procaspase 9 Search Results


33 murine procaspase 9  (Enzo Biochem)
90
Enzo Biochem 33 murine procaspase 9
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
33 Murine Procaspase 9, supplied by Enzo Biochem, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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33 murine procaspase 9 - by Bioz Stars, 2026-09
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procaspase-9  (SMAC Corp)
90
SMAC Corp procaspase-9
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
Procaspase 9, supplied by SMAC Corp, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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procaspase-9  (MBL Life science)
90
MBL Life science procaspase-9
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
Procaspase 9, supplied by MBL Life science, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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primary rabbit monoclonal antibody to procaspase-9  (GeneTex)
90
GeneTex primary rabbit monoclonal antibody to procaspase-9
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
Primary Rabbit Monoclonal Antibody To Procaspase 9, supplied by GeneTex, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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polyclonal anti-rabbit procaspase-9 antibody  (Becton Dickinson)
90
Becton Dickinson polyclonal anti-rabbit procaspase-9 antibody
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
Polyclonal Anti Rabbit Procaspase 9 Antibody, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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monoclonal human anti-procaspase 9  (Becton Dickinson)
90
Becton Dickinson monoclonal human anti-procaspase 9
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
Monoclonal Human Anti Procaspase 9, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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antibody against oncogene procaspase-9 (catalogue # 556585)  (Becton Dickinson)
90
Becton Dickinson antibody against oncogene procaspase-9 (catalogue # 556585)
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
Antibody Against Oncogene Procaspase 9 (Catalogue # 556585), supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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antibody rabbit anti-procaspase 9  (GeneTex)
90
GeneTex antibody rabbit anti-procaspase 9
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
Antibody Rabbit Anti Procaspase 9, supplied by GeneTex, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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antibody rabbit anti-procaspase 9 - by Bioz Stars, 2026-09
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procaspase-9  (Becton Dickinson)
90
Becton Dickinson procaspase-9
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
Procaspase 9, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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anti-procaspase-9 antibody (cat. no. adi-aam-139)  (Enzo Biochem)
90
Enzo Biochem anti-procaspase-9 antibody (cat. no. adi-aam-139)
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
Anti Procaspase 9 Antibody (Cat. No. Adi Aam 139), supplied by Enzo Biochem, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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procaspase 9/ caspase 9  (Becton Dickinson)
90
Becton Dickinson procaspase 9/ caspase 9
Cyt C-mediates <t>procaspase</t> <t>9</t> persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.
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Cyt C-mediates procaspase 9 persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.

Journal: ACS chemical biology

Article Title: Cytochrome c Reduction by H 2 S Potentiates Sulfide Signaling

doi: 10.1021/acschembio.8b00463

Figure Lengend Snippet: Cyt C-mediates procaspase 9 persulfdation and activity. (A) Murine recombinant procaspase 9 (1 μM) was incubated with 5 μM H2S for 30 min at 37 °C in the absence (lane 1) or presence of 0.5 (lane 2) and 2 μM (lane 3) Cyt C. Persulfidation was detected by the CN-Cy3-based tag switch method (top panel), while equal loading was confirmed by Coomassie blue staining (bottom panel). (B) H2S inhibits Cyt C-induced caspase 9 activation in cell lysates. Cyt C (500 nM) was added to freshly prepared HeLa cell lysates, and caspase 9 activity was monitored using a fluorescence caspase 9 kit. Simultaneous addition of Cyt C and H2S (10 μM) inhibited the Cyt C-induced increase in fluorescence. (C) Cyt C-induced caspase 9 activation in Jurkat cells is inhibited by H2S donors. Apoptosis was induced by staurosporine (ST, 2.5 μM), and caspase 9 activity was monitored by flow cytometry using the caspase 9 FITC staining kit 4 and 6 h after induction. Treatment of cells with GYY4137 (100 μM) or ammonium tetrathiomolybdate (ATTM, 200 μM) reduced fluorescence. (D) Inhibition of caspase 3 activation in cells treated with antimycin A and the slow-releasing H2S donor, GYY4137 (100 μM). HeLa cells were incubated with antimycin A (300 μM) overnight to induce the release of Cyt C from mitochondria. GYY4137, when used, was added 2 h prior to antimycin A. Caspase 3 activity was monitored in cell lysates as described for panel B. (E) Persulfidation of procaspase 9 in HeLa cells treated with 2.5 μM ST (1) that were either pretreated for 2 h (2) or treated for the last 2 h (3) with GYY4137 (100 μM): (left) total procaspase 9 levels and (right) persulfidation (PSSH) levels of procaspase 9. PSSH levels were quantified by measuring the difference in fluorescence between DTT-treated and untreated samples and normalizing this to the total procaspase 9 levels. The strategy used for sample processing to detect total versus persulfidated procaspase 9 is explained in the Supporting Information and Supporting Figure 5.

Article Snippet: 33 Murine procaspase 9 was from Enzo Life Science.

Techniques: Activity Assay, Recombinant, Incubation, Staining, Activation Assay, Fluorescence, Flow Cytometry, Inhibition