP‐1 (G) concentration in serum samples was measured. The IL‐6 and MCP‐1 levels were normalized to the protein concentrations of each sample. Bars show mean and standard deviation. n = 3–8, * p < 0.05, ** p < 0.01; unpaired t test. Each dot (= n ) is representing a single mouse. Mice for the experiments came from three litters (A–G).We further investigated systemic inflammation during neonatal sepsis with and without depletion of Ly6G‐expressing NNC by measuring the levels of the proinflammatory cytokines IL‐6 and MCP‐1 in serum of the animals. Both, IL‐6 and MCP‐1 levels were significantly increased after Ly6G‐depletion both after sepsis induction with 10,000 (IL‐6: 187 ng/g vs. 443 ng/g; n = 7, 8; p < 0.05; MCP‐1: 3231 ng/g vs. 5737 ng/g; n = 3; ** p < 0.01) or 30.000 CFU E. coli (IL‐6: 422 ng/g vs. 552 ng/g, n = 3, p < 0.05; MCP‐1: 3808 g/g vs. 6470 g/g, n = 3; ** p < 0.01) (Figure ).

Article Snippet: To study the effect of NNC depletion on the course of neonatal sepsis, newborn mice were administered either an intraperitoneal injection of 10 μg/mouse of anti‐Ly6G (1A8, anti‐mouse Ly6G, Bio X Cell Lebanon, USA) to deplete Ly6G‐expressing cells, or 10 μg/mouse of isotype control antibody (2A3, rat IgG2a isotype control, Bio X Cell Lebanon, USA) 1 day prior to sepsis induction.

Techniques: Injection, Control, Negative Control, Incubation, Standard Deviation, MANN-WHITNEY, Concentration Assay, Enzyme-linked Immunosorbent Assay, Expressing

Journal: European Journal of Immunology

Article Title: Depletion of Neonatal Neutrophilic Cells Worsens the Outcome of E. coli Sepsis in Newborn Mice

doi: 10.1002/eji.70171

Figure Lengend Snippet: Immune cell populations in spleens after depletion of Ly6G‐positive NNC. Newborn mice received either an intraperitoneal injection of an anti‐Ly6G antibody (anti Ly6G) to deplete NNC or an isotype control antibody (Isotype) at (P1). At P2 mice were injected subcutaneously with either 10,000 CFU or 30,000 CFU E. coli . Injection of PBS served as negative control. The pups were sacrificed when reaching the criteria for critical illness or 48 h after sepsis induction. Spleens were collected and homogenized to obtain single cell suspensions. Immune cell subsets were analyzed by flow cytometry. Scatter diagrams with bars showing percentages of macrophages (A), dendritic cells (B), monocytes (C), B cells (D), T cells (E), and NK cells (F) and activated T cells of leukocytes of control mice (isotype, white bars) or NNC‐depleted mice (anti‐Ly6G, striped bars) without sepsis induction (control) or with sepsis induction with 10,000 CFU or 30,000 CFU. n = 6–7, unpaired t test. Each dot (= n ) is representing a single mouse. Mice for the experiments came from three litters.

Article Snippet: To study the effect of NNC depletion on the course of neonatal sepsis, newborn mice were administered either an intraperitoneal injection of 10 μg/mouse of anti‐Ly6G (1A8, anti‐mouse Ly6G, Bio X Cell Lebanon, USA) to deplete Ly6G‐expressing cells, or 10 μg/mouse of isotype control antibody (2A3, rat IgG2a isotype control, Bio X Cell Lebanon, USA) 1 day prior to sepsis induction.

Techniques: Injection, Control, Negative Control, Single Cell, Flow Cytometry