BD PharmingenTM Purified Rabbit Anti-Active Caspase-3

Description

Flow cytometric analysis of Active Caspase-3 expression by cells treated to induce apoptosis. Human Jurkat Cells - Cells from the Human Jurkat T-cell leukemia, ATCC TIB-152 cell line were left Untreated or Treated 6 h with 12 muM Camptothecin to induce apoptosis as indicated. Cells were washed, fixed with BD Cytofix Fixation Buffer Cat. No. 554655 and permeabilized with BD Perm/Wash Buffer Cat. No. 554723 . The cells were subsequently stained with Purified Rabbit Anti-Active Caspase-3 antibody Cat. No. 570524/570525 at 0.125 mug/test and secondarily stained with FITC Goat Anti-Rabbit IgG FITC-Anti-Ig; Cat. No. 554020 at 0.06 mug/test. The fluorescence histograms showing Active Caspase-3 expression were derived from gated events with the light-scatter characteristics of single cells. Mouse Thymocytes - C57BL/6 Mouse thymocytes were left Untreated or treated 5 h with 1 muM of Dexamethasone to induce apoptosis as indicated. The cells were similarly washed, fixed, permeabilized and stained as the Human Jurkat T cells. The fluorescence histograms showing Active Caspase-3 expression were derived from gated events with the light-scatter characteristics of single thymocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa X-20 Cell Analyzer System and FlowJo software. Data shown on this Technical Data Sheet are not lot specific. Analyses of Active Caspase-3 expression by Jurkat cells treated to induce apoptosis. Left Panel: Immunofluorescent Analysis: Jurkat cells T-cell leukemia, ATCC TIB-152 were left Untreated Left Image or Treated Right Image with 6 muM Camptothecin for 4 hours to induce apoptosis. Cytospins of the Jurkat cells were fixed with BD Cytofix Fixation Buffer Cat. No. 554655 and permeabilized with BD Perm/Wash Buffer Cat. No. 554723 . and stained with Purified Rabbit Anti-Active Caspase-3 antibody Cat. No. 570524/570525 . After washing, cells were secondarily stained with BD Horizon BV480 Goat Anti-Rabbit IgG antibody Cat. No. 564879; pseudocolored green and with DRAQ7 cat. no. 564904, pseudocolored red to stain nuclei. Active Caspase-3 staining was almost exclusively identified in cells treated to undergo apoptosis. Images acquired with 20x objective. Right Panel: Immunoprecipitation and Western blot analysis of Active Caspase-3 present in lysates from Human Jurkat cells treated to induce apoptosis. Cells from the Human Jurkat T-cell leukemia, ATCC TIB-152 cell line were left Untreated Left Panel or Treated with 12 muM of Camptothecin for 6 hours to induce apoptosis Right Panel . Immunoprecipitation: Cell lysates were immunoprecipitated with 2 or 4 mug/ml of Purified Rabbit Anti-Active Caspase-3 antibody Cat. No. 570524/570525 , Lanes 1 and 3 4 mug and lanes 2 and 4 2 mug . Western blot: Active Caspase-3 was detected by Western blot analysis with an antibody that recognizes both the Pro-enzyme and Active Caspase-3 forms, reflecting the presence or absence of the caspase-3 pro-domain lanes 1-4 . The results show that the Purified Rabbit Anti-Active Caspase-3 antibody immunoprecipitated only the active form of Caspase-3. Flow cytometric analysis of Active Caspase-3 expression by cells treated to induce apoptosis. Human Jurkat Cells - Cells from the Human Jurkat T-cell leukemia, ATCC TIB-152 cell line were left Untreated or Treated 6 h with 12 muM Camptothecin to induce apoptosis as indicated. Cells were washed, fixed with BD Cytofix Fixation Buffer Cat. No. 554655 and permeabilized with BD Perm/Wash Buffer Cat. No. 554723 . The cells were subsequently stained with Purified Rabbit Anti-Active Caspase-3 antibody Cat. No. 570524/570525 at 0.125 mug/test and secondarily stained with FITC Goat Anti-Rabbit IgG FITC-Anti-Ig; Cat. No. 554020 at 0.06 mug/test. The fluorescence histograms showing Active Caspase-3 expression were derived from gated events with the light-scatter characteristics of single cells. Mouse Thymocytes - C57BL/6 Mouse thymocytes were left Untreated or treated 5 h with 1 muM of Dexamethasone to induce apoptosis as indicated. The cells were similarly washed, fixed, permeabilized and stained as the Human Jurkat T cells. The fluorescence histograms showing Active Caspase-3 expression were derived from gated events with the light-scatter characteristics of single thymocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa X-20 Cell Analyzer System and FlowJo software. Data shown on this Technical Data Sheet are not lot specific. Show More Analyses of Active Caspase-3 expression by Jurkat cells treated to induce apoptosis. Left Panel: Immunofluorescent Analysis: Jurkat cells T-cell leukemia, ATCC TIB-152 were left Untreated Left Image or Treated Right Image with 6 muM Camptothecin for 4 hours to induce apoptosis. Cytospins of the Jurkat cells were fixed with BD Cytofix Fixation Buffer Cat. No. 554655 and permeabilized with BD Perm/Wash Buffer Cat. No. 554723 . and stained with Purified Rabbit Anti-Active Caspase-3 antibody Cat. No. 570524/570525 . After washing, cells were secondarily stained with BD Horizon BV480 Goat Anti-Rabbit IgG antibody Cat. No. 564879; pseudocolored green and with DRAQ7 cat. no. 564904, pseudocolored red to stain nuclei. Active Caspase-3 staining was almost exclusively identified in cells treated to undergo apoptosis. Images acquired with 20x objective. Right Panel: Immunoprecipitation and Western blot analysis of Active Caspase-3 present in lysates from Human Jurkat cells treated to induce apoptosis. Cells from the Human Jurkat T-cell leukemia, ATCC TIB-152 cell line were left Untreated Left Panel or Treated with 12 muM of Camptothecin for 6 hours to induce apoptosis Right Panel . Immunoprecipitation: Cell lysates were immunoprecipitated with 2 or 4 mug/ml of Purified Rabbit Anti-Active Caspase-3 antibody Cat. No. 570524/570525 , Lanes 1 and 3 4 mug and lanes 2 and 4 2 mug . Western blot: Active Caspase-3 was detected by Western blot analysis with an antibody that recognizes both the Pro-enzyme and Active Caspase-3 forms, reflecting the presence or absence of the caspase-3 pro-domain lanes 1-4 . The results show that the Purified Rabbit Anti-Active Caspase-3 antibody immunoprecipitated only the active form of Caspase-3. Show More Flow cytometric analysis of Active Caspase-3 expression by cells treated to induce apoptosis. Human Jurkat Cells - Cells from the Human Jurkat T-cell leukemia, ATCC TIB-152 cell line were left Untreated or Treated 6 h with 12 muM Camptothecin to induce apoptosis as indicated. Cells were washed, fixed with BD Cytofix Fixation Buffer Cat. No. 554655 and permeabilized with BD Perm/Wash Buffer Cat. No. 554723 . The cells were subsequently stained with Purified Rabbit Anti-Active Caspase-3 antibody Cat. No. 570524/570525 at 0.125 mug/test and secondarily stained with FITC Goat Anti-Rabbit IgG FITC-Anti-Ig; Cat. No. 554020 at 0.06 mug/test. The fluorescence histograms showing Active Caspase-3 expression were derived from gated events with the light-scatter characteristics of single cells. Mouse Thymocytes - C57BL/6 Mouse thymocytes were left Untreated or treated 5 h with 1 muM of Dexamethasone to induce apoptosis as indicated. The cells were similarly washed, fixed, permeabilized and stained as the Human Jurkat T cells. The fluorescence histograms showing Active Caspase-3 expression were derived from gated events with the light-scatter characteristics of single thymocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa X-20 Cell Analyzer System and FlowJo software. Data shown on this Technical Data Sheet are not lot specific. Analyses of Active Caspase-3 expression by Jurkat cells treated to induce apoptosis. Left Panel: Immunofluorescent Analysis: Jurkat cells T-cell leukemia, ATCC TIB-152 were left Untreated Left Image or Treated Right Image with 6 muM Camptothecin for 4 hours to induce apoptosis. Cytospins of the Jurkat cells were fixed with BD Cytofix Fixation Buffer Cat. No. 554655 and permeabilized with BD Perm/Wash Buffer Cat. No. 554723 . and stained with Purified Rabbit Anti-Active Caspase-3 antibody Cat. No. 570524/570525 . After washing, cells were secondarily stained with BD Horizon BV480 Goat Anti-Rabbit IgG antibody Cat. No. 564879; pseudocolored green and with DRAQ7 cat. no. 564904, pseudocolored red to stain nuclei. Active Caspase-3 staining was almost exclusively identified in cells treated to undergo apoptosis. Images acquired with 20x objective. Right Panel: Immunoprecipitation and Western blot analysis of Active Caspase-3 present in lysates from Human Jurkat cells treated to induce apoptosis. Cells from the Human Jurkat T-cell leukemia, ATCC TIB-152 cell line were left Untreated Left Panel or Treated with 12 muM of Camptothecin for 6 hours to induce apoptosis Right Panel . Immunoprecipitation: Cell lysates were immunoprecipitated with 2 or 4 mug/ml of Purified Rabbit Anti-Active Caspase-3 antibody Cat. No. 570524/570525 , Lanes 1 and 3 4 mug and lanes 2 and 4 2 mug . Western blot: Active Caspase-3 was detected by Western blot analysis with an antibody that recognizes both the Pro-enzyme and Active Caspase-3 forms, reflecting the presence or absence of the caspase-3 pro-domain lanes 1-4 . The results show that the Purified Rabbit Anti-Active Caspase-3 antibody immunoprecipitated only the active form of Caspase-3.