cleaved caspase 3 asp175 d3e9  (Cell Signaling Technology Inc)


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    Structured Review

    Cell Signaling Technology Inc cleaved caspase 3 asp175 d3e9
    Docetaxel overcomes EZH2 inhibitor resistance in PTEN-mutated tumors in mice. (A) C4-2 cells were infected with lentivirus expressing non-specific shRNA (shNS) or a pool of FOXO1-specific shRNAs and selected with puromycin for stable cell lines. Cells were treated with 5 μM of control ASO or EZH2-specific ASO and/or 2 nM of DTX and MTS assay was performed at different time points. (B, C) C4-2 cells were infected with lentivirus expressing shNS or a pool of FOXO1-specific shRNAs as indicated and selected with puromycin. The stable cells (5×10 6 /group) were injected subcutaneously into the right flank of NSG mice. When tumors reached the size of ~100 mm 3 , mice were treated with vehicle (V) (0.9% saline) plus 50 mg/kg of control ASO, vehicle plus 50 mg/kg of EZH2 ASO, 5 mg/kg of DTX plus 50 mg/kg of control ASO or 50 mg/kg of EZH2 ASO plus 5 mg/kg of DTX by i.p. injection twice a week (the 1 st and 4 th day of week). The tumor volume at each time point was documented (B) and tumors in each group were harvested and photographed at day 24 (C) . (D) Western blot analysis of protein expression in xenografts harvested from mice. ERK2 was used as a loading control. The cPARP stands for cleaved PARP. (E, F) H&E and IHC analysis of expression of Ki-67 and cleaved <t>Caspase-3</t> in xenograft sections from mice with indicated treatment. Representative images are shown in (E) and quantification of Ki67 and cleaved Caspase-3 positive cells from the tissue sections (n = 6) are shown in (F) . The number of positive cells from at least five fields were counted and analyzed. Data are shown as means ± SEM. The P value was performed by the unpaired two-tailed Student's t-test. * P <0.05; ** P <0.01; *** P <0.001; n.s., no significance.
    Cleaved Caspase 3 Asp175 D3e9, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "Overcoming EZH2 Inhibitor Resistance by Taxane in PTEN-Mutated Cancer"

    Article Title: Overcoming EZH2 Inhibitor Resistance by Taxane in PTEN-Mutated Cancer

    Journal: Theranostics

    doi: 10.7150/thno.34700

    Docetaxel overcomes EZH2 inhibitor resistance in PTEN-mutated tumors in mice. (A) C4-2 cells were infected with lentivirus expressing non-specific shRNA (shNS) or a pool of FOXO1-specific shRNAs and selected with puromycin for stable cell lines. Cells were treated with 5 μM of control ASO or EZH2-specific ASO and/or 2 nM of DTX and MTS assay was performed at different time points. (B, C) C4-2 cells were infected with lentivirus expressing shNS or a pool of FOXO1-specific shRNAs as indicated and selected with puromycin. The stable cells (5×10 6 /group) were injected subcutaneously into the right flank of NSG mice. When tumors reached the size of ~100 mm 3 , mice were treated with vehicle (V) (0.9% saline) plus 50 mg/kg of control ASO, vehicle plus 50 mg/kg of EZH2 ASO, 5 mg/kg of DTX plus 50 mg/kg of control ASO or 50 mg/kg of EZH2 ASO plus 5 mg/kg of DTX by i.p. injection twice a week (the 1 st and 4 th day of week). The tumor volume at each time point was documented (B) and tumors in each group were harvested and photographed at day 24 (C) . (D) Western blot analysis of protein expression in xenografts harvested from mice. ERK2 was used as a loading control. The cPARP stands for cleaved PARP. (E, F) H&E and IHC analysis of expression of Ki-67 and cleaved Caspase-3 in xenograft sections from mice with indicated treatment. Representative images are shown in (E) and quantification of Ki67 and cleaved Caspase-3 positive cells from the tissue sections (n = 6) are shown in (F) . The number of positive cells from at least five fields were counted and analyzed. Data are shown as means ± SEM. The P value was performed by the unpaired two-tailed Student's t-test. * P <0.05; ** P <0.01; *** P <0.001; n.s., no significance.
    Figure Legend Snippet: Docetaxel overcomes EZH2 inhibitor resistance in PTEN-mutated tumors in mice. (A) C4-2 cells were infected with lentivirus expressing non-specific shRNA (shNS) or a pool of FOXO1-specific shRNAs and selected with puromycin for stable cell lines. Cells were treated with 5 μM of control ASO or EZH2-specific ASO and/or 2 nM of DTX and MTS assay was performed at different time points. (B, C) C4-2 cells were infected with lentivirus expressing shNS or a pool of FOXO1-specific shRNAs as indicated and selected with puromycin. The stable cells (5×10 6 /group) were injected subcutaneously into the right flank of NSG mice. When tumors reached the size of ~100 mm 3 , mice were treated with vehicle (V) (0.9% saline) plus 50 mg/kg of control ASO, vehicle plus 50 mg/kg of EZH2 ASO, 5 mg/kg of DTX plus 50 mg/kg of control ASO or 50 mg/kg of EZH2 ASO plus 5 mg/kg of DTX by i.p. injection twice a week (the 1 st and 4 th day of week). The tumor volume at each time point was documented (B) and tumors in each group were harvested and photographed at day 24 (C) . (D) Western blot analysis of protein expression in xenografts harvested from mice. ERK2 was used as a loading control. The cPARP stands for cleaved PARP. (E, F) H&E and IHC analysis of expression of Ki-67 and cleaved Caspase-3 in xenograft sections from mice with indicated treatment. Representative images are shown in (E) and quantification of Ki67 and cleaved Caspase-3 positive cells from the tissue sections (n = 6) are shown in (F) . The number of positive cells from at least five fields were counted and analyzed. Data are shown as means ± SEM. The P value was performed by the unpaired two-tailed Student's t-test. * P <0.05; ** P <0.01; *** P <0.001; n.s., no significance.

    Techniques Used: Infection, Expressing, shRNA, Stable Transfection, MTS Assay, Injection, Western Blot, Two Tailed Test

    cleaved caspase 3 asp175 d3e9  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc cleaved caspase 3 asp175 d3e9
    Docetaxel overcomes EZH2 inhibitor resistance in PTEN-mutated tumors in mice. (A) C4-2 cells were infected with lentivirus expressing non-specific shRNA (shNS) or a pool of FOXO1-specific shRNAs and selected with puromycin for stable cell lines. Cells were treated with 5 μM of control ASO or EZH2-specific ASO and/or 2 nM of DTX and MTS assay was performed at different time points. (B, C) C4-2 cells were infected with lentivirus expressing shNS or a pool of FOXO1-specific shRNAs as indicated and selected with puromycin. The stable cells (5×10 6 /group) were injected subcutaneously into the right flank of NSG mice. When tumors reached the size of ~100 mm 3 , mice were treated with vehicle (V) (0.9% saline) plus 50 mg/kg of control ASO, vehicle plus 50 mg/kg of EZH2 ASO, 5 mg/kg of DTX plus 50 mg/kg of control ASO or 50 mg/kg of EZH2 ASO plus 5 mg/kg of DTX by i.p. injection twice a week (the 1 st and 4 th day of week). The tumor volume at each time point was documented (B) and tumors in each group were harvested and photographed at day 24 (C) . (D) Western blot analysis of protein expression in xenografts harvested from mice. ERK2 was used as a loading control. The cPARP stands for cleaved PARP. (E, F) H&E and IHC analysis of expression of Ki-67 and cleaved <t>Caspase-3</t> in xenograft sections from mice with indicated treatment. Representative images are shown in (E) and quantification of Ki67 and cleaved Caspase-3 positive cells from the tissue sections (n = 6) are shown in (F) . The number of positive cells from at least five fields were counted and analyzed. Data are shown as means ± SEM. The P value was performed by the unpaired two-tailed Student's t-test. * P <0.05; ** P <0.01; *** P <0.001; n.s., no significance.
    Cleaved Caspase 3 Asp175 D3e9, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cleaved caspase 3 asp175 d3e9/product/Cell Signaling Technology Inc
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    cleaved caspase 3 asp175 d3e9 - by Bioz Stars, 2023-01
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    Images

    1) Product Images from "Overcoming EZH2 Inhibitor Resistance by Taxane in PTEN-Mutated Cancer"

    Article Title: Overcoming EZH2 Inhibitor Resistance by Taxane in PTEN-Mutated Cancer

    Journal: Theranostics

    doi: 10.7150/thno.34700

    Docetaxel overcomes EZH2 inhibitor resistance in PTEN-mutated tumors in mice. (A) C4-2 cells were infected with lentivirus expressing non-specific shRNA (shNS) or a pool of FOXO1-specific shRNAs and selected with puromycin for stable cell lines. Cells were treated with 5 μM of control ASO or EZH2-specific ASO and/or 2 nM of DTX and MTS assay was performed at different time points. (B, C) C4-2 cells were infected with lentivirus expressing shNS or a pool of FOXO1-specific shRNAs as indicated and selected with puromycin. The stable cells (5×10 6 /group) were injected subcutaneously into the right flank of NSG mice. When tumors reached the size of ~100 mm 3 , mice were treated with vehicle (V) (0.9% saline) plus 50 mg/kg of control ASO, vehicle plus 50 mg/kg of EZH2 ASO, 5 mg/kg of DTX plus 50 mg/kg of control ASO or 50 mg/kg of EZH2 ASO plus 5 mg/kg of DTX by i.p. injection twice a week (the 1 st and 4 th day of week). The tumor volume at each time point was documented (B) and tumors in each group were harvested and photographed at day 24 (C) . (D) Western blot analysis of protein expression in xenografts harvested from mice. ERK2 was used as a loading control. The cPARP stands for cleaved PARP. (E, F) H&E and IHC analysis of expression of Ki-67 and cleaved Caspase-3 in xenograft sections from mice with indicated treatment. Representative images are shown in (E) and quantification of Ki67 and cleaved Caspase-3 positive cells from the tissue sections (n = 6) are shown in (F) . The number of positive cells from at least five fields were counted and analyzed. Data are shown as means ± SEM. The P value was performed by the unpaired two-tailed Student's t-test. * P <0.05; ** P <0.01; *** P <0.001; n.s., no significance.
    Figure Legend Snippet: Docetaxel overcomes EZH2 inhibitor resistance in PTEN-mutated tumors in mice. (A) C4-2 cells were infected with lentivirus expressing non-specific shRNA (shNS) or a pool of FOXO1-specific shRNAs and selected with puromycin for stable cell lines. Cells were treated with 5 μM of control ASO or EZH2-specific ASO and/or 2 nM of DTX and MTS assay was performed at different time points. (B, C) C4-2 cells were infected with lentivirus expressing shNS or a pool of FOXO1-specific shRNAs as indicated and selected with puromycin. The stable cells (5×10 6 /group) were injected subcutaneously into the right flank of NSG mice. When tumors reached the size of ~100 mm 3 , mice were treated with vehicle (V) (0.9% saline) plus 50 mg/kg of control ASO, vehicle plus 50 mg/kg of EZH2 ASO, 5 mg/kg of DTX plus 50 mg/kg of control ASO or 50 mg/kg of EZH2 ASO plus 5 mg/kg of DTX by i.p. injection twice a week (the 1 st and 4 th day of week). The tumor volume at each time point was documented (B) and tumors in each group were harvested and photographed at day 24 (C) . (D) Western blot analysis of protein expression in xenografts harvested from mice. ERK2 was used as a loading control. The cPARP stands for cleaved PARP. (E, F) H&E and IHC analysis of expression of Ki-67 and cleaved Caspase-3 in xenograft sections from mice with indicated treatment. Representative images are shown in (E) and quantification of Ki67 and cleaved Caspase-3 positive cells from the tissue sections (n = 6) are shown in (F) . The number of positive cells from at least five fields were counted and analyzed. Data are shown as means ± SEM. The P value was performed by the unpaired two-tailed Student's t-test. * P <0.05; ** P <0.01; *** P <0.001; n.s., no significance.

    Techniques Used: Infection, Expressing, shRNA, Stable Transfection, MTS Assay, Injection, Western Blot, Two Tailed Test

    cleaved caspase 3  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc cleaved caspase 3
    Effects of LINC00673 on pancreatic cancer cell apoptosis, viability, migration. (A)Validation of LINC00673 overexpression or knockdown in CanPan-1 cells as determined by qRT-PCR. (B) Transwell invasion assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (C) CCK-8 proliferation assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (D)Expression Cleaved-Caspase 3 in LINC00673 over-expression or knock-down in CanPan-1 cells as determined by western blot. (E)Flow cytometric analysis of apoptosis in LINC00673 over-expression or knock-down in CanPan-1 cells. *P<0.05, ** P<0.01
    Cleaved Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96/100 stars

    Images

    1) Product Images from "LNC00673 suppresses proliferation and metastasis of pancreatic cancer via target miR-504/ HNF1A"

    Article Title: LNC00673 suppresses proliferation and metastasis of pancreatic cancer via target miR-504/ HNF1A

    Journal: Journal of Cancer

    doi: 10.7150/jca.32855

    Effects of LINC00673 on pancreatic cancer cell apoptosis, viability, migration. (A)Validation of LINC00673 overexpression or knockdown in CanPan-1 cells as determined by qRT-PCR. (B) Transwell invasion assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (C) CCK-8 proliferation assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (D)Expression Cleaved-Caspase 3 in LINC00673 over-expression or knock-down in CanPan-1 cells as determined by western blot. (E)Flow cytometric analysis of apoptosis in LINC00673 over-expression or knock-down in CanPan-1 cells. *P<0.05, ** P<0.01
    Figure Legend Snippet: Effects of LINC00673 on pancreatic cancer cell apoptosis, viability, migration. (A)Validation of LINC00673 overexpression or knockdown in CanPan-1 cells as determined by qRT-PCR. (B) Transwell invasion assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (C) CCK-8 proliferation assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (D)Expression Cleaved-Caspase 3 in LINC00673 over-expression or knock-down in CanPan-1 cells as determined by western blot. (E)Flow cytometric analysis of apoptosis in LINC00673 over-expression or knock-down in CanPan-1 cells. *P<0.05, ** P<0.01

    Techniques Used: Migration, Over Expression, Quantitative RT-PCR, Transwell Invasion Assay, CCK-8 Assay, Proliferation Assay, Expressing, Western Blot

    HNF1A suppression inhibited cell progression in pancreatic cancer cells. CCK-8 proliferation assay of specified plasmid or RNA transfection transfected in CanPan-1 cells. (B) Transwell invasion assay in specified plasmid or RNA transfection in CanPan-1 cells. (C) Expression Cleaved-Caspase 3 in specified plasmid or RNA transfected in CanPan-1 cells as determined by western blot. (D)Flow cytometric analysis of apoptosis in specified plasmid or RNA transfected in CanPan-1cells. *P<0.05, ** P<0.01
    Figure Legend Snippet: HNF1A suppression inhibited cell progression in pancreatic cancer cells. CCK-8 proliferation assay of specified plasmid or RNA transfection transfected in CanPan-1 cells. (B) Transwell invasion assay in specified plasmid or RNA transfection in CanPan-1 cells. (C) Expression Cleaved-Caspase 3 in specified plasmid or RNA transfected in CanPan-1 cells as determined by western blot. (D)Flow cytometric analysis of apoptosis in specified plasmid or RNA transfected in CanPan-1cells. *P<0.05, ** P<0.01

    Techniques Used: CCK-8 Assay, Proliferation Assay, Plasmid Preparation, Transfection, Transwell Invasion Assay, Expressing, Western Blot

    cleaved caspase 3  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc cleaved caspase 3
    HQH promotes apoptosis of ALL cells by upregulating GRα and downregulating pERK. BAX and <t>cleaved-caspase-3</t> were upregulated and Bcl-2 was downregulated in both Jurkat ( A ) and Nalm-6 ( B ) cells. Additionally, HQH increased apoptosis by upregulating GRα and downregulating pERK levels in Jurkat ( C ) and Nalm-6 ( D ) cells. The loading control was β-actin. HQH – Huai Qi Huang; ALL – acute lymphoblastic leukemia; pERK – phospho-ERK; GR – glucocorticoid receptor; t-ERK – total ERK.
    Cleaved Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96/100 stars

    Images

    1) Product Images from "Huai Qi Huang Potentiates Dexamethasone-Mediated Lethality in Acute Lymphoblastic Leukemia Cells by Upregulating Glucocorticoid Receptor α"

    Article Title: Huai Qi Huang Potentiates Dexamethasone-Mediated Lethality in Acute Lymphoblastic Leukemia Cells by Upregulating Glucocorticoid Receptor α

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    doi: 10.12659/MSM.921649

    HQH promotes apoptosis of ALL cells by upregulating GRα and downregulating pERK. BAX and cleaved-caspase-3 were upregulated and Bcl-2 was downregulated in both Jurkat ( A ) and Nalm-6 ( B ) cells. Additionally, HQH increased apoptosis by upregulating GRα and downregulating pERK levels in Jurkat ( C ) and Nalm-6 ( D ) cells. The loading control was β-actin. HQH – Huai Qi Huang; ALL – acute lymphoblastic leukemia; pERK – phospho-ERK; GR – glucocorticoid receptor; t-ERK – total ERK.
    Figure Legend Snippet: HQH promotes apoptosis of ALL cells by upregulating GRα and downregulating pERK. BAX and cleaved-caspase-3 were upregulated and Bcl-2 was downregulated in both Jurkat ( A ) and Nalm-6 ( B ) cells. Additionally, HQH increased apoptosis by upregulating GRα and downregulating pERK levels in Jurkat ( C ) and Nalm-6 ( D ) cells. The loading control was β-actin. HQH – Huai Qi Huang; ALL – acute lymphoblastic leukemia; pERK – phospho-ERK; GR – glucocorticoid receptor; t-ERK – total ERK.

    Techniques Used:

    Co-treatment of cells with DEX and HQH enhances BAX and cleaved-caspase-3 expression and reduces Bcl-2 and pERK expression. Cells were cultured with 100 μmol/L DEX alone or DEX combined with 4.3 mg/mL HQH for Jurkat cells and 1.5 mg/mL HQH for Nalm-6 cells for 24 hours. We used western blotting to detect protein expression levels of BAX, cleaved-caspase-3, Bcl-2, t-ERK and pERK in Jurkat ( A, C ) and Nalm-6 ( B, D ) cells. 100 nmol/L DEX and the same dose of HQH as above were used to treat ALL cells for 24 hours, and western blotting was used to detect protein expression levels of BAX, cleaved-caspase-3, Bcl-2, t-ERK and pERK in Jurkat ( E ) and Nalm-6 ( F ) cells. The loading control was β-actin. DEX – dexamethasone; HQH – Huai Qi Huang; pERK – phospho-ERK; t-ERK – total ERK.
    Figure Legend Snippet: Co-treatment of cells with DEX and HQH enhances BAX and cleaved-caspase-3 expression and reduces Bcl-2 and pERK expression. Cells were cultured with 100 μmol/L DEX alone or DEX combined with 4.3 mg/mL HQH for Jurkat cells and 1.5 mg/mL HQH for Nalm-6 cells for 24 hours. We used western blotting to detect protein expression levels of BAX, cleaved-caspase-3, Bcl-2, t-ERK and pERK in Jurkat ( A, C ) and Nalm-6 ( B, D ) cells. 100 nmol/L DEX and the same dose of HQH as above were used to treat ALL cells for 24 hours, and western blotting was used to detect protein expression levels of BAX, cleaved-caspase-3, Bcl-2, t-ERK and pERK in Jurkat ( E ) and Nalm-6 ( F ) cells. The loading control was β-actin. DEX – dexamethasone; HQH – Huai Qi Huang; pERK – phospho-ERK; t-ERK – total ERK.

    Techniques Used: Expressing, Cell Culture, Western Blot

    HQH inhibits ERK phosphorylation through inhibition of MEK in Jurkat and Nalm-6 cells. ALL cells were cultured with 50 μmol/L of the MEK inhibitor PD98059 alone, or combination of 4.3 mg/mL HQH for Jurkat cells and 1.5 mg/mL HQH for Nalm-6 cells and PD98059 for 24 hours. We used western blotting to detect the proteins of cleaved-caspase 3, BAX, Bcl-2, t-ERK and pERK in Jurkat ( A, C ) and Nalm-6 ( B, D ) cells. β-actin was used as the internal control for western blotting. PD – PD98059; ALL – acute lymphoblastic leukemia; HQH – Huai Qi Huang; pERK – phospho-ERK; t-ERK – total ERK.
    Figure Legend Snippet: HQH inhibits ERK phosphorylation through inhibition of MEK in Jurkat and Nalm-6 cells. ALL cells were cultured with 50 μmol/L of the MEK inhibitor PD98059 alone, or combination of 4.3 mg/mL HQH for Jurkat cells and 1.5 mg/mL HQH for Nalm-6 cells and PD98059 for 24 hours. We used western blotting to detect the proteins of cleaved-caspase 3, BAX, Bcl-2, t-ERK and pERK in Jurkat ( A, C ) and Nalm-6 ( B, D ) cells. β-actin was used as the internal control for western blotting. PD – PD98059; ALL – acute lymphoblastic leukemia; HQH – Huai Qi Huang; pERK – phospho-ERK; t-ERK – total ERK.

    Techniques Used: Inhibition, Cell Culture, Western Blot

    cleaved caspase 3  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc cleaved caspase 3
    lncRNA-XLOC_006390 downregulation induced cancer cell apoptosis resulting in decline of cell viability. (a) Silencing of lncRNA-XLOC_006390 in HCT-15 and CaCo 2 cancer cell lines; (b) determination of viability of HCT-15 and CaCo 2 cancer cells repressing lncRNA-XLOC_006390 and respective control cells by MTT assay; (c) AO/EB staining of HCT-15 and CaCo 2 cancer cells repressing lncRNA-XLOC_006390 for the estimation of relative cell death with reference to control cancer cells; (d) estimation of apoptosis of HCT-15 and CaCo 2 cancer cells under lncRNA-XLOC_006390 silencing and respective control cell lines through flow cytometry; (e) western blot analysis of expression of bax, <t>Bcl-2,</t> <t>caspase-3</t> and cleaved caspase-3 proteins from lncRNA-XLOC_006390 repressing HCT-15, and CaCo 2 along with respective negative control cells. The values represent mean ± SD of three biological replicates ( ∗ P < 0.05).
    Cleaved Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Long Noncoding XLOC_006390 Regulates the Proliferation and Metastasis of Human Colorectal Cancer via miR-296/ONECUT2 Axis"

    Article Title: Long Noncoding XLOC_006390 Regulates the Proliferation and Metastasis of Human Colorectal Cancer via miR-296/ONECUT2 Axis

    Journal: Journal of Oncology

    doi: 10.1155/2022/4897201

    lncRNA-XLOC_006390 downregulation induced cancer cell apoptosis resulting in decline of cell viability. (a) Silencing of lncRNA-XLOC_006390 in HCT-15 and CaCo 2 cancer cell lines; (b) determination of viability of HCT-15 and CaCo 2 cancer cells repressing lncRNA-XLOC_006390 and respective control cells by MTT assay; (c) AO/EB staining of HCT-15 and CaCo 2 cancer cells repressing lncRNA-XLOC_006390 for the estimation of relative cell death with reference to control cancer cells; (d) estimation of apoptosis of HCT-15 and CaCo 2 cancer cells under lncRNA-XLOC_006390 silencing and respective control cell lines through flow cytometry; (e) western blot analysis of expression of bax, Bcl-2, caspase-3 and cleaved caspase-3 proteins from lncRNA-XLOC_006390 repressing HCT-15, and CaCo 2 along with respective negative control cells. The values represent mean ± SD of three biological replicates ( ∗ P < 0.05).
    Figure Legend Snippet: lncRNA-XLOC_006390 downregulation induced cancer cell apoptosis resulting in decline of cell viability. (a) Silencing of lncRNA-XLOC_006390 in HCT-15 and CaCo 2 cancer cell lines; (b) determination of viability of HCT-15 and CaCo 2 cancer cells repressing lncRNA-XLOC_006390 and respective control cells by MTT assay; (c) AO/EB staining of HCT-15 and CaCo 2 cancer cells repressing lncRNA-XLOC_006390 for the estimation of relative cell death with reference to control cancer cells; (d) estimation of apoptosis of HCT-15 and CaCo 2 cancer cells under lncRNA-XLOC_006390 silencing and respective control cell lines through flow cytometry; (e) western blot analysis of expression of bax, Bcl-2, caspase-3 and cleaved caspase-3 proteins from lncRNA-XLOC_006390 repressing HCT-15, and CaCo 2 along with respective negative control cells. The values represent mean ± SD of three biological replicates ( ∗ P < 0.05).

    Techniques Used: MTT Assay, Staining, Flow Cytometry, Western Blot, Expressing, Negative Control

    Silencing of lncRNA-XLOC_006390 reduced in vivo tumorigenesis. (a) Analysis of relative size of tumor xenografts under lncRNA-XLOC_006390 downregulation with reference to negative control mice tumors; (b) analysis of relative volume of tumor xenografts under lncRNA-XLOC_006390 downregulation with reference to negative control mice tumors; (c) analysis of average weight of tumor xenografts under lncRNA-XLOC_006390 downregulation with reference to negative control mice tumors; (d)immuno-histochemical staining of ki67 and cleaved caspase-3 proteins from tumor xenografts under lncRNA-XLOC_006390 downregulation with reference to negative control mice tumors.
    Figure Legend Snippet: Silencing of lncRNA-XLOC_006390 reduced in vivo tumorigenesis. (a) Analysis of relative size of tumor xenografts under lncRNA-XLOC_006390 downregulation with reference to negative control mice tumors; (b) analysis of relative volume of tumor xenografts under lncRNA-XLOC_006390 downregulation with reference to negative control mice tumors; (c) analysis of average weight of tumor xenografts under lncRNA-XLOC_006390 downregulation with reference to negative control mice tumors; (d)immuno-histochemical staining of ki67 and cleaved caspase-3 proteins from tumor xenografts under lncRNA-XLOC_006390 downregulation with reference to negative control mice tumors.

    Techniques Used: In Vivo, Negative Control, Staining

    cleaved caspase 3  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc cleaved caspase 3
    Allicin treatment alleviated AOPP-induced human NP cell apoptosis. (a) The human NP cells were pretreated with various concentrations of allicin (0, 5, 10, and 20 μ M) for 2 h, followed by stimulation with 400 μ g/ml AOPP for 24 h. And the rate of cell apoptosis was detected by flow cytometry with Annexin V-FITC/PI dual staining. The proportion of apoptotic cells in the first and fourth quadrant was measured for analysis. (b–h) The protein levels of Bax, Bcl-2, <t>c-caspase</t> <t>3,</t> c-caspase 9, mitochondrial Cyt-c, and cytoplasmic Cyt-c were determined using Western blotting analysis (b) and quantified in (c–h). (i) Representative images of immunofluorescence staining for cleaved <t>caspase-3</t> in each group, with the relative fluorescence intensity quantified. Original magnification: ×400. Data were represented as mean ± SD. ## p < 0.01 versus the control group; ∗ p < 0.05 and ∗∗ p < 0.01 versus the AOPP alone treatment group, n = 3.
    Cleaved Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Allicin Attenuated Advanced Oxidation Protein Product-Induced Oxidative Stress and Mitochondrial Apoptosis in Human Nucleus Pulposus Cells"

    Article Title: Allicin Attenuated Advanced Oxidation Protein Product-Induced Oxidative Stress and Mitochondrial Apoptosis in Human Nucleus Pulposus Cells

    Journal: Oxidative Medicine and Cellular Longevity

    doi: 10.1155/2020/6685043

    Allicin treatment alleviated AOPP-induced human NP cell apoptosis. (a) The human NP cells were pretreated with various concentrations of allicin (0, 5, 10, and 20 μ M) for 2 h, followed by stimulation with 400 μ g/ml AOPP for 24 h. And the rate of cell apoptosis was detected by flow cytometry with Annexin V-FITC/PI dual staining. The proportion of apoptotic cells in the first and fourth quadrant was measured for analysis. (b–h) The protein levels of Bax, Bcl-2, c-caspase 3, c-caspase 9, mitochondrial Cyt-c, and cytoplasmic Cyt-c were determined using Western blotting analysis (b) and quantified in (c–h). (i) Representative images of immunofluorescence staining for cleaved caspase-3 in each group, with the relative fluorescence intensity quantified. Original magnification: ×400. Data were represented as mean ± SD. ## p < 0.01 versus the control group; ∗ p < 0.05 and ∗∗ p < 0.01 versus the AOPP alone treatment group, n = 3.
    Figure Legend Snippet: Allicin treatment alleviated AOPP-induced human NP cell apoptosis. (a) The human NP cells were pretreated with various concentrations of allicin (0, 5, 10, and 20 μ M) for 2 h, followed by stimulation with 400 μ g/ml AOPP for 24 h. And the rate of cell apoptosis was detected by flow cytometry with Annexin V-FITC/PI dual staining. The proportion of apoptotic cells in the first and fourth quadrant was measured for analysis. (b–h) The protein levels of Bax, Bcl-2, c-caspase 3, c-caspase 9, mitochondrial Cyt-c, and cytoplasmic Cyt-c were determined using Western blotting analysis (b) and quantified in (c–h). (i) Representative images of immunofluorescence staining for cleaved caspase-3 in each group, with the relative fluorescence intensity quantified. Original magnification: ×400. Data were represented as mean ± SD. ## p < 0.01 versus the control group; ∗ p < 0.05 and ∗∗ p < 0.01 versus the AOPP alone treatment group, n = 3.

    Techniques Used: Flow Cytometry, Staining, Western Blot, Immunofluorescence, Fluorescence

    Allicin alleviated AOPP-induced oxidative stress and mitochondrial dysfunction via p38-MAPK pathway in human NP cells. (a) The human NP cells were pretreated with allicin (10 μ M) or p38-MAPK inhibitor SB202190 (10 μ M) for 2 h, then treated with AOPP (400 μ g/ml) for 24 h. The cell proliferation was determined using EdU staining combined with DAPI staining for the nuclei under fluorescence microscope, with the EdU positive cells quantitated. Original magnification: ×200. (b) Representative images of immunofluorescence staining for cleaved caspase-3 in each group, with the relative fluorescence intensity quantified. Original magnification: ×400. (c) The intracellular ROS levels of the NP cells for each group were detected by ROS-specific fluorescent probe DHE and measured by subsequent flow cytometry analysis. (d) The mitochondrial membrane potential of human NP cells in each group was examined by JC-1 staining and measured by subsequent flow cytometry analysis. Representative scatter plots of flow cytometry and the quantitative analysis of red fluorescence to green fluorescence ratio were shown. Data were represented as mean ± SD. # p < 0.05 and ## p < 0.01 versus the control group; ∗ p < 0.05 and ∗∗ p < 0.01 versus the AOPP alone treatment group, n = 3.
    Figure Legend Snippet: Allicin alleviated AOPP-induced oxidative stress and mitochondrial dysfunction via p38-MAPK pathway in human NP cells. (a) The human NP cells were pretreated with allicin (10 μ M) or p38-MAPK inhibitor SB202190 (10 μ M) for 2 h, then treated with AOPP (400 μ g/ml) for 24 h. The cell proliferation was determined using EdU staining combined with DAPI staining for the nuclei under fluorescence microscope, with the EdU positive cells quantitated. Original magnification: ×200. (b) Representative images of immunofluorescence staining for cleaved caspase-3 in each group, with the relative fluorescence intensity quantified. Original magnification: ×400. (c) The intracellular ROS levels of the NP cells for each group were detected by ROS-specific fluorescent probe DHE and measured by subsequent flow cytometry analysis. (d) The mitochondrial membrane potential of human NP cells in each group was examined by JC-1 staining and measured by subsequent flow cytometry analysis. Representative scatter plots of flow cytometry and the quantitative analysis of red fluorescence to green fluorescence ratio were shown. Data were represented as mean ± SD. # p < 0.05 and ## p < 0.01 versus the control group; ∗ p < 0.05 and ∗∗ p < 0.01 versus the AOPP alone treatment group, n = 3.

    Techniques Used: Staining, Fluorescence, Microscopy, Immunofluorescence, Flow Cytometry

    cleaved caspase 3  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc cleaved caspase 3
    Allicin treatment alleviated AOPP-induced human NP cell apoptosis. (a) The human NP cells were pretreated with various concentrations of allicin (0, 5, 10, and 20 μ M) for 2 h, followed by stimulation with 400 μ g/ml AOPP for 24 h. And the rate of cell apoptosis was detected by flow cytometry with Annexin V-FITC/PI dual staining. The proportion of apoptotic cells in the first and fourth quadrant was measured for analysis. (b–h) The protein levels of Bax, Bcl-2, <t>c-caspase</t> <t>3,</t> c-caspase 9, mitochondrial Cyt-c, and cytoplasmic Cyt-c were determined using Western blotting analysis (b) and quantified in (c–h). (i) Representative images of immunofluorescence staining for cleaved <t>caspase-3</t> in each group, with the relative fluorescence intensity quantified. Original magnification: ×400. Data were represented as mean ± SD. ## p < 0.01 versus the control group; ∗ p < 0.05 and ∗∗ p < 0.01 versus the AOPP alone treatment group, n = 3.
    Cleaved Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "Allicin Attenuated Advanced Oxidation Protein Product-Induced Oxidative Stress and Mitochondrial Apoptosis in Human Nucleus Pulposus Cells"

    Article Title: Allicin Attenuated Advanced Oxidation Protein Product-Induced Oxidative Stress and Mitochondrial Apoptosis in Human Nucleus Pulposus Cells

    Journal: Oxidative Medicine and Cellular Longevity

    doi: 10.1155/2020/6685043

    Allicin treatment alleviated AOPP-induced human NP cell apoptosis. (a) The human NP cells were pretreated with various concentrations of allicin (0, 5, 10, and 20 μ M) for 2 h, followed by stimulation with 400 μ g/ml AOPP for 24 h. And the rate of cell apoptosis was detected by flow cytometry with Annexin V-FITC/PI dual staining. The proportion of apoptotic cells in the first and fourth quadrant was measured for analysis. (b–h) The protein levels of Bax, Bcl-2, c-caspase 3, c-caspase 9, mitochondrial Cyt-c, and cytoplasmic Cyt-c were determined using Western blotting analysis (b) and quantified in (c–h). (i) Representative images of immunofluorescence staining for cleaved caspase-3 in each group, with the relative fluorescence intensity quantified. Original magnification: ×400. Data were represented as mean ± SD. ## p < 0.01 versus the control group; ∗ p < 0.05 and ∗∗ p < 0.01 versus the AOPP alone treatment group, n = 3.
    Figure Legend Snippet: Allicin treatment alleviated AOPP-induced human NP cell apoptosis. (a) The human NP cells were pretreated with various concentrations of allicin (0, 5, 10, and 20 μ M) for 2 h, followed by stimulation with 400 μ g/ml AOPP for 24 h. And the rate of cell apoptosis was detected by flow cytometry with Annexin V-FITC/PI dual staining. The proportion of apoptotic cells in the first and fourth quadrant was measured for analysis. (b–h) The protein levels of Bax, Bcl-2, c-caspase 3, c-caspase 9, mitochondrial Cyt-c, and cytoplasmic Cyt-c were determined using Western blotting analysis (b) and quantified in (c–h). (i) Representative images of immunofluorescence staining for cleaved caspase-3 in each group, with the relative fluorescence intensity quantified. Original magnification: ×400. Data were represented as mean ± SD. ## p < 0.01 versus the control group; ∗ p < 0.05 and ∗∗ p < 0.01 versus the AOPP alone treatment group, n = 3.

    Techniques Used: Flow Cytometry, Staining, Western Blot, Immunofluorescence, Fluorescence

    Allicin alleviated AOPP-induced oxidative stress and mitochondrial dysfunction via p38-MAPK pathway in human NP cells. (a) The human NP cells were pretreated with allicin (10 μ M) or p38-MAPK inhibitor SB202190 (10 μ M) for 2 h, then treated with AOPP (400 μ g/ml) for 24 h. The cell proliferation was determined using EdU staining combined with DAPI staining for the nuclei under fluorescence microscope, with the EdU positive cells quantitated. Original magnification: ×200. (b) Representative images of immunofluorescence staining for cleaved caspase-3 in each group, with the relative fluorescence intensity quantified. Original magnification: ×400. (c) The intracellular ROS levels of the NP cells for each group were detected by ROS-specific fluorescent probe DHE and measured by subsequent flow cytometry analysis. (d) The mitochondrial membrane potential of human NP cells in each group was examined by JC-1 staining and measured by subsequent flow cytometry analysis. Representative scatter plots of flow cytometry and the quantitative analysis of red fluorescence to green fluorescence ratio were shown. Data were represented as mean ± SD. # p < 0.05 and ## p < 0.01 versus the control group; ∗ p < 0.05 and ∗∗ p < 0.01 versus the AOPP alone treatment group, n = 3.
    Figure Legend Snippet: Allicin alleviated AOPP-induced oxidative stress and mitochondrial dysfunction via p38-MAPK pathway in human NP cells. (a) The human NP cells were pretreated with allicin (10 μ M) or p38-MAPK inhibitor SB202190 (10 μ M) for 2 h, then treated with AOPP (400 μ g/ml) for 24 h. The cell proliferation was determined using EdU staining combined with DAPI staining for the nuclei under fluorescence microscope, with the EdU positive cells quantitated. Original magnification: ×200. (b) Representative images of immunofluorescence staining for cleaved caspase-3 in each group, with the relative fluorescence intensity quantified. Original magnification: ×400. (c) The intracellular ROS levels of the NP cells for each group were detected by ROS-specific fluorescent probe DHE and measured by subsequent flow cytometry analysis. (d) The mitochondrial membrane potential of human NP cells in each group was examined by JC-1 staining and measured by subsequent flow cytometry analysis. Representative scatter plots of flow cytometry and the quantitative analysis of red fluorescence to green fluorescence ratio were shown. Data were represented as mean ± SD. # p < 0.05 and ## p < 0.01 versus the control group; ∗ p < 0.05 and ∗∗ p < 0.01 versus the AOPP alone treatment group, n = 3.

    Techniques Used: Staining, Fluorescence, Microscopy, Immunofluorescence, Flow Cytometry

    cleaved caspase 3 antibody  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc cleaved caspase 3 antibody
    High-fat diet (HFD; ■) consumption does not modify enteroendocrine cell proliferation or apoptosis in mice. Representative double immunofluorescent staining ( n 7–8) of glucagon-like peptide-1 (GLP-1) (green) and bromodeoxyuridine (BrdU) (red) with DAPI (4′,6-diamidino-2-phenylindole) staining (blue) in the jejunum (a) of mice fed a control diet (CD; □) or the HFD for 2 weeks. The lower panels of (a) show GLP-1 and BrdU double-positive cells whereas the upper panels of (a) show GLP-1-positive BrdU-negative cells (highlighted with dotted circles). The scale bar represents 10 μm. Quantification as percentage of proliferating GLP-1 cells over total GLP-1 cells in the jejunum (b) and colon (c). Representative immunostaining ( n 6–8) of cleaved <t>caspase-3</t> positive cells in the jejunum (d) of mice fed the CD or HFD for 2 weeks. The scale bar represents 20 μm. Quantification of stained cells in the jejunum (e) and colon (f). Values are means, with standard errors represented by vertical bars. Statistical analysis was performed using a non-parametric Mann–Whitney test; there were no significant differences between the groups.
    Cleaved Caspase 3 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Lipid-rich diet enhances L-cell density in obese subjects and in mice through improved L-cell differentiation"

    Article Title: Lipid-rich diet enhances L-cell density in obese subjects and in mice through improved L-cell differentiation

    Journal: Journal of Nutritional Science

    doi: 10.1017/jns.2015.11

    High-fat diet (HFD; ■) consumption does not modify enteroendocrine cell proliferation or apoptosis in mice. Representative double immunofluorescent staining ( n 7–8) of glucagon-like peptide-1 (GLP-1) (green) and bromodeoxyuridine (BrdU) (red) with DAPI (4′,6-diamidino-2-phenylindole) staining (blue) in the jejunum (a) of mice fed a control diet (CD; □) or the HFD for 2 weeks. The lower panels of (a) show GLP-1 and BrdU double-positive cells whereas the upper panels of (a) show GLP-1-positive BrdU-negative cells (highlighted with dotted circles). The scale bar represents 10 μm. Quantification as percentage of proliferating GLP-1 cells over total GLP-1 cells in the jejunum (b) and colon (c). Representative immunostaining ( n 6–8) of cleaved caspase-3 positive cells in the jejunum (d) of mice fed the CD or HFD for 2 weeks. The scale bar represents 20 μm. Quantification of stained cells in the jejunum (e) and colon (f). Values are means, with standard errors represented by vertical bars. Statistical analysis was performed using a non-parametric Mann–Whitney test; there were no significant differences between the groups.
    Figure Legend Snippet: High-fat diet (HFD; ■) consumption does not modify enteroendocrine cell proliferation or apoptosis in mice. Representative double immunofluorescent staining ( n 7–8) of glucagon-like peptide-1 (GLP-1) (green) and bromodeoxyuridine (BrdU) (red) with DAPI (4′,6-diamidino-2-phenylindole) staining (blue) in the jejunum (a) of mice fed a control diet (CD; □) or the HFD for 2 weeks. The lower panels of (a) show GLP-1 and BrdU double-positive cells whereas the upper panels of (a) show GLP-1-positive BrdU-negative cells (highlighted with dotted circles). The scale bar represents 10 μm. Quantification as percentage of proliferating GLP-1 cells over total GLP-1 cells in the jejunum (b) and colon (c). Representative immunostaining ( n 6–8) of cleaved caspase-3 positive cells in the jejunum (d) of mice fed the CD or HFD for 2 weeks. The scale bar represents 20 μm. Quantification of stained cells in the jejunum (e) and colon (f). Values are means, with standard errors represented by vertical bars. Statistical analysis was performed using a non-parametric Mann–Whitney test; there were no significant differences between the groups.

    Techniques Used: Staining, Immunostaining, MANN-WHITNEY

    caspase 3 antibody  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc caspase 3 antibody
    Caspase 3 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    caspase 3  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc caspase 3
    A) Percentage of positive cells for OCT3/4 and SOX2 in cells with reduced OCT3/4 and SOX2 levels compared to cells transfected with control siRNA in the NCCIT cells in three different time-points post transfection based on immunohistochemistry; The controls are set to 100 in all cases. B) Relative expression pattern of 32 genes representing targets for pluripotency and differentiation (ectoderm, mesoderm and endoderm). The expression levels are normalized based on the housekeeping gene HPRT ; C) Percentage of living NCCIT cells with reduced level of OCT3/4 and SOX2 compared to cells transfected with control siRNA at each time point based on Trypan blue measurement; D) Percentage of positive NCCIT cells for Ki67 in untreated cells, cells transfected with control siRNA and cells with reduced levels of OCT3/4 and SOX2 at 48+12 h post transfection; E) Percentage of positive cells for <t>Caspase</t> <t>3</t> in untreated NCCIT cells, Cells transfected with control siRNA show a reduced level of OCT3/4 and SOX2 at all three time points after the transfection; F) FACS analysis with Propidium Iodide staining in cells transfected with control siRNA, cells transfected with SOX2 and OCT3/4 siRNA at 60 h post transfection. NCCIT cells transfected with control siRNA show the presence of 78% living cells, while cells transfected with SOX2 siRNA show 51% living cells, NCCIT cells with OCT3/4kd show 62% live cells. G) Annexin V assay results in cells with siRNA control and cells with reduced SOX2 and OCT3/4 at 60 h after transfection. In the control, percentages of living-annexin negative cells are 89.6%, while dead-annexin positive cells are 12.2%. In SOX2kd cells, the amount of living-annexin negative cells is 57.5% while the amount of dead-annexin positive cells is 17.2%. In cells transfected with OCT3/4 siRNA, living-annexin negative cells are 68.3% while dead-annexin positive cells are 12.8%. The differences between the percentages of live and dead cells in all experiments are due to using independent and various methods in order to prove apoptosis and different sensitivity of the materials and methods used. As it is demonstrated, OCT3/4 knock-down also induces apoptosis, however, this effect is not as dramatic as SOX2 knock-down. The FACS analysis were performed in triplicate.
    Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Role of SOX2 in the Etiology of Embryonal Carcinoma, Based on Analysis of the NCCIT and NT2 Cell Lines"

    Article Title: Role of SOX2 in the Etiology of Embryonal Carcinoma, Based on Analysis of the NCCIT and NT2 Cell Lines

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0083585

    A) Percentage of positive cells for OCT3/4 and SOX2 in cells with reduced OCT3/4 and SOX2 levels compared to cells transfected with control siRNA in the NCCIT cells in three different time-points post transfection based on immunohistochemistry; The controls are set to 100 in all cases. B) Relative expression pattern of 32 genes representing targets for pluripotency and differentiation (ectoderm, mesoderm and endoderm). The expression levels are normalized based on the housekeeping gene HPRT ; C) Percentage of living NCCIT cells with reduced level of OCT3/4 and SOX2 compared to cells transfected with control siRNA at each time point based on Trypan blue measurement; D) Percentage of positive NCCIT cells for Ki67 in untreated cells, cells transfected with control siRNA and cells with reduced levels of OCT3/4 and SOX2 at 48+12 h post transfection; E) Percentage of positive cells for Caspase 3 in untreated NCCIT cells, Cells transfected with control siRNA show a reduced level of OCT3/4 and SOX2 at all three time points after the transfection; F) FACS analysis with Propidium Iodide staining in cells transfected with control siRNA, cells transfected with SOX2 and OCT3/4 siRNA at 60 h post transfection. NCCIT cells transfected with control siRNA show the presence of 78% living cells, while cells transfected with SOX2 siRNA show 51% living cells, NCCIT cells with OCT3/4kd show 62% live cells. G) Annexin V assay results in cells with siRNA control and cells with reduced SOX2 and OCT3/4 at 60 h after transfection. In the control, percentages of living-annexin negative cells are 89.6%, while dead-annexin positive cells are 12.2%. In SOX2kd cells, the amount of living-annexin negative cells is 57.5% while the amount of dead-annexin positive cells is 17.2%. In cells transfected with OCT3/4 siRNA, living-annexin negative cells are 68.3% while dead-annexin positive cells are 12.8%. The differences between the percentages of live and dead cells in all experiments are due to using independent and various methods in order to prove apoptosis and different sensitivity of the materials and methods used. As it is demonstrated, OCT3/4 knock-down also induces apoptosis, however, this effect is not as dramatic as SOX2 knock-down. The FACS analysis were performed in triplicate.
    Figure Legend Snippet: A) Percentage of positive cells for OCT3/4 and SOX2 in cells with reduced OCT3/4 and SOX2 levels compared to cells transfected with control siRNA in the NCCIT cells in three different time-points post transfection based on immunohistochemistry; The controls are set to 100 in all cases. B) Relative expression pattern of 32 genes representing targets for pluripotency and differentiation (ectoderm, mesoderm and endoderm). The expression levels are normalized based on the housekeeping gene HPRT ; C) Percentage of living NCCIT cells with reduced level of OCT3/4 and SOX2 compared to cells transfected with control siRNA at each time point based on Trypan blue measurement; D) Percentage of positive NCCIT cells for Ki67 in untreated cells, cells transfected with control siRNA and cells with reduced levels of OCT3/4 and SOX2 at 48+12 h post transfection; E) Percentage of positive cells for Caspase 3 in untreated NCCIT cells, Cells transfected with control siRNA show a reduced level of OCT3/4 and SOX2 at all three time points after the transfection; F) FACS analysis with Propidium Iodide staining in cells transfected with control siRNA, cells transfected with SOX2 and OCT3/4 siRNA at 60 h post transfection. NCCIT cells transfected with control siRNA show the presence of 78% living cells, while cells transfected with SOX2 siRNA show 51% living cells, NCCIT cells with OCT3/4kd show 62% live cells. G) Annexin V assay results in cells with siRNA control and cells with reduced SOX2 and OCT3/4 at 60 h after transfection. In the control, percentages of living-annexin negative cells are 89.6%, while dead-annexin positive cells are 12.2%. In SOX2kd cells, the amount of living-annexin negative cells is 57.5% while the amount of dead-annexin positive cells is 17.2%. In cells transfected with OCT3/4 siRNA, living-annexin negative cells are 68.3% while dead-annexin positive cells are 12.8%. The differences between the percentages of live and dead cells in all experiments are due to using independent and various methods in order to prove apoptosis and different sensitivity of the materials and methods used. As it is demonstrated, OCT3/4 knock-down also induces apoptosis, however, this effect is not as dramatic as SOX2 knock-down. The FACS analysis were performed in triplicate.

    Techniques Used: Transfection, Immunohistochemistry, Expressing, Staining, Annexin V Assay

    anti cleaved caspase 3  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc anti cleaved caspase 3
    Anti Cleaved Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc cleaved caspase 3 asp175 d3e9
    Docetaxel overcomes EZH2 inhibitor resistance in PTEN-mutated tumors in mice. (A) C4-2 cells were infected with lentivirus expressing non-specific shRNA (shNS) or a pool of FOXO1-specific shRNAs and selected with puromycin for stable cell lines. Cells were treated with 5 μM of control ASO or EZH2-specific ASO and/or 2 nM of DTX and MTS assay was performed at different time points. (B, C) C4-2 cells were infected with lentivirus expressing shNS or a pool of FOXO1-specific shRNAs as indicated and selected with puromycin. The stable cells (5×10 6 /group) were injected subcutaneously into the right flank of NSG mice. When tumors reached the size of ~100 mm 3 , mice were treated with vehicle (V) (0.9% saline) plus 50 mg/kg of control ASO, vehicle plus 50 mg/kg of EZH2 ASO, 5 mg/kg of DTX plus 50 mg/kg of control ASO or 50 mg/kg of EZH2 ASO plus 5 mg/kg of DTX by i.p. injection twice a week (the 1 st and 4 th day of week). The tumor volume at each time point was documented (B) and tumors in each group were harvested and photographed at day 24 (C) . (D) Western blot analysis of protein expression in xenografts harvested from mice. ERK2 was used as a loading control. The cPARP stands for cleaved PARP. (E, F) H&E and IHC analysis of expression of Ki-67 and cleaved <t>Caspase-3</t> in xenograft sections from mice with indicated treatment. Representative images are shown in (E) and quantification of Ki67 and cleaved Caspase-3 positive cells from the tissue sections (n = 6) are shown in (F) . The number of positive cells from at least five fields were counted and analyzed. Data are shown as means ± SEM. The P value was performed by the unpaired two-tailed Student's t-test. * P <0.05; ** P <0.01; *** P <0.001; n.s., no significance.
    Cleaved Caspase 3 Asp175 D3e9, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc cleaved caspase 3
    Effects of LINC00673 on pancreatic cancer cell apoptosis, viability, migration. (A)Validation of LINC00673 overexpression or knockdown in CanPan-1 cells as determined by qRT-PCR. (B) Transwell invasion assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (C) CCK-8 proliferation assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (D)Expression Cleaved-Caspase 3 in LINC00673 over-expression or knock-down in CanPan-1 cells as determined by western blot. (E)Flow cytometric analysis of apoptosis in LINC00673 over-expression or knock-down in CanPan-1 cells. *P<0.05, ** P<0.01
    Cleaved Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc cleaved caspase 3 antibody
    High-fat diet (HFD; ■) consumption does not modify enteroendocrine cell proliferation or apoptosis in mice. Representative double immunofluorescent staining ( n 7–8) of glucagon-like peptide-1 (GLP-1) (green) and bromodeoxyuridine (BrdU) (red) with DAPI (4′,6-diamidino-2-phenylindole) staining (blue) in the jejunum (a) of mice fed a control diet (CD; □) or the HFD for 2 weeks. The lower panels of (a) show GLP-1 and BrdU double-positive cells whereas the upper panels of (a) show GLP-1-positive BrdU-negative cells (highlighted with dotted circles). The scale bar represents 10 μm. Quantification as percentage of proliferating GLP-1 cells over total GLP-1 cells in the jejunum (b) and colon (c). Representative immunostaining ( n 6–8) of cleaved <t>caspase-3</t> positive cells in the jejunum (d) of mice fed the CD or HFD for 2 weeks. The scale bar represents 20 μm. Quantification of stained cells in the jejunum (e) and colon (f). Values are means, with standard errors represented by vertical bars. Statistical analysis was performed using a non-parametric Mann–Whitney test; there were no significant differences between the groups.
    Cleaved Caspase 3 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    High-fat diet (HFD; ■) consumption does not modify enteroendocrine cell proliferation or apoptosis in mice. Representative double immunofluorescent staining ( n 7–8) of glucagon-like peptide-1 (GLP-1) (green) and bromodeoxyuridine (BrdU) (red) with DAPI (4′,6-diamidino-2-phenylindole) staining (blue) in the jejunum (a) of mice fed a control diet (CD; □) or the HFD for 2 weeks. The lower panels of (a) show GLP-1 and BrdU double-positive cells whereas the upper panels of (a) show GLP-1-positive BrdU-negative cells (highlighted with dotted circles). The scale bar represents 10 μm. Quantification as percentage of proliferating GLP-1 cells over total GLP-1 cells in the jejunum (b) and colon (c). Representative immunostaining ( n 6–8) of cleaved <t>caspase-3</t> positive cells in the jejunum (d) of mice fed the CD or HFD for 2 weeks. The scale bar represents 20 μm. Quantification of stained cells in the jejunum (e) and colon (f). Values are means, with standard errors represented by vertical bars. Statistical analysis was performed using a non-parametric Mann–Whitney test; there were no significant differences between the groups.
    Caspase 3 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    A) Percentage of positive cells for OCT3/4 and SOX2 in cells with reduced OCT3/4 and SOX2 levels compared to cells transfected with control siRNA in the NCCIT cells in three different time-points post transfection based on immunohistochemistry; The controls are set to 100 in all cases. B) Relative expression pattern of 32 genes representing targets for pluripotency and differentiation (ectoderm, mesoderm and endoderm). The expression levels are normalized based on the housekeeping gene HPRT ; C) Percentage of living NCCIT cells with reduced level of OCT3/4 and SOX2 compared to cells transfected with control siRNA at each time point based on Trypan blue measurement; D) Percentage of positive NCCIT cells for Ki67 in untreated cells, cells transfected with control siRNA and cells with reduced levels of OCT3/4 and SOX2 at 48+12 h post transfection; E) Percentage of positive cells for <t>Caspase</t> <t>3</t> in untreated NCCIT cells, Cells transfected with control siRNA show a reduced level of OCT3/4 and SOX2 at all three time points after the transfection; F) FACS analysis with Propidium Iodide staining in cells transfected with control siRNA, cells transfected with SOX2 and OCT3/4 siRNA at 60 h post transfection. NCCIT cells transfected with control siRNA show the presence of 78% living cells, while cells transfected with SOX2 siRNA show 51% living cells, NCCIT cells with OCT3/4kd show 62% live cells. G) Annexin V assay results in cells with siRNA control and cells with reduced SOX2 and OCT3/4 at 60 h after transfection. In the control, percentages of living-annexin negative cells are 89.6%, while dead-annexin positive cells are 12.2%. In SOX2kd cells, the amount of living-annexin negative cells is 57.5% while the amount of dead-annexin positive cells is 17.2%. In cells transfected with OCT3/4 siRNA, living-annexin negative cells are 68.3% while dead-annexin positive cells are 12.8%. The differences between the percentages of live and dead cells in all experiments are due to using independent and various methods in order to prove apoptosis and different sensitivity of the materials and methods used. As it is demonstrated, OCT3/4 knock-down also induces apoptosis, however, this effect is not as dramatic as SOX2 knock-down. The FACS analysis were performed in triplicate.
    Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    A) Percentage of positive cells for OCT3/4 and SOX2 in cells with reduced OCT3/4 and SOX2 levels compared to cells transfected with control siRNA in the NCCIT cells in three different time-points post transfection based on immunohistochemistry; The controls are set to 100 in all cases. B) Relative expression pattern of 32 genes representing targets for pluripotency and differentiation (ectoderm, mesoderm and endoderm). The expression levels are normalized based on the housekeeping gene HPRT ; C) Percentage of living NCCIT cells with reduced level of OCT3/4 and SOX2 compared to cells transfected with control siRNA at each time point based on Trypan blue measurement; D) Percentage of positive NCCIT cells for Ki67 in untreated cells, cells transfected with control siRNA and cells with reduced levels of OCT3/4 and SOX2 at 48+12 h post transfection; E) Percentage of positive cells for <t>Caspase</t> <t>3</t> in untreated NCCIT cells, Cells transfected with control siRNA show a reduced level of OCT3/4 and SOX2 at all three time points after the transfection; F) FACS analysis with Propidium Iodide staining in cells transfected with control siRNA, cells transfected with SOX2 and OCT3/4 siRNA at 60 h post transfection. NCCIT cells transfected with control siRNA show the presence of 78% living cells, while cells transfected with SOX2 siRNA show 51% living cells, NCCIT cells with OCT3/4kd show 62% live cells. G) Annexin V assay results in cells with siRNA control and cells with reduced SOX2 and OCT3/4 at 60 h after transfection. In the control, percentages of living-annexin negative cells are 89.6%, while dead-annexin positive cells are 12.2%. In SOX2kd cells, the amount of living-annexin negative cells is 57.5% while the amount of dead-annexin positive cells is 17.2%. In cells transfected with OCT3/4 siRNA, living-annexin negative cells are 68.3% while dead-annexin positive cells are 12.8%. The differences between the percentages of live and dead cells in all experiments are due to using independent and various methods in order to prove apoptosis and different sensitivity of the materials and methods used. As it is demonstrated, OCT3/4 knock-down also induces apoptosis, however, this effect is not as dramatic as SOX2 knock-down. The FACS analysis were performed in triplicate.
    Anti Cleaved Caspase 3, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Docetaxel overcomes EZH2 inhibitor resistance in PTEN-mutated tumors in mice. (A) C4-2 cells were infected with lentivirus expressing non-specific shRNA (shNS) or a pool of FOXO1-specific shRNAs and selected with puromycin for stable cell lines. Cells were treated with 5 μM of control ASO or EZH2-specific ASO and/or 2 nM of DTX and MTS assay was performed at different time points. (B, C) C4-2 cells were infected with lentivirus expressing shNS or a pool of FOXO1-specific shRNAs as indicated and selected with puromycin. The stable cells (5×10 6 /group) were injected subcutaneously into the right flank of NSG mice. When tumors reached the size of ~100 mm 3 , mice were treated with vehicle (V) (0.9% saline) plus 50 mg/kg of control ASO, vehicle plus 50 mg/kg of EZH2 ASO, 5 mg/kg of DTX plus 50 mg/kg of control ASO or 50 mg/kg of EZH2 ASO plus 5 mg/kg of DTX by i.p. injection twice a week (the 1 st and 4 th day of week). The tumor volume at each time point was documented (B) and tumors in each group were harvested and photographed at day 24 (C) . (D) Western blot analysis of protein expression in xenografts harvested from mice. ERK2 was used as a loading control. The cPARP stands for cleaved PARP. (E, F) H&E and IHC analysis of expression of Ki-67 and cleaved Caspase-3 in xenograft sections from mice with indicated treatment. Representative images are shown in (E) and quantification of Ki67 and cleaved Caspase-3 positive cells from the tissue sections (n = 6) are shown in (F) . The number of positive cells from at least five fields were counted and analyzed. Data are shown as means ± SEM. The P value was performed by the unpaired two-tailed Student's t-test. * P <0.05; ** P <0.01; *** P <0.001; n.s., no significance.

    Journal: Theranostics

    Article Title: Overcoming EZH2 Inhibitor Resistance by Taxane in PTEN-Mutated Cancer

    doi: 10.7150/thno.34700

    Figure Lengend Snippet: Docetaxel overcomes EZH2 inhibitor resistance in PTEN-mutated tumors in mice. (A) C4-2 cells were infected with lentivirus expressing non-specific shRNA (shNS) or a pool of FOXO1-specific shRNAs and selected with puromycin for stable cell lines. Cells were treated with 5 μM of control ASO or EZH2-specific ASO and/or 2 nM of DTX and MTS assay was performed at different time points. (B, C) C4-2 cells were infected with lentivirus expressing shNS or a pool of FOXO1-specific shRNAs as indicated and selected with puromycin. The stable cells (5×10 6 /group) were injected subcutaneously into the right flank of NSG mice. When tumors reached the size of ~100 mm 3 , mice were treated with vehicle (V) (0.9% saline) plus 50 mg/kg of control ASO, vehicle plus 50 mg/kg of EZH2 ASO, 5 mg/kg of DTX plus 50 mg/kg of control ASO or 50 mg/kg of EZH2 ASO plus 5 mg/kg of DTX by i.p. injection twice a week (the 1 st and 4 th day of week). The tumor volume at each time point was documented (B) and tumors in each group were harvested and photographed at day 24 (C) . (D) Western blot analysis of protein expression in xenografts harvested from mice. ERK2 was used as a loading control. The cPARP stands for cleaved PARP. (E, F) H&E and IHC analysis of expression of Ki-67 and cleaved Caspase-3 in xenograft sections from mice with indicated treatment. Representative images are shown in (E) and quantification of Ki67 and cleaved Caspase-3 positive cells from the tissue sections (n = 6) are shown in (F) . The number of positive cells from at least five fields were counted and analyzed. Data are shown as means ± SEM. The P value was performed by the unpaired two-tailed Student's t-test. * P <0.05; ** P <0.01; *** P <0.001; n.s., no significance.

    Article Snippet: The antibodies used are as follows: EZH2 (5246S), FOXO1 (2880S), Myc tag (2278S), cleaved caspase-3 (Asp175) (D3E9) (9579S) and cleaved PARP (ASP214) (D64E10) (5625S) were purchased from Cell Signaling Technology; Ki67 (ab15580), H3K27me3 (ab6002) and H3K27ac (ab177178) from Abcam; EZH1 (sc-515817), ERK 2 (D-2) (sc-1647) and total H3 (sc-10809) from Santa Cruz; EED (05-1327) and SUZ12 (3C1.2) (05-1320) from Millipore; β-Tubulin (DHSB E7, AB-528499) from Developmental Studies Hybridoma Bank (DSHB); E-cadherin (BD 610181) from BD Biosciences; BIM (AB17003) from Chemicon International.

    Techniques: Infection, Expressing, shRNA, Stable Transfection, MTS Assay, Injection, Western Blot, Two Tailed Test

    Effects of LINC00673 on pancreatic cancer cell apoptosis, viability, migration. (A)Validation of LINC00673 overexpression or knockdown in CanPan-1 cells as determined by qRT-PCR. (B) Transwell invasion assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (C) CCK-8 proliferation assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (D)Expression Cleaved-Caspase 3 in LINC00673 over-expression or knock-down in CanPan-1 cells as determined by western blot. (E)Flow cytometric analysis of apoptosis in LINC00673 over-expression or knock-down in CanPan-1 cells. *P<0.05, ** P<0.01

    Journal: Journal of Cancer

    Article Title: LNC00673 suppresses proliferation and metastasis of pancreatic cancer via target miR-504/ HNF1A

    doi: 10.7150/jca.32855

    Figure Lengend Snippet: Effects of LINC00673 on pancreatic cancer cell apoptosis, viability, migration. (A)Validation of LINC00673 overexpression or knockdown in CanPan-1 cells as determined by qRT-PCR. (B) Transwell invasion assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (C) CCK-8 proliferation assay of LINC00673 over-expression or knock-down in CanPan-1 cells. (D)Expression Cleaved-Caspase 3 in LINC00673 over-expression or knock-down in CanPan-1 cells as determined by western blot. (E)Flow cytometric analysis of apoptosis in LINC00673 over-expression or knock-down in CanPan-1 cells. *P<0.05, ** P<0.01

    Article Snippet: Antibody dilution ratio is as follows Cleaved-caspase-3 (1:1000, CST, #9579), HNF1A (1:1500, abcam, #ab204306), GAPDH (1:5000, CST, #51332).

    Techniques: Migration, Over Expression, Quantitative RT-PCR, Transwell Invasion Assay, CCK-8 Assay, Proliferation Assay, Expressing, Western Blot

    HNF1A suppression inhibited cell progression in pancreatic cancer cells. CCK-8 proliferation assay of specified plasmid or RNA transfection transfected in CanPan-1 cells. (B) Transwell invasion assay in specified plasmid or RNA transfection in CanPan-1 cells. (C) Expression Cleaved-Caspase 3 in specified plasmid or RNA transfected in CanPan-1 cells as determined by western blot. (D)Flow cytometric analysis of apoptosis in specified plasmid or RNA transfected in CanPan-1cells. *P<0.05, ** P<0.01

    Journal: Journal of Cancer

    Article Title: LNC00673 suppresses proliferation and metastasis of pancreatic cancer via target miR-504/ HNF1A

    doi: 10.7150/jca.32855

    Figure Lengend Snippet: HNF1A suppression inhibited cell progression in pancreatic cancer cells. CCK-8 proliferation assay of specified plasmid or RNA transfection transfected in CanPan-1 cells. (B) Transwell invasion assay in specified plasmid or RNA transfection in CanPan-1 cells. (C) Expression Cleaved-Caspase 3 in specified plasmid or RNA transfected in CanPan-1 cells as determined by western blot. (D)Flow cytometric analysis of apoptosis in specified plasmid or RNA transfected in CanPan-1cells. *P<0.05, ** P<0.01

    Article Snippet: Antibody dilution ratio is as follows Cleaved-caspase-3 (1:1000, CST, #9579), HNF1A (1:1500, abcam, #ab204306), GAPDH (1:5000, CST, #51332).

    Techniques: CCK-8 Assay, Proliferation Assay, Plasmid Preparation, Transfection, Transwell Invasion Assay, Expressing, Western Blot

    High-fat diet (HFD; ■) consumption does not modify enteroendocrine cell proliferation or apoptosis in mice. Representative double immunofluorescent staining ( n 7–8) of glucagon-like peptide-1 (GLP-1) (green) and bromodeoxyuridine (BrdU) (red) with DAPI (4′,6-diamidino-2-phenylindole) staining (blue) in the jejunum (a) of mice fed a control diet (CD; □) or the HFD for 2 weeks. The lower panels of (a) show GLP-1 and BrdU double-positive cells whereas the upper panels of (a) show GLP-1-positive BrdU-negative cells (highlighted with dotted circles). The scale bar represents 10 μm. Quantification as percentage of proliferating GLP-1 cells over total GLP-1 cells in the jejunum (b) and colon (c). Representative immunostaining ( n 6–8) of cleaved caspase-3 positive cells in the jejunum (d) of mice fed the CD or HFD for 2 weeks. The scale bar represents 20 μm. Quantification of stained cells in the jejunum (e) and colon (f). Values are means, with standard errors represented by vertical bars. Statistical analysis was performed using a non-parametric Mann–Whitney test; there were no significant differences between the groups.

    Journal: Journal of Nutritional Science

    Article Title: Lipid-rich diet enhances L-cell density in obese subjects and in mice through improved L-cell differentiation

    doi: 10.1017/jns.2015.11

    Figure Lengend Snippet: High-fat diet (HFD; ■) consumption does not modify enteroendocrine cell proliferation or apoptosis in mice. Representative double immunofluorescent staining ( n 7–8) of glucagon-like peptide-1 (GLP-1) (green) and bromodeoxyuridine (BrdU) (red) with DAPI (4′,6-diamidino-2-phenylindole) staining (blue) in the jejunum (a) of mice fed a control diet (CD; □) or the HFD for 2 weeks. The lower panels of (a) show GLP-1 and BrdU double-positive cells whereas the upper panels of (a) show GLP-1-positive BrdU-negative cells (highlighted with dotted circles). The scale bar represents 10 μm. Quantification as percentage of proliferating GLP-1 cells over total GLP-1 cells in the jejunum (b) and colon (c). Representative immunostaining ( n 6–8) of cleaved caspase-3 positive cells in the jejunum (d) of mice fed the CD or HFD for 2 weeks. The scale bar represents 20 μm. Quantification of stained cells in the jejunum (e) and colon (f). Values are means, with standard errors represented by vertical bars. Statistical analysis was performed using a non-parametric Mann–Whitney test; there were no significant differences between the groups.

    Article Snippet: A cleaved caspase-3 antibody (no. 9579; Cell Signaling) labelled apoptotic cells.

    Techniques: Staining, Immunostaining, MANN-WHITNEY

    A) Percentage of positive cells for OCT3/4 and SOX2 in cells with reduced OCT3/4 and SOX2 levels compared to cells transfected with control siRNA in the NCCIT cells in three different time-points post transfection based on immunohistochemistry; The controls are set to 100 in all cases. B) Relative expression pattern of 32 genes representing targets for pluripotency and differentiation (ectoderm, mesoderm and endoderm). The expression levels are normalized based on the housekeeping gene HPRT ; C) Percentage of living NCCIT cells with reduced level of OCT3/4 and SOX2 compared to cells transfected with control siRNA at each time point based on Trypan blue measurement; D) Percentage of positive NCCIT cells for Ki67 in untreated cells, cells transfected with control siRNA and cells with reduced levels of OCT3/4 and SOX2 at 48+12 h post transfection; E) Percentage of positive cells for Caspase 3 in untreated NCCIT cells, Cells transfected with control siRNA show a reduced level of OCT3/4 and SOX2 at all three time points after the transfection; F) FACS analysis with Propidium Iodide staining in cells transfected with control siRNA, cells transfected with SOX2 and OCT3/4 siRNA at 60 h post transfection. NCCIT cells transfected with control siRNA show the presence of 78% living cells, while cells transfected with SOX2 siRNA show 51% living cells, NCCIT cells with OCT3/4kd show 62% live cells. G) Annexin V assay results in cells with siRNA control and cells with reduced SOX2 and OCT3/4 at 60 h after transfection. In the control, percentages of living-annexin negative cells are 89.6%, while dead-annexin positive cells are 12.2%. In SOX2kd cells, the amount of living-annexin negative cells is 57.5% while the amount of dead-annexin positive cells is 17.2%. In cells transfected with OCT3/4 siRNA, living-annexin negative cells are 68.3% while dead-annexin positive cells are 12.8%. The differences between the percentages of live and dead cells in all experiments are due to using independent and various methods in order to prove apoptosis and different sensitivity of the materials and methods used. As it is demonstrated, OCT3/4 knock-down also induces apoptosis, however, this effect is not as dramatic as SOX2 knock-down. The FACS analysis were performed in triplicate.

    Journal: PLoS ONE

    Article Title: Role of SOX2 in the Etiology of Embryonal Carcinoma, Based on Analysis of the NCCIT and NT2 Cell Lines

    doi: 10.1371/journal.pone.0083585

    Figure Lengend Snippet: A) Percentage of positive cells for OCT3/4 and SOX2 in cells with reduced OCT3/4 and SOX2 levels compared to cells transfected with control siRNA in the NCCIT cells in three different time-points post transfection based on immunohistochemistry; The controls are set to 100 in all cases. B) Relative expression pattern of 32 genes representing targets for pluripotency and differentiation (ectoderm, mesoderm and endoderm). The expression levels are normalized based on the housekeeping gene HPRT ; C) Percentage of living NCCIT cells with reduced level of OCT3/4 and SOX2 compared to cells transfected with control siRNA at each time point based on Trypan blue measurement; D) Percentage of positive NCCIT cells for Ki67 in untreated cells, cells transfected with control siRNA and cells with reduced levels of OCT3/4 and SOX2 at 48+12 h post transfection; E) Percentage of positive cells for Caspase 3 in untreated NCCIT cells, Cells transfected with control siRNA show a reduced level of OCT3/4 and SOX2 at all three time points after the transfection; F) FACS analysis with Propidium Iodide staining in cells transfected with control siRNA, cells transfected with SOX2 and OCT3/4 siRNA at 60 h post transfection. NCCIT cells transfected with control siRNA show the presence of 78% living cells, while cells transfected with SOX2 siRNA show 51% living cells, NCCIT cells with OCT3/4kd show 62% live cells. G) Annexin V assay results in cells with siRNA control and cells with reduced SOX2 and OCT3/4 at 60 h after transfection. In the control, percentages of living-annexin negative cells are 89.6%, while dead-annexin positive cells are 12.2%. In SOX2kd cells, the amount of living-annexin negative cells is 57.5% while the amount of dead-annexin positive cells is 17.2%. In cells transfected with OCT3/4 siRNA, living-annexin negative cells are 68.3% while dead-annexin positive cells are 12.8%. The differences between the percentages of live and dead cells in all experiments are due to using independent and various methods in order to prove apoptosis and different sensitivity of the materials and methods used. As it is demonstrated, OCT3/4 knock-down also induces apoptosis, however, this effect is not as dramatic as SOX2 knock-down. The FACS analysis were performed in triplicate.

    Article Snippet: Antibodies used were SOX2 (1∶250, AF2018; R&D System, USA), OCT3/4 (sc-5279; 1∶350; sc-Santa Cruz Biotechnology, Santa Cruz, CA, USA), Ki67 (1∶50, A047; Dako, CA, USA) and Caspase 3 (1∶500, 9579; Cell Signaling).

    Techniques: Transfection, Immunohistochemistry, Expressing, Staining, Annexin V Assay