human notch2 igg 1 blocking antibody  (Novus Biologicals)

Journal: PLoS ONE

Article Title: Notch2 Controls Prolactin and Insulin-Like Growth Factor Binding Protein-1 Expression in Decidualizing Human Stromal Cells of Early Pregnancy

doi: 10.1371/journal.pone.0112723

Figure Lengend Snippet: Serial sectioning of paraffin-embedded tissues and immunofluorescence were performed as described in . Representative examples (8 th week of pregnancy) of 5 different deciduae analyzed are depicted. DSC, decidual stromal cells; GEC, glandular epithelial cell; LC, leukocytes; Scale bars represent 100 µm. (A) Localization of Notch family members in human deidual stromal cells. Double staining with antibodies recognizing vimentin (Vim, green), Notch2 (green) and CD45 (red) mark decidual stromal cells and leukocytes, respectively. The respective counterstaining with DAPI is depicted on the right hand side. Double stainings with the appropriate isotype-specific monoclonal (mAb, green, insert picture) and polyclonal (pAb, green, insert picture) controls with vimentin (Vim, red) are shown. (B) Glandular expression of Notch receptors and ligands. Double staining with antibodies recognizing vimentin (Vim, green) or cytokeratin 7 (KRT7, red) was used to depict DSC and GEC, respectively. Co-staining of Notch receptor or ligand (both green) with nuclear staining (DAPI, blue) is shown.

Article Snippet: To specifically inhibit the Notch2 receptor, seeded cells were treated with 0.4 μg/ml human IgG isotype control (Novus Biologicals) or 0.4 μg/ml therapeutic human Notch2 IgG 1 blocking antibody, which binds and stabilizes the auto-inhibited, negative regulatory region (NRR) of the receptor, preventing conformational changes and therefore ADAM-mediated cleavage .

Techniques: Immunofluorescence, Double Staining, Expressing, Staining

Journal: PLoS ONE

Article Title: Notch2 Controls Prolactin and Insulin-Like Growth Factor Binding Protein-1 Expression in Decidualizing Human Stromal Cells of Early Pregnancy

doi: 10.1371/journal.pone.0112723

Figure Lengend Snippet: Cells were stimulated with cAMP and/or E2P4 for 3 and 6 days. Total protein lysates were analyzed by western blotting and quantitated using densitometry as described in . A representative example out of 3 different experiments is shown. n.c., non-stimulated controls; (A) Western blot showing specific signals for Notch2, Jagged1, DLL1 and DLL4 (marked by arrows). GAPDH was used as a loading control, * indicates unspecific band. (B) Densitometrical quantification of western blot signals normalized to GAPDH. Bars (n = 3) represent mean values ± S.D. For relative quantification of protein expression, values of n.c. at day 3 were arbitrarily set to 100%. * indicates p≤0.05 compared to n.c. of day 3; n.s., not significant.

Article Snippet: To specifically inhibit the Notch2 receptor, seeded cells were treated with 0.4 μg/ml human IgG isotype control (Novus Biologicals) or 0.4 μg/ml therapeutic human Notch2 IgG 1 blocking antibody, which binds and stabilizes the auto-inhibited, negative regulatory region (NRR) of the receptor, preventing conformational changes and therefore ADAM-mediated cleavage .

Techniques: Western Blot, Control, Quantitative Proteomics, Expressing

Journal: PLoS ONE

Article Title: Notch2 Controls Prolactin and Insulin-Like Growth Factor Binding Protein-1 Expression in Decidualizing Human Stromal Cells of Early Pregnancy

doi: 10.1371/journal.pone.0112723

Figure Lengend Snippet: Cells were cultured in the absence or presence of Notch2 blocking antibodies (Notch2 ab) (A) or si-RNAs targeting Notch2 (si-Notch2) (B). (A) Luciferase activity of the canonical Notch reporter in the presence of Notch2 ab or IgG controls. Reporter expression was normalized to constitutive β-Gal activity. Mean values ± S.D. of 3 experiments performed in duplicates are shown. IgG ctrl, IgG control; * indicates p≤0.05 compared to IgG-treated n.c. Significant changes (*, p≤0.05) between DAPT-treated (open bars) and untreated (black bars) cells are indicated by brackets. (B) Western blot showing Notch2 protein expression in siRNA-treated HDSC. Specific Notch2 signals are marked by an arrow (110 kDa). GAPDH was used as a loading control. Representative examples of 3 independent experiments are depicted. Lower panel shows the densitometrical quantification calculated from 3 independent experiments. si-ntc, si-non targeting control. * indicates p≤0.05 compared to si-ntc of the same day.

Article Snippet: To specifically inhibit the Notch2 receptor, seeded cells were treated with 0.4 μg/ml human IgG isotype control (Novus Biologicals) or 0.4 μg/ml therapeutic human Notch2 IgG 1 blocking antibody, which binds and stabilizes the auto-inhibited, negative regulatory region (NRR) of the receptor, preventing conformational changes and therefore ADAM-mediated cleavage .

Techniques: Cell Culture, Blocking Assay, Luciferase, Activity Assay, Expressing, Control, Western Blot

Journal: PLoS ONE

Article Title: Notch2 Controls Prolactin and Insulin-Like Growth Factor Binding Protein-1 Expression in Decidualizing Human Stromal Cells of Early Pregnancy

doi: 10.1371/journal.pone.0112723

Figure Lengend Snippet: HDSC were differentiated with cAMP/E2P4 for 6 days in the absence or presence of Notch2-blocking antibodies or siRNAs as mentioned in . Quantitative real-time PCR analyses showing mRNA expression of the Notch target gene HES1 and the markers of decidualization, IGFBP1 and PRL, upon addition of Notch2 blocking antibodies (A) or Notch2 siRNAs (B). For relative quantification non-stimulated controls (n.c.) were arbitrarily set to 1. Bars indicate mean values ± S.D. of 3 different experiments performed in duplicates. * indicates p≤0.05 compared to n.c.; Significant changes (*, p≤0.05) between Notch2 ab or si-Notch2-treated cells (open bars) and untreated (black bars) cells are indicated by brackets. IgG ctrl, IgG control; si-ntc, si-non targeting control.

Article Snippet: To specifically inhibit the Notch2 receptor, seeded cells were treated with 0.4 μg/ml human IgG isotype control (Novus Biologicals) or 0.4 μg/ml therapeutic human Notch2 IgG 1 blocking antibody, which binds and stabilizes the auto-inhibited, negative regulatory region (NRR) of the receptor, preventing conformational changes and therefore ADAM-mediated cleavage .

Techniques: Blocking Assay, Real-time Polymerase Chain Reaction, Expressing, Quantitative Proteomics, Control

Journal: PLoS ONE

Article Title: Notch2 Controls Prolactin and Insulin-Like Growth Factor Binding Protein-1 Expression in Decidualizing Human Stromal Cells of Early Pregnancy

doi: 10.1371/journal.pone.0112723

Figure Lengend Snippet: Cells were incubated with Notch2 blocking antibodies (A) or Notch2 siRNAs (B) and differentiated for 6 days in the presence of cAMP/E2P4. Supernatants were collected and prolactin (PRL) concentrations were measured by ELISA. Normalization to cellular protein concentrations was performed as mentioned in . Bars represent mean values ± S.D. of each 3 different experiments performed in duplicates. * indicates p≤0.05 compared to non-stimulated control (n.c.); Significant changes (*, p≤0.05) between Notch2 ab or si-Notch2-treated cells (open bars) and untreated (black bars) cells are indicated by brackets. IgG ctrl, IgG control; si-ntc, si-non targeting control.

Article Snippet: To specifically inhibit the Notch2 receptor, seeded cells were treated with 0.4 μg/ml human IgG isotype control (Novus Biologicals) or 0.4 μg/ml therapeutic human Notch2 IgG 1 blocking antibody, which binds and stabilizes the auto-inhibited, negative regulatory region (NRR) of the receptor, preventing conformational changes and therefore ADAM-mediated cleavage .

Techniques: Incubation, Blocking Assay, Enzyme-linked Immunosorbent Assay, Control