Mouse/Rat Cathepsin L Antibody

Cathepsin L in Mouse Thymus.

Cathepsin L was detected in perfusion fixed frozen sections of mouse thymus using 15 µg/mL Goat Anti-Mouse Cathepsin L Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1515) overnight at 4 °C. Tissue was stained with the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; CTS008) and counterstained with hematoxylin (blue). View our protocol for Chromogenic IHC Staining of Frozen Tissue Sections.

Detection of Mouse Cathepsin L by Immunocytochemistry/ Immunofluorescence

Myc induces cathepsin L expression in beta-cells of pancreatic Islets.(A) Immunohistochemical analyses for CTS B, C, L or S expression (all in red) in combination with staining for the pan-leukocyte marker CD45 (green) in pancreatic islet tumors from the MycERTAM;Bcl-xL animals. Pancreata were harvested from the MycERTAM;Bcl-xL mice treated for 7 d with TAM (Myc-On, 7 days) or control vehicle in place of TAM (Myc-OFF). The islet area is indicated by dotted line. The asterisks indicate the area of tumor represented in the insets. The panels are representatives of at least three animals assayed at each data point, all immunohistochemical analyses were done in duplicate; eight randomized fields per analysis were examined. Scale bars, 100μm. (B) Immunohistochemical analysis for cathepsin L expression in beta-cells of pancreatic islets from MycERTAM;Bcl-xL animals identified by insulin expression. Pancreata were collected from the animals described above. Scale bars represent 25μm. The panels are representatives of three animals assayed at each data point, all immunohistochemical analyses were done in duplicate; ten randomized fields per analysis were examined. Image collected and cropped by CiteAb from the following open publication (https://dx.plos.org/10.1371/journal.pone.0120348), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse Cathepsin L by Immunocytochemistry/ Immunofluorescence

Myc induces cathepsin L expression in beta-cells of pancreatic Islets.(A) Immunohistochemical analyses for CTS B, C, L or S expression (all in red) in combination with staining for the pan-leukocyte marker CD45 (green) in pancreatic islet tumors from the MycERTAM;Bcl-xL animals. Pancreata were harvested from the MycERTAM;Bcl-xL mice treated for 7 d with TAM (Myc-On, 7 days) or control vehicle in place of TAM (Myc-OFF). The islet area is indicated by dotted line. The asterisks indicate the area of tumor represented in the insets. The panels are representatives of at least three animals assayed at each data point, all immunohistochemical analyses were done in duplicate; eight randomized fields per analysis were examined. Scale bars, 100μm. (B) Immunohistochemical analysis for cathepsin L expression in beta-cells of pancreatic islets from MycERTAM;Bcl-xL animals identified by insulin expression. Pancreata were collected from the animals described above. Scale bars represent 25μm. The panels are representatives of three animals assayed at each data point, all immunohistochemical analyses were done in duplicate; ten randomized fields per analysis were examined. Image collected and cropped by CiteAb from the following open publication (https://dx.plos.org/10.1371/journal.pone.0120348), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse Cathepsin L by Immunocytochemistry/ Immunofluorescence

Myc induces cathepsin L expression in beta-cells of pancreatic Islets.(A) Immunohistochemical analyses for CTS B, C, L or S expression (all in red) in combination with staining for the pan-leukocyte marker CD45 (green) in pancreatic islet tumors from the MycERTAM;Bcl-xL animals. Pancreata were harvested from the MycERTAM;Bcl-xL mice treated for 7 d with TAM (Myc-On, 7 days) or control vehicle in place of TAM (Myc-OFF). The islet area is indicated by dotted line. The asterisks indicate the area of tumor represented in the insets. The panels are representatives of at least three animals assayed at each data point, all immunohistochemical analyses were done in duplicate; eight randomized fields per analysis were examined. Scale bars, 100μm. (B) Immunohistochemical analysis for cathepsin L expression in beta-cells of pancreatic islets from MycERTAM;Bcl-xL animals identified by insulin expression. Pancreata were collected from the animals described above. Scale bars represent 25μm. The panels are representatives of three animals assayed at each data point, all immunohistochemical analyses were done in duplicate; ten randomized fields per analysis were examined. Image collected and cropped by CiteAb from the following open publication (https://dx.plos.org/10.1371/journal.pone.0120348), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Cathepsin L by Western Blot

Acute, first window RIC activates autophagy signaling via p-AMPK upregulation and concomitant downregulation of mTOR.(A) Western blots for autophagy related signaling proteins. (B) Quantification of the protein fold change in 1W RIC compared to 1W controls. Values are means ± S.E.M. n = 6–8 per group. An (*) denotes a statistically significant difference (P<0.05) compared to control. (P-: phospho-). Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/25347774), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Cathepsin L by Western Blot

Chronic, third window RIC increases the expression of autophagosome proteins, LC3I/II and Atg5.(A) Western blots for autophagy related signaling proteins. (B) Quantification of the protein fold change in 3W RIC compared to 3W controls. Values are means ± S.E.M. n = 6–8 per group. An (*) denotes a statistically significant difference (P<0.05) compared to control. (P-: phospho-). Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/25347774), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Cathepsin L by Western Blot

Inhibition of CTSV induces cellular senescence. (A) HUVECs were treated with si-negative control (NC), si-CTSV, si-ALDH1A2 or si-CTSV + si-ALDH1A2 for 24 h and simulated with 100 nM doxorubicin (DOX) for 24 h after incubated in complete growth medium for 24 h. SA-beta -gal activity was analyzed (blue staining for the senescent cells, scale bar = 50 μm, n = 4). (B) EdU assay of the cell proliferation ability in HUVECs treated with DOX, si-CTSV and si-ALDH1A2 (red staining for the EdU, blue staining for Hoechst, scale bar = 25 μm, n = 4). (C) Representative image of the wound healing assay and the quantification of wound area in HUVECs (scale bar = 100 μm, n = 4). (D) Western blot of CTSV, ALDH1A2, P21, p-ERK1/2, ERK1/2, p-AKT, AKT, CTSL, P53 and P16 in HUVECs treated with DOX and siRNAs (n = 4). (E) Relative mRNA level of CTSV, ALDH1A2, P21, IL-1 beta, IL-6 and ICAM-1 (n = 4). (F) RA concentration in HUVECs treated with DOX and siRNAs (n = 4). Data are presented as mean ± SEM. One-way ANOVA test was used. * 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/36671735), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Cathepsin L by Immunohistochemistry

Ctsl expression is decreased in senescent aorta. (A) Representative images of hematoxylin-eosin staining in the thoracic aortas of aged (18-month-old) and young (8-week-old) mice (scale bar = 200 μm in top and 50 μm in bottom, respectively). (B) Representative images of Masson’s trichrome stain in the thoracic aortas (blue-gray staining for the collagens, scale bar = 200 μm in top and 50 μm in bottom, respectively). (C) Quantification of media thickness (MT), the ratio of media thickness to lumen diameter (MT/LD), media area (MA), and the ratio of collagen to media area (collagen/MA) using hematoxylin-eosin staining and Masson’s trichrome stain (n = 4). (D). Immunohistochemistry of Ctsl in the thoracic aortas of aged and young mice (brown staining for Ctsl, scale bar = 50 μm, n = 4). (E) Western Blot of Ctsl and p21 in young and senescent aortas (n = 3–4). Data were presented as mean ± SEM. T-test was used. * p < 0.05 vs. young, ** p < 0.01 vs. young, *** p < 0.001 vs. young. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36671735), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Cathepsin L by Western Blot

CTSV attenuates DOX-induced cellular senescence. (A). HUVECs were treated with ad-negative control (NC) or ad-CTSV (MOI = 10) for 8 h and simulated with 100 nM doxorubicin (DOX) and 1 μM RA for 24 h after incubated in complete growth medium for 40 h. SA-beta -gal activity was analyzed (blue staining for the senescent cells, scale bar = 50 μm, n = 4). (B) EdU assay of the cell proliferation ability in HUVECs treated with DOX, RA and ad-CTSV (red staining for the EdU, blue staining for Hoechst, scale bar = 25 μm, n = 4). (C) Representative image of the wound healing assay and the quantification of wound area in HUVECs (scale bar = 100 μm, n = 4). (D) Western blot of CTSV, ALDH1A2, P21, p-ERK1/2, ERK1/2, p-AKT, AKT, CTSL, P53, and P16 in HUVECs treated with DOX, RA and ad-CTSV (n = 4). (E) Relative mRNA level of CTSV, ALDH1A2, P21, IL-1 beta, IL-6, and ICAM-1 (n = 4). (F) RA concentration in HUVECs treated with DOX, RA, and ad-CTSV (n = 4). Data are presented as mean ± SEM. One-way ANOVA test was used. * 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/36671735), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Cathepsin L by Immunohistochemistry

Ctsl expression is decreased in senescent aorta. (A) Representative images of hematoxylin-eosin staining in the thoracic aortas of aged (18-month-old) and young (8-week-old) mice (scale bar = 200 μm in top and 50 μm in bottom, respectively). (B) Representative images of Masson’s trichrome stain in the thoracic aortas (blue-gray staining for the collagens, scale bar = 200 μm in top and 50 μm in bottom, respectively). (C) Quantification of media thickness (MT), the ratio of media thickness to lumen diameter (MT/LD), media area (MA), and the ratio of collagen to media area (collagen/MA) using hematoxylin-eosin staining and Masson’s trichrome stain (n = 4). (D). Immunohistochemistry of Ctsl in the thoracic aortas of aged and young mice (brown staining for Ctsl, scale bar = 50 μm, n = 4). (E) Western Blot of Ctsl and p21 in young and senescent aortas (n = 3–4). Data were presented as mean ± SEM. T-test was used. * p < 0.05 vs. young, ** p < 0.01 vs. young, *** p < 0.001 vs. young. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36671735), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Cathepsin L by Western Blot

Inhibition of CTSV induces cellular senescence. (A) HUVECs were treated with si-negative control (NC), si-CTSV, si-ALDH1A2 or si-CTSV + si-ALDH1A2 for 24 h and simulated with 100 nM doxorubicin (DOX) for 24 h after incubated in complete growth medium for 24 h. SA-beta -gal activity was analyzed (blue staining for the senescent cells, scale bar = 50 μm, n = 4). (B) EdU assay of the cell proliferation ability in HUVECs treated with DOX, si-CTSV and si-ALDH1A2 (red staining for the EdU, blue staining for Hoechst, scale bar = 25 μm, n = 4). (C) Representative image of the wound healing assay and the quantification of wound area in HUVECs (scale bar = 100 μm, n = 4). (D) Western blot of CTSV, ALDH1A2, P21, p-ERK1/2, ERK1/2, p-AKT, AKT, CTSL, P53 and P16 in HUVECs treated with DOX and siRNAs (n = 4). (E) Relative mRNA level of CTSV, ALDH1A2, P21, IL-1 beta, IL-6 and ICAM-1 (n = 4). (F) RA concentration in HUVECs treated with DOX and siRNAs (n = 4). Data are presented as mean ± SEM. One-way ANOVA test was used. * 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/36671735), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Cathepsin L by Western Blot

CTSV attenuates DOX-induced cellular senescence. (A). HUVECs were treated with ad-negative control (NC) or ad-CTSV (MOI = 10) for 8 h and simulated with 100 nM doxorubicin (DOX) and 1 μM RA for 24 h after incubated in complete growth medium for 40 h. SA-beta -gal activity was analyzed (blue staining for the senescent cells, scale bar = 50 μm, n = 4). (B) EdU assay of the cell proliferation ability in HUVECs treated with DOX, RA and ad-CTSV (red staining for the EdU, blue staining for Hoechst, scale bar = 25 μm, n = 4). (C) Representative image of the wound healing assay and the quantification of wound area in HUVECs (scale bar = 100 μm, n = 4). (D) Western blot of CTSV, ALDH1A2, P21, p-ERK1/2, ERK1/2, p-AKT, AKT, CTSL, P53, and P16 in HUVECs treated with DOX, RA and ad-CTSV (n = 4). (E) Relative mRNA level of CTSV, ALDH1A2, P21, IL-1 beta, IL-6, and ICAM-1 (n = 4). (F) RA concentration in HUVECs treated with DOX, RA, and ad-CTSV (n = 4). Data are presented as mean ± SEM. One-way ANOVA test was used. * 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/36671735), licensed under a CC-BY license. Not internally tested by R&D Systems.