Human/Mouse Brachyury Antibody Best Seller

Brachyury in BG01V Human Stem Cells.

Brachyury was detected in immersion fixed BG01V human embryonic stem cells differentiated into mesoderm using Goat Anti-Human Brachyury Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2085) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the Northern-Lights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; NL001) and counter-stained with DAPI (blue). Specific staining was localized to nuclei. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

Brachyury in Embryonic Mouse Notochord.

Brachyury was detected in immersion fixed frozen sections of embryonic mouse notochord (E9.5) using 10 µg/mL Goat Anti-Human Brachyury Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2085) overnight at 4 °C. Tissue was stained with the Northern-Lights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; NL001) and counter-stained with DAPI (blue). View our protocol for Fluorescent IHC Staining of Frozen Tissue Sections.

Detection of Brachyury-regulated Genes by Chromatin Immunoprecipitation.

Mesoderm-differentiated BG01V human embryonic stem cells were fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. Brachyury/DNA complexes were immunoprecipitated using 5 µg Goat Anti-Human/Mouse Brachyury Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2085) or control antibody (AB-108-C) for 15 minutes in an ultrasonic bath, followed by Biotinylated Anti-Goat IgG Secondary Antibody (BAF109). Immunocomplexes were captured using 50 µL of MagCellect Streptavidin Ferrofluid (MAG999) and DNA was purified using chelating resin solution. TheVEGFpromoter was detected by standard PCR.

Detection of Brachyury in NCI-H460 cells by Flow Cytometry.

NCI-H460 cells were stained with Goat Anti-Human/Mouse Brachyury Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2085, filled histogram) or isotype control antibody (Catalog # AB-108-C, open histogram), followed by Phycoerythrin-conjugated Anti-Goat IgG Secondary Antibody (Catalog # F0107). To facilitate intracellular staining, cells were fixed and permeabilized with FlowX FoxP3 Fixation & Permeabilization Buffer Kit (Catalog # FC012). View our protocol for Staining Intracellular Molecules.

Detection of Human Brachyury by Immunocytochemistry/Immunofluorescence

Generation and characterization of systemic lupus erythematosus (SLE)‐specific human‐induced pluripotent stem cells (hiPSCs). (A) Dermal fibroblasts derived from patient with SLE were reprogrammed into iPSCs using Sendai virus vectors and three clones (#1, #2 and #3) were characterized. RT‐PCR confirms the loss of transgenes in hiPSCs‐SLE (lanes 1, 2 and 3), presence (lane 4) in infected fibroblasts (ipF‐SLE) and absence of Sendai viral transgenes in parental fibroblasts (pF‐SLE) (lane 5). Full‐length gels are presented in File S2. (B) The hiPSCs‐SLE colonies expressed alkaline phosphatase. Scale bar, 500 μm. (C) RT‐qPCR analysis of pluripotency genes OCT4, NANOG, SOX2 and REX1 was performed in fibroblasts and in hiPSCs derived from patient with SLE. All expression values are normalized to GAPDH and relative donor fibroblasts. Data are mean ± SEM and all statistical analysis was made between hiPSCs‐SLE clones and relative fibroblasts by Student's t test showing P‐values ≤ .05 in each comparison. (D) PluriTest assays combines novelty score (blue) on x‐axis and pluripotency score (red) on y‐axis. hiPSCs‐SLE localize in the red cloud suggesting the empirical distribution of pluripotent cells compared to non‐pluripotent blue cloud. (E) Representative images of M‐FISH staining show normal karyotypes of hiPSCs‐SLE clones. (F) Immunofluorescence analysis of pluripotent stem cell markers Nanog (green), Oct4 (red) and co‐staining with DAPI (blue) in hiPSCs‐SLE. Scale bar, 50 μm. (G) Representative images of floating and adherent EBs derived from hiPSCs‐SLE at differentiation day 8 and 18, respectively. Scale bar, 500 μm. (H) RT‐qPCR results confirm the capability of hiPSCs‐SLE to differentiate into all three germ layers. The expression levels of GATA4, HAND1 and PAX6 in EBs are relative to undifferentiated hiPSCs. All expression values are normalized to GAPDH and relative hiPSCs. Data are mean ± SEM and all statistical analysis was made between EBs‐SLE and relative hiPSCs‐SLE clones by Student's t test showing P‐values ≤ .05 in each comparison. (I) Immunostaining of Otx2 (ectoderm marker), Sox17 (endoderm marker), Brachyury (mesoderm marker) and co‐staining with DAPI (blue) in EBs derived from hiPSCs‐SLE. Scale bar, 50 μm Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/31536674), licensed under a CC-BY license. Not internally tested by R&D Systems.