rat anti wnt5a monoclonal  (R&D Systems)

Journal: Cell Genomics

Article Title: Single-cell and spatial transcriptomic profiling revealed niche interactions sustaining growth of endometriotic lesions

doi: 10.1016/j.xgen.2024.100737

Figure Lengend Snippet:

Article Snippet: mouse monoclonal anti-WNT5A , Santa Cruz Biotechnology , Cat#sc-365370; RRID:AB_10846090.

Techniques: Recombinant, Red Blood Cell Lysis, Blocking Assay, RNA Sequencing, Gene Expression, Control, Software

Journal: bioRxiv

Article Title: Cancer-associated fibroblast-derived ROR2 induces WNT/PCP activation and polarized migration in receiving gastric cancer cells

doi: 10.1101/2022.04.07.487474

Figure Lengend Snippet: ROR2/WNT5A complexes are transferred from producing cells to receiving cells via cytonemes. a . WNT5A-GFP and ROR2-mCherry were overexpressed in producing AGS cells, and co-cultured with untransfected receiving AGS cells for 24 hours prior to imaging with confocal microscopy. Blue dashed line indicates the position of the receiving cell as detected by the brightfield image. Co-localising complexes in the receving cell are indicated with a white triangle. b . WNT5A-GFP and ROR2-mCherry were overexpressed in producing pCAF2 cells, and co-cultured with untransfected receiving AGS cells for 24 hours prior to imaging with confocal microscopy. Blue dashed line indictaes the position of the receiving cell as detected by the brightfield image. Scale bar indicates 10μm.

Article Snippet: Cells were washed again, incubated with anti-Wnt5a primary mouse monoclonal antibody clone A-5 (Santa Cruz) for 30 minutes on ice, washed in 0.5% BSA PBS buffer and incubated with goat anti-mouse IgG AF647 (Abcam) for 30 minutes on ice.

Techniques: Cell Culture, Imaging, Confocal Microscopy

Journal: bioRxiv

Article Title: Cancer-associated fibroblast-derived ROR2 induces WNT/PCP activation and polarized migration in receiving gastric cancer cells

doi: 10.1101/2022.04.07.487474

Figure Lengend Snippet: a . mRNA levels of the indicated gene shown as fold change away from AGS. Wnt5a is shown in red, values on the left y-axis. Ror2 is shown in blue, values on the right y-axis. gNM - primary gastric Normal Myofibroblasts, gCAM6 – primary gastric cancer-associated myofibroblast patient 6, gCAM13 - primary gastric cancer-associated myofibroblast patient 13, pCAF2 – pancreatic cancer-associated fibroblast cell line 2. Values shown were calculated using the 2 -ΔΔCt method and are plotted on a log10 scale. b . Protein expression of WNT5A as detected using flow cytometry. Cells as indicated on left panels. The grey histogram indicates control staining, and the black histogram indicates WNT5A staining. c . Immunofluorescence on pCAF2 cells fixed using a previously described method to preserve filopodia. The left-hand column indicates WNT5A/B antibody staining, the center panel indicates ROR2 antibody staining, and the right-hand panel indicates WNT5A/B and ROR2 antibody staining (blue and red, respectively) merged with Phalloidin FITC shown in green. Three representative images are shown. The scale bar represents 10 µm.

Article Snippet: Cells were washed again, incubated with anti-Wnt5a primary mouse monoclonal antibody clone A-5 (Santa Cruz) for 30 minutes on ice, washed in 0.5% BSA PBS buffer and incubated with goat anti-mouse IgG AF647 (Abcam) for 30 minutes on ice.

Techniques: Expressing, Flow Cytometry, Control, Staining, Immunofluorescence

Journal: bioRxiv

Article Title: Cancer-associated fibroblast-derived ROR2 induces WNT/PCP activation and polarized migration in receiving gastric cancer cells

doi: 10.1101/2022.04.07.487474

Figure Lengend Snippet: a . Confocal image of an AGS cell transiently transfected with ROR2-BFP, WNT5A-GFP, and membrane mCherry. ROR2/WNT5A/membrane complexes in a receiving, untransfected AGS cell are indicated by white arrows. The scale bar represents 10 µm. b . AGS cell expresses ROR2-mCherry and WNT5A-GFP on cytoneme tips (white arrow), which can be seen contacting a neighboring AGS receiving cell indicated by the blue dashed line (see also ). Time-lapse images, as indicated, show a ROR2/WNT5A complex leaving the cytoneme tip and co-localizing in the receiving cell. Other ROR2/WNT5A complexes from the producing cell can localize in the receiving cell, as indicated by the yellow arrow. c . pCAF2 cell expresses ROR2-mCherry and WNT5A-GFP on cytoneme tips (white arrow), which can be seen contacting a neighboring receiving AGS cell indicated by the blue dashed line (see also ). Time-lapse images, as indicated, show a ROR2/WNT5A complex leaving the cytoneme tip and co-localizing in the receiving cell. Other ROR2/WNT5A complexes from the producing cell can be seen localizing in the receiving cell, as indicated by the yellow arrow.

Article Snippet: Cells were washed again, incubated with anti-Wnt5a primary mouse monoclonal antibody clone A-5 (Santa Cruz) for 30 minutes on ice, washed in 0.5% BSA PBS buffer and incubated with goat anti-mouse IgG AF647 (Abcam) for 30 minutes on ice.

Techniques: Transfection, Membrane

Journal: bioRxiv

Article Title: Cancer-associated fibroblast-derived ROR2 induces WNT/PCP activation and polarized migration in receiving gastric cancer cells

doi: 10.1101/2022.04.07.487474

Figure Lengend Snippet: a . Cartoon depicting JNK signaling assay (adapted from . Upon activation of the JNK signaling pathway, the mCherry reporter protein translocates from the nucleus to the cytoplasm of the cell. b . Representative confocal images of AGS-B18 cells co-cultured with wild-type pCAF2 transfected with either membrane GFP (left panel), ROR2 BFP (center panel), or dCD ROR2 BFP (right panel). The scale bar represents 10μm. c . Quantification of the cytoplasmic to nuclear ratio of JNK reporter signal in receiving AGS-B18 cells co-cultured with pCAF2 cells as described in b. Boxes represent 95% quartile, the centerline indicates the mean, and whiskers indicate the range. d . Quantification of the cytoplasmic to nuclear ratio of JNK reporter signal in AGS-B18 cells co-cultured with AGS cells transfected with either membrane GFP (control), ROR2, ROR2, and WNT5A, or dominant negative ROR23i plus WNT5A as indicated. e . The intensity of ROR2-BFP was quantified in AGS-B18 receiving cells following co-culture with pCAF2 transfected with ROR2-BFP. Values were plotted against corresponding cytoplasmic/nuclear JNK reporter localization, and a simple linear regression was performed.

Article Snippet: Cells were washed again, incubated with anti-Wnt5a primary mouse monoclonal antibody clone A-5 (Santa Cruz) for 30 minutes on ice, washed in 0.5% BSA PBS buffer and incubated with goat anti-mouse IgG AF647 (Abcam) for 30 minutes on ice.

Techniques: Activation Assay, Cell Culture, Transfection, Membrane, Control, Dominant Negative Mutation, Co-Culture Assay

Journal: bioRxiv

Article Title: Cancer-associated fibroblast-derived ROR2 induces WNT/PCP activation and polarized migration in receiving gastric cancer cells

doi: 10.1101/2022.04.07.487474

Figure Lengend Snippet: ROR2/WNT5A overexpression induces a directional migration in AGS cells. a . Control AGS or AGS cells transfected with Ror2 and Wnt5a expression plasmids were cultured in Ibidi wound healing chambers for 24 hours prior to mitomycin C treatment and release. Dishes were imaged at release (0 hrs) and 18 hrs later. b . Area of the wound at 18 hours was measured at the start and end of the experiment. c . Time lapses images taken at the indicated times of pCAF2 expressing ROR2 mCherry (red cells) co-cultured with AGS cells expressing membrane GFP. An individual AGS receiving cell in close proximity to a pCAF2 cell is highlighted with a pink dot, whereas an individual AGS receiving cell in that starts at a distance to pCAF2 cells is highlighted with a light blue dot

Article Snippet: Cells were washed again, incubated with anti-Wnt5a primary mouse monoclonal antibody clone A-5 (Santa Cruz) for 30 minutes on ice, washed in 0.5% BSA PBS buffer and incubated with goat anti-mouse IgG AF647 (Abcam) for 30 minutes on ice.

Techniques: Over Expression, Migration, Control, Transfection, Expressing, Cell Culture, Membrane