Anti-Wnt1 antibody

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Wnt1 antibody (AB15251)

Breast carcinoma cells stained with ab15251 (Immunohistochemistry, paraffin sections);

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Wnt1 antibody (AB15251)

ICC/IF image of ab15251 stained mouse 3T3 cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab15251, 1μg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iT^TM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Wnt1 antibody (AB15251)

ab15251 staining Wnt1 in breast carcinoma by Immunohistochemistry (FFPE-sections).

• WB

Unknown

Western blot - Anti-Wnt1 antibody (AB15251)

All lanes:

Western blot - Anti-Wnt1 antibody (ab15251) at 1/25 dilution

All lanes:

NIH3T3 cell lysate

Predicted band size: 40 kDa

Observed band size: 41 kDa

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• IHC

CiteAb

Immunohistochemistry - Anti-Wnt1 antibody (AB15251)

Immunohistochemistry-immunofluorescence using Anti-Wnt1 antibody, ab15251. Publication image from Eiraku, M. et al., 2017, Nat Commun, 29109536. Legend direct from paper.

Genome-wide analysis of the polarized tissue reveals differential expression of Wnt signaling components in the caudal region. a Fluorescent images of a day-5 aggregate using a Fgf5 : : Turq//Six3 : : Venus//Irx3 : : Tomato ESC line. Fgf5 : : Turq signals were enhanced by the auto contrast function of ImageJ software due to its dim expression at culture day 5. b Schematic diagram of microarray sample collection. c A validation of microarray samples via RT-qPCR, showing rax and irx3 expression in FACS-sorted GFP+ and GFP− cells. d Volcano plot drawn by plotting points of all probe sets with log2 fold-changes of GFP− expression values to GFP+ (X-axis) and p-values of significant differences (Y-axis). Blue dots indicate log2 fold-change of more than 1 (i.e., more than two-fold). e, f Quantification of Wnt and Fgf signaling-related-gene expressions via RT-qPCR, following FACS sorting of GFP+ and GFP- cells. g Immunohistochemistry was performed on cryosections of day-5 Rax : : GFP aggregates with antibodies recognizing Wnt1, Irx3 and GFP, showing Wnt1, Irx3 and Rax : : GFP signals. h, i Quantification of gene expressions via RT-qPCR, following time-specific treatment of bFGF and PD173074. j Schematic of Fgf/FGFR and Wnt signaling relationship in ESC-derived caudal differentiation in vitro. Error bars indicate s.e.m of each experiment (c, e, f, h, i). Significance was determined using unpaired t-test. *P < 0.05; **P < 0.01; ***P < 0.001; ns, not significant (e, f, h, i). Scale bars, 100 µm (a, g). These images were one of n = 3 experiments (a, g)

• IHC

CiteAb

Immunohistochemistry - Anti-Wnt1 antibody (AB15251)

Immunohistochemistry-immunofluorescence using Anti-Wnt1 antibody, ab15251. Publication image from Eiraku, M. et al., 2017, Nat Commun, 29109536. Legend direct from paper.

Wnt signaling components wnt1/4/8b, sfrp1, dkk1 and axin2 are spatially distributed along the R-C polarized neuroectoderm. a Montage of images taken from Supplementary Movie 5, showing upper panel; merged image of DIC (differential interference contrast), Rax : : GFP and 7TCF : : Cherry, lower panel; merged image of Rax : : GFP and 7TCF : : Cherry. b Kymograph analysis of time-lapse images by culture day 7, showing Rax : : GFP and 7TCF : : Cherry. White solid line indicates the time when Rax : : GFP+ signals started to polarize. c Four way fractionation of day-6 cells by FACS, depending on Rax : : GFP expression levels. d Confirmation of four fractions via FACS. e Quantification of Wnt signaling-related gene expression via RT-qPCR : wnt1, wnt4, wnt8b, axin2, dkk1, and sfrp1. Error bars indicate s.e.m of each FACS sorting experiment. f, g, h Immunohistochemistry was performed on cryosections of day-6 aggregates, showing Wnt1, 7TCF : : H2B-Tomato, Sfrp1, Dkk1, Six3 : : Venus and Irx3 : : Tomato expressions. Dkk1 signals in Fig. 5h were also used to show in green in Supplementary Fig. 5J. i Schematic diagram of Wnt component distribution. R, rostral; C, caudal. Scale bars, 100 µm (a, f, g). These images were one of n = 3 experiments (a, f, g, h)

• WB

CiteAb

Western blot - Anti-Wnt1 antibody (AB15251)

Western Blotting using Anti-Wnt1 antibody, ab15251. Publication image from Soumelis, V. et al., 2019, Nat Commun, 30926812. Legend direct from paper.

Wnt1 silencing attenuates Wnt pathway activation in human LUAD cDCs. a Human CD1−cDCs were sorted from primary lung adenocarcinomas and paired juxta-tumor healthy lung and analyzed by RNAseq. Gene Set Enrichment Analysis (GSEA) enrichment plot (n = 4 per group). b Primary human lung adenocarcinomas were dispersed and total cells cultured in the presence of Wnt1-silencing RNA (siWnt1) or control siRNA loaded nanoparticles (Up). Wnt1 silencing was confirmed by western blot (bottom). Representative histograms showing active b-catenin in cDCs (up) and extracellular CD107 in CD8+ T cells (bottom). Data are representative of 3 independent experiments. Source data are provided as a Source Data file. Sequencing data are available with the accession code GSE124199

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• WB

CiteAb

Western blot - Anti-Wnt1 antibody (AB15251)

Western Blotting using Anti-Wnt1 antibody, ab15251. Publication image from Soumelis, V. et al., 2019, Nat Commun, 30926812. Legend direct from paper.

Wnt1 impairs adaptive immune surveillance in syngeneic models. a Expression of Wnt1 and active b-catenin (western blot) in LLC cells transduced with Wnt1 (Wnt1) or Empty (Empty) viral vectors (Up). In vitro proliferation (MTT assay) (Bottom). b, c Tumor burden (total absolute number of LLC cells) after intrathoracic (i.t.) implantation or intravenous (i.v.) administration. d Cellular profiles of lung tumors. Pies depict mean percentages among CD45+ cells. e Numbers of intratumoral cDCs, CD8 T, CD4 T, and B cells per cancer cell. f Tumor growth after subcutaneous implantation of ovalbumin (OVA)-LLC cells in immunocompetent vs. immunodeficient RAG mice. g Numbers of OVA-specific (dextramer+) T cytotoxic cells per cancer cell and T cell CD44 expression. h Expression of Wnt1 and active b-catenin (western blot) in LLC cells transduced with shWnt1 or Scramble viral vectors (left). In vitro proliferation (MTT assay) (right). i Tumor burden (total absolute number of OVA-LLC cells) after intrathoracic (i.t.) implantation in untreated vs. aCD8-treated mice (left). Numbers of dextramer+ T cytotoxic cells per cancer cell and T cell PD1 expression (Right). a–i Error bars represent mean with SEM. b–g, i Cell numbers and profiles were assessed by FACS. Data are representative or cumulative of at least two independent experiments with 5–9 mice per group. *p < 0.05; **p < 0.01, Mann–Whitney. Source data are provided as a Source Data file

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