Western blot shows lysates of mouse embryo tissue. PVDF membrane was probed with 1 µg/mL of Goat Anti-Human/Mouse GLI‑3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3690) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF019). A specific band was detected for GLI‑3 at approximately 85 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 8.
GLI‑3 was detected in immersion fixed HeLa human cervical epithelial carcinoma cell line using Human/Mouse GLI‑3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3690) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (yellow; Catalog # NL001) and counterstained with DAPI (blue). View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
Jurkat human acute T cell leukemia cell line treated with 50 ng/mL PMA and 200 ng/mL calcium ionomycin for 30 minutes was fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. GLI‑3/DNA complexes were immunoprecipitated using 5 μg Goat Anti-Human/Mouse GLI‑3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3690) or control antibody (Catalog # AB-108-C) for 15 minutes in an ultrasonic bath, followed by Biotinylated Anti-Goat IgG Secondary Antibody (Catalog # BAF109). Immunocomplexes were captured using 50 μL of MagCellect Streptavidin Ferrofluid (Catalog # MAG999) and DNA was purified using chelating resin solution. The gli-1 promoter was detected by standard PCR.