Mouse Cathepsin D Antibody

Detection of Mouse Cathepsin D by Western Blot. Western blot shows lysate of mouse intestine tissue. PVDF membrane was probed with 1 µg/mL of Goat Anti-Mouse Cathepsin D Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1029) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (HAF017). Specific bands were detected for Cathepsin D at approximately 28 and 45 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Cathepsin D in RAW 264.7 Mouse Cell Line. Cathepsin D was detected in immersion fixed RAW 264.7 mouse monocyte/macrophage cell line using Goat Anti-Mouse Cathepsin D Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1029) at 5 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; NL001) and counterstained with DAPI (blue). Specific staining was localized to lysosomes. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

Cathepsin D in Mouse Intestine. Cathepsin D was detected in perfusion fixed frozen sections of mouse intestine using 15 µg/mL Goat Anti-Mouse Cathepsin D Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1029) overnight at 4 °C. Tissue was stained with the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; CTS008) and counter-stained with hematoxylin (blue). Specific labeling was localized to epithelial cells in intestinal villi. View our protocol for Chromogenic IHC Staining of Frozen Tissue Sections.

Detection of Human Cathepsin D by Western Blot Chronic treatment with BACE1 inhibitors causes accumulation of CatD.(a) Representative western blot of brain homogenates prepared from mice dosed daily for 30 days with PF-9283 (80 mg kg−1) showed increased levels of CatD heavy and light chains, but not the CatD single chain, compared with a vehicle control. Data are presented as mean (normalized to actin)±s.e.m.; n=5 (PF-9283) and n=6 (veh) per group. (b) ELISA of live ARPE-19 cells incubated with BACE1 inhibitors for 7 days showed accumulation of CatD; the extent of accumulation was consistent with cellular CatD IC50 values (Supplementary Table 4). Data are presented as mean±s.e.m. for three independent experiments. (c) Proteomic analysis of RPE tissue prepared from rats dosed daily for 14 days with 80 mg kg−1 PF-9283. Peptide spectral matches (PSM) of 11 of 391 identified proteins were changed more than 1.5-fold (P<0.01, red symbols, see also Supplementary Table 5) compared with vehicle controls including a 1.7-fold increase in CatD (***P=0.0002). Average PSM were calculated from six LC-MS runs consisting of two technical replicates of n=3 RPE tissues per group. For a–c, significant differences were determined using a two-tailed t-test; *P<0.05, **P<0.01, ***P<0.001. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/27727204), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human Cathepsin D by Western Blot Chronic treatment with BACE1 inhibitors causes accumulation of CatD.(a) Representative western blot of brain homogenates prepared from mice dosed daily for 30 days with PF-9283 (80 mg kg−1) showed increased levels of CatD heavy and light chains, but not the CatD single chain, compared with a vehicle control. Data are presented as mean (normalized to actin)±s.e.m.; n=5 (PF-9283) and n=6 (veh) per group. (b) ELISA of live ARPE-19 cells incubated with BACE1 inhibitors for 7 days showed accumulation of CatD; the extent of accumulation was consistent with cellular CatD IC50 values (Supplementary Table 4). Data are presented as mean±s.e.m. for three independent experiments. (c) Proteomic analysis of RPE tissue prepared from rats dosed daily for 14 days with 80 mg kg−1 PF-9283. Peptide spectral matches (PSM) of 11 of 391 identified proteins were changed more than 1.5-fold (P<0.01, red symbols, see also Supplementary Table 5) compared with vehicle controls including a 1.7-fold increase in CatD (***P=0.0002). Average PSM were calculated from six LC-MS runs consisting of two technical replicates of n=3 RPE tissues per group. For a–c, significant differences were determined using a two-tailed t-test; *P<0.05, **P<0.01, ***P<0.001. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/27727204), licensed under a CC-BY license. Not internally tested by R&D Systems.