Human/Mouse Gremlin Antibody

Gremlin in Human Breast Cancer Tissue. Gremlin was detected in immersion fixed paraffin-embedded sections of human breast cancer tissue using Goat Anti-Human/Mouse Gremlin Antigen Affinity-purified Polyclonal Antibody (Catalog # AF956) at 3 µg/mL for 1 hour at room temperature followed by incubation with the Anti-Goat IgG VisUCyte™ HRP Polymer Antibody (Catalog # VC004). Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to cytoplasm. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

Gremlin in Embryonic Mouse Ribs. Gremlin was detected in immersion fixed frozen sections of embryonic mouse ribs (15 d.p.c.) using 15 µg/mL Goat Anti-Human/Mouse Gremlin Antigen Affinity-purified Polyclonal Antibody (Catalog # AF956) overnight at 4 °C. Tissue was stained with the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS008) and counterstained with hematoxylin (blue). View our protocol for Chromogenic IHC Staining of Frozen Tissue Sections.

Detection of Gremlin by Immunohistochemistry Inverse relationship between expression of Grem1 protein and pSmad1/5 staining in mouse intestine. Sequential FFPE sections (5 μm) of mouse intestine were stained for Grem1 (A) or pSmad1/5 (B) as described in Methods. (C) Positively stained cells (empty bars) or villi tips (filled bars) were quantified in the indicated regions using Image J and data were plotted as mean pixel intensity –/+ SEM (n = 3 mice, 3 independent regions of intestine quantified per mouse). *p< 0.05, ***p< 0.001 using two-way ANOVA and Bonferroni post hoc test. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/31384391), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Gremlin by Immunohistochemistry Distinct pattern of endogenous Grem1 expression in mouse colon. Sections (5 μm) from FFPE colon samples (n = 4) from wild-type or Grem1-/- mice were processed for in situ hybridisation (A left) and immunohistochemistry (A right; B.) as described in Methods. Positive Grem1 mRNA and protein staining was imaged using DAB (brown) and sections were counterstained using haematoxylin and imaged using PathXL. (A) Grem1 mRNA is visible as brown, punctate staining in the muscularis mucosa layer. Scale bars 100 μm. (B) Grem1 protein staining is evident as brown staining in the muscularis layer and the base of the colonic crypts (left; scale bar 100 μm). Grem1 protein staining in the musclaris layer as well as cells of the colonic crypt. CBCC, crypt base columnar cells; P, Paneth cells; TA, transit-amplifying cells. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/31384391), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Gremlin by Western Blot VEGFR2-MEK-ERK axis mediated the enhancement of FAO from Gremlin 1. (A). Cell viability assay of human intestinal fibroblast cells CCD-18Co and CCD-112Co cells treated with 200 ng/ml Gremlin 1 and Rapamycin (10 nM), LY3214996 (10 nM) or stattic (10 μM) for 24 h. (B). Immunoblotting analysis of VEGFR2-MEK-ERK signaling in human intestinal fibroblast cells CCD-18Co and CCD-112Co cells treated with 200 ng/ml Gremlin 1 and LY3214996 (10 nM) for 24 h. Actin acted as loading control and re-used for illustrative purposes. (C). Immunofluorescence staining of a-SMA in human intestinal fibroblast cells CCD-18Co and CCD-112Co cells treated with 200 ng/ml Gremlin 1 and LY3214996 (10 nM). (D). The intensity statistical results of a-SMA immunofluorescence staining from five random fields of slides. E. OCR measurement of CCD-18Co and CCD-112Co cells treated with 200 ng/ml Gremlin 1 and LY3214996 (10 nM) for 24 h. p values are derived from one-way ANOVA analysis. Dunnett’s test was used to analysis the difference between control group to the other groups. **p ≤ 0.01; ***p ≤ 0.001. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/33967807), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Gremlin by Western Blot BMP2, TGF-beta 1 and BMP8A show different effects on the expressional induction of Nog, Grem1 and Grem2. C3H10T1/2 cells were treated with BMP2 (1 nM), TGF-beta 1 (0.3 nM), BMP2 plus with TGF-beta 1 or BMP8A (1 nM) for 24 h. The transcript levels (upper panel) and encoding protein amounts (lower panel) of Nog (A), Grem1 (B) and Grem2 (C) were then evaluated. beta -actin amounts served as loading controls in immunoblotting. D To explore how BMP8A induces Grem1 expression, cells were treated with BMP8A in the absence or presence of SB431542 (SB) or dorsomorphin (DM) as indicated. Quantification data are presented as means ± SEM. *P < 0.05, ***P < 0.001 compared to no-treatment control. # P < 0.001 compared between indicated groups Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36443839), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Gremlin by Western Blot BMP2, TGF-beta 1 and BMP8A show different effects on the expressional induction of Nog, Grem1 and Grem2. C3H10T1/2 cells were treated with BMP2 (1 nM), TGF-beta 1 (0.3 nM), BMP2 plus with TGF-beta 1 or BMP8A (1 nM) for 24 h. The transcript levels (upper panel) and encoding protein amounts (lower panel) of Nog (A), Grem1 (B) and Grem2 (C) were then evaluated. beta -actin amounts served as loading controls in immunoblotting. D To explore how BMP8A induces Grem1 expression, cells were treated with BMP8A in the absence or presence of SB431542 (SB) or dorsomorphin (DM) as indicated. Quantification data are presented as means ± SEM. *P < 0.05, ***P < 0.001 compared to no-treatment control. # P < 0.001 compared between indicated groups Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36443839), licensed under a CC-BY license. Not internally tested by R&D Systems.