Intracellular Staining by Flow Cytometry
|Detection of Patched 2 in WS‑1 Human Cell Line by Flow Cytometry. WS‑1 human fetal skin fibroblast cell line was stained with Human Patched 2 (First Extracellular Loop) Antigen Affinity‑purified Polyclonal Antibody (Catalog # AF4078, filled histogram) or isotype control antibody (Catalog # AB-108-C, open histogram), followed by Allophycocyanin-conjugated Anti-Goat IgG Secondary Antibody (Catalog # F0108). To facilitate intracellular staining, cells were fixed with paraformaldehyde and permeabilized with saponin.|
|Patched 2 in Human Basal Cell Carcinoma. Patched 2 was detected in immersion fixed paraffin-embedded sections of human basal cell carcinoma using 3 µg/mL Goat Anti-Human Patched 2 (First Extracellular Loop) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF4078) overnight at 4 °C. Before incubation with the primary antibody tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained with the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS008) and counterstained with hematoxylin (blue). Specific labeling was localized to the cytoplasm of keratinocytes. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.|
Patched 2 (Ptch2) is a 180-200 kDa 12 transmembrane member of the Patched family. Human PTCH2 is 1203 amino acids (aa) in length. It has two terminal cytoplasmic domains, one between aa 1-57, and a second between aa 1115-1203. Together with Smoothened (SMO), Ptch2 forms a membrane complex that serves as a receptor for Sonic Hedgehog (SHH). Human Ptch2 likely has two isoforms. One shows an eight aa extension of the C-terminus, while a second shows a four aa substitution for the C-terminal 61 amino acids (aa 1143-1203). Over the first extracellular loop human Patched 2 shares 95% and 96% aa sequence identity with mouse and canine Patched 2, respectively.