igg  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc igg
    Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc igg
    Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc igg
    Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc igg
    Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc anti igg
    GFPT1 is critical for endogenous glutamate to determine ferroptosis sensitivity. (A) Schematic overview of the HBP signaling from glucose to O-GlcNAcylation. (B) Endogenous glutamate affects HBP metabolites. Indicated metabolites were measured in H1975-based GGG cells under the same treatment as that in Figure D. (C) Relative GFPT1 activity was measured in GGG cells under the same treatment as that in Figure D but treated with or without erastin (10 µM) for 8 h. The GFPT1 activity was normalized between those treated with erastin and DMSO. (D) Metabolites downstream of GFPT1 <t>restore</t> <t>YAP</t> alteration by erastin. p-YAP S127 , O-YAP T241 and YAP were measured by IB in H1975 cells treated with or without erastin (10 µM), glucose (25 mM), GlcN (5 mM) or GlcNAc (5 mM) for 8 h. (E-F) GlcN and GlcNAc diminish ferroptosis sensitivity. LUAD cells were treated with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM), in the presence or absence of GlcN (5 mM) or GlcNAc (5 mM). Cell death was measured after treatment for 24 h (E). MDA was measured after treatment for 16 h (F). (G) GFPT1 S205A restores YAP alteration by erastin. FLAG-GFPT1 S205A was ectopically expressed in H1975 cells prior to the treatment with or without erastin (10 µM) for 8 h. Indicated proteins were analyzed by IB. (H) GFPT1 boosts protein stability of YAP. CHX (10 µg/ml) chase experiments were performed in H1975 cells with or without GFPT1 overexpression or knockout. The relative protein levels of YAP were shown as the ratios between YAP and GAPDH, and the “0 h” points were arbitrarily set to 100%. (I) GFPT1 reduces ubiquitination of YAP. The ubiquitination of YAP was measured in control cells and H1975 cells with or without GFPT1 overexpression or knockout using anti-Ub antibodies following immunoprecipitation using anti-YAP antibodies. <t>IgG</t> antibodies were used as negative controls. (J-K) Restore of GFPT1 diminishes ferroptosis sensitivity. FLAG-GFPT1 S205A was ectopically expressed in LUAD cells before treatment with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM). Cell death was measured after treatment for 24 h by SYTOX green staining followed by flow cytometry (J). MDA was measured after treatment for 16 h (K). The data are shown as the mean ± SD from three biological replicates (including IB). **P < 0.01 indicates statistical significance. Data in B were analyzed using a one-way ANOVA test. Data in C, E, F, J, K were analyzed using a two-way ANOVA test.
    Anti Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti igg/product/Cell Signaling Technology Inc
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    Images

    1) Product Images from "Endogenous glutamate determines ferroptosis sensitivity via ADCY10-dependent YAP suppression in lung adenocarcinoma"

    Article Title: Endogenous glutamate determines ferroptosis sensitivity via ADCY10-dependent YAP suppression in lung adenocarcinoma

    Journal: Theranostics

    doi: 10.7150/thno.55482

    GFPT1 is critical for endogenous glutamate to determine ferroptosis sensitivity. (A) Schematic overview of the HBP signaling from glucose to O-GlcNAcylation. (B) Endogenous glutamate affects HBP metabolites. Indicated metabolites were measured in H1975-based GGG cells under the same treatment as that in Figure D. (C) Relative GFPT1 activity was measured in GGG cells under the same treatment as that in Figure D but treated with or without erastin (10 µM) for 8 h. The GFPT1 activity was normalized between those treated with erastin and DMSO. (D) Metabolites downstream of GFPT1 restore YAP alteration by erastin. p-YAP S127 , O-YAP T241 and YAP were measured by IB in H1975 cells treated with or without erastin (10 µM), glucose (25 mM), GlcN (5 mM) or GlcNAc (5 mM) for 8 h. (E-F) GlcN and GlcNAc diminish ferroptosis sensitivity. LUAD cells were treated with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM), in the presence or absence of GlcN (5 mM) or GlcNAc (5 mM). Cell death was measured after treatment for 24 h (E). MDA was measured after treatment for 16 h (F). (G) GFPT1 S205A restores YAP alteration by erastin. FLAG-GFPT1 S205A was ectopically expressed in H1975 cells prior to the treatment with or without erastin (10 µM) for 8 h. Indicated proteins were analyzed by IB. (H) GFPT1 boosts protein stability of YAP. CHX (10 µg/ml) chase experiments were performed in H1975 cells with or without GFPT1 overexpression or knockout. The relative protein levels of YAP were shown as the ratios between YAP and GAPDH, and the “0 h” points were arbitrarily set to 100%. (I) GFPT1 reduces ubiquitination of YAP. The ubiquitination of YAP was measured in control cells and H1975 cells with or without GFPT1 overexpression or knockout using anti-Ub antibodies following immunoprecipitation using anti-YAP antibodies. IgG antibodies were used as negative controls. (J-K) Restore of GFPT1 diminishes ferroptosis sensitivity. FLAG-GFPT1 S205A was ectopically expressed in LUAD cells before treatment with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM). Cell death was measured after treatment for 24 h by SYTOX green staining followed by flow cytometry (J). MDA was measured after treatment for 16 h (K). The data are shown as the mean ± SD from three biological replicates (including IB). **P < 0.01 indicates statistical significance. Data in B were analyzed using a one-way ANOVA test. Data in C, E, F, J, K were analyzed using a two-way ANOVA test.
    Figure Legend Snippet: GFPT1 is critical for endogenous glutamate to determine ferroptosis sensitivity. (A) Schematic overview of the HBP signaling from glucose to O-GlcNAcylation. (B) Endogenous glutamate affects HBP metabolites. Indicated metabolites were measured in H1975-based GGG cells under the same treatment as that in Figure D. (C) Relative GFPT1 activity was measured in GGG cells under the same treatment as that in Figure D but treated with or without erastin (10 µM) for 8 h. The GFPT1 activity was normalized between those treated with erastin and DMSO. (D) Metabolites downstream of GFPT1 restore YAP alteration by erastin. p-YAP S127 , O-YAP T241 and YAP were measured by IB in H1975 cells treated with or without erastin (10 µM), glucose (25 mM), GlcN (5 mM) or GlcNAc (5 mM) for 8 h. (E-F) GlcN and GlcNAc diminish ferroptosis sensitivity. LUAD cells were treated with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM), in the presence or absence of GlcN (5 mM) or GlcNAc (5 mM). Cell death was measured after treatment for 24 h (E). MDA was measured after treatment for 16 h (F). (G) GFPT1 S205A restores YAP alteration by erastin. FLAG-GFPT1 S205A was ectopically expressed in H1975 cells prior to the treatment with or without erastin (10 µM) for 8 h. Indicated proteins were analyzed by IB. (H) GFPT1 boosts protein stability of YAP. CHX (10 µg/ml) chase experiments were performed in H1975 cells with or without GFPT1 overexpression or knockout. The relative protein levels of YAP were shown as the ratios between YAP and GAPDH, and the “0 h” points were arbitrarily set to 100%. (I) GFPT1 reduces ubiquitination of YAP. The ubiquitination of YAP was measured in control cells and H1975 cells with or without GFPT1 overexpression or knockout using anti-Ub antibodies following immunoprecipitation using anti-YAP antibodies. IgG antibodies were used as negative controls. (J-K) Restore of GFPT1 diminishes ferroptosis sensitivity. FLAG-GFPT1 S205A was ectopically expressed in LUAD cells before treatment with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM). Cell death was measured after treatment for 24 h by SYTOX green staining followed by flow cytometry (J). MDA was measured after treatment for 16 h (K). The data are shown as the mean ± SD from three biological replicates (including IB). **P < 0.01 indicates statistical significance. Data in B were analyzed using a one-way ANOVA test. Data in C, E, F, J, K were analyzed using a two-way ANOVA test.

    Techniques Used: Activity Assay, Over Expression, Knock-Out, Immunoprecipitation, Staining, Flow Cytometry

    igg  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc igg
    GFPT1 is critical for endogenous glutamate to determine ferroptosis sensitivity. (A) Schematic overview of the HBP signaling from glucose to O-GlcNAcylation. (B) Endogenous glutamate affects HBP metabolites. Indicated metabolites were measured in H1975-based GGG cells under the same treatment as that in Figure D. (C) Relative GFPT1 activity was measured in GGG cells under the same treatment as that in Figure D but treated with or without erastin (10 µM) for 8 h. The GFPT1 activity was normalized between those treated with erastin and DMSO. (D) Metabolites downstream of GFPT1 restore YAP alteration by erastin. p-YAP S127 , O-YAP T241 and YAP were measured by IB in H1975 cells treated with or without erastin (10 µM), glucose (25 mM), GlcN (5 mM) or GlcNAc (5 mM) for 8 h. (E-F) GlcN and GlcNAc diminish ferroptosis sensitivity. LUAD cells were treated with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM), in the presence or absence of GlcN (5 mM) or GlcNAc (5 mM). Cell death was measured after treatment for 24 h (E). MDA was measured after treatment for 16 h (F). (G) GFPT1 S205A restores YAP alteration by erastin. FLAG-GFPT1 S205A was ectopically expressed in H1975 cells prior to the treatment with or without erastin (10 µM) for 8 h. Indicated proteins were analyzed by IB. (H) GFPT1 boosts protein stability of YAP. CHX (10 µg/ml) chase experiments were performed in H1975 cells with or without GFPT1 overexpression or knockout. The relative protein levels of YAP were shown as the ratios between YAP and GAPDH, and the “0 h” points were arbitrarily set to 100%. (I) GFPT1 reduces ubiquitination of YAP. The ubiquitination of YAP was measured in control cells and H1975 cells with or without GFPT1 overexpression or knockout using <t>anti-Ub</t> <t>antibodies</t> following immunoprecipitation using anti-YAP antibodies. <t>IgG</t> antibodies were used as negative controls. (J-K) Restore of GFPT1 diminishes ferroptosis sensitivity. FLAG-GFPT1 S205A was ectopically expressed in LUAD cells before treatment with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM). Cell death was measured after treatment for 24 h by SYTOX green staining followed by flow cytometry (J). MDA was measured after treatment for 16 h (K). The data are shown as the mean ± SD from three biological replicates (including IB). **P < 0.01 indicates statistical significance. Data in B were analyzed using a one-way ANOVA test. Data in C, E, F, J, K were analyzed using a two-way ANOVA test.
    Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg/product/Cell Signaling Technology Inc
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    Images

    1) Product Images from "Endogenous glutamate determines ferroptosis sensitivity via ADCY10-dependent YAP suppression in lung adenocarcinoma"

    Article Title: Endogenous glutamate determines ferroptosis sensitivity via ADCY10-dependent YAP suppression in lung adenocarcinoma

    Journal: Theranostics

    doi: 10.7150/thno.55482

    GFPT1 is critical for endogenous glutamate to determine ferroptosis sensitivity. (A) Schematic overview of the HBP signaling from glucose to O-GlcNAcylation. (B) Endogenous glutamate affects HBP metabolites. Indicated metabolites were measured in H1975-based GGG cells under the same treatment as that in Figure D. (C) Relative GFPT1 activity was measured in GGG cells under the same treatment as that in Figure D but treated with or without erastin (10 µM) for 8 h. The GFPT1 activity was normalized between those treated with erastin and DMSO. (D) Metabolites downstream of GFPT1 restore YAP alteration by erastin. p-YAP S127 , O-YAP T241 and YAP were measured by IB in H1975 cells treated with or without erastin (10 µM), glucose (25 mM), GlcN (5 mM) or GlcNAc (5 mM) for 8 h. (E-F) GlcN and GlcNAc diminish ferroptosis sensitivity. LUAD cells were treated with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM), in the presence or absence of GlcN (5 mM) or GlcNAc (5 mM). Cell death was measured after treatment for 24 h (E). MDA was measured after treatment for 16 h (F). (G) GFPT1 S205A restores YAP alteration by erastin. FLAG-GFPT1 S205A was ectopically expressed in H1975 cells prior to the treatment with or without erastin (10 µM) for 8 h. Indicated proteins were analyzed by IB. (H) GFPT1 boosts protein stability of YAP. CHX (10 µg/ml) chase experiments were performed in H1975 cells with or without GFPT1 overexpression or knockout. The relative protein levels of YAP were shown as the ratios between YAP and GAPDH, and the “0 h” points were arbitrarily set to 100%. (I) GFPT1 reduces ubiquitination of YAP. The ubiquitination of YAP was measured in control cells and H1975 cells with or without GFPT1 overexpression or knockout using anti-Ub antibodies following immunoprecipitation using anti-YAP antibodies. IgG antibodies were used as negative controls. (J-K) Restore of GFPT1 diminishes ferroptosis sensitivity. FLAG-GFPT1 S205A was ectopically expressed in LUAD cells before treatment with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM). Cell death was measured after treatment for 24 h by SYTOX green staining followed by flow cytometry (J). MDA was measured after treatment for 16 h (K). The data are shown as the mean ± SD from three biological replicates (including IB). **P < 0.01 indicates statistical significance. Data in B were analyzed using a one-way ANOVA test. Data in C, E, F, J, K were analyzed using a two-way ANOVA test.
    Figure Legend Snippet: GFPT1 is critical for endogenous glutamate to determine ferroptosis sensitivity. (A) Schematic overview of the HBP signaling from glucose to O-GlcNAcylation. (B) Endogenous glutamate affects HBP metabolites. Indicated metabolites were measured in H1975-based GGG cells under the same treatment as that in Figure D. (C) Relative GFPT1 activity was measured in GGG cells under the same treatment as that in Figure D but treated with or without erastin (10 µM) for 8 h. The GFPT1 activity was normalized between those treated with erastin and DMSO. (D) Metabolites downstream of GFPT1 restore YAP alteration by erastin. p-YAP S127 , O-YAP T241 and YAP were measured by IB in H1975 cells treated with or without erastin (10 µM), glucose (25 mM), GlcN (5 mM) or GlcNAc (5 mM) for 8 h. (E-F) GlcN and GlcNAc diminish ferroptosis sensitivity. LUAD cells were treated with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM), in the presence or absence of GlcN (5 mM) or GlcNAc (5 mM). Cell death was measured after treatment for 24 h (E). MDA was measured after treatment for 16 h (F). (G) GFPT1 S205A restores YAP alteration by erastin. FLAG-GFPT1 S205A was ectopically expressed in H1975 cells prior to the treatment with or without erastin (10 µM) for 8 h. Indicated proteins were analyzed by IB. (H) GFPT1 boosts protein stability of YAP. CHX (10 µg/ml) chase experiments were performed in H1975 cells with or without GFPT1 overexpression or knockout. The relative protein levels of YAP were shown as the ratios between YAP and GAPDH, and the “0 h” points were arbitrarily set to 100%. (I) GFPT1 reduces ubiquitination of YAP. The ubiquitination of YAP was measured in control cells and H1975 cells with or without GFPT1 overexpression or knockout using anti-Ub antibodies following immunoprecipitation using anti-YAP antibodies. IgG antibodies were used as negative controls. (J-K) Restore of GFPT1 diminishes ferroptosis sensitivity. FLAG-GFPT1 S205A was ectopically expressed in LUAD cells before treatment with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM). Cell death was measured after treatment for 24 h by SYTOX green staining followed by flow cytometry (J). MDA was measured after treatment for 16 h (K). The data are shown as the mean ± SD from three biological replicates (including IB). **P < 0.01 indicates statistical significance. Data in B were analyzed using a one-way ANOVA test. Data in C, E, F, J, K were analyzed using a two-way ANOVA test.

    Techniques Used: Activity Assay, Over Expression, Knock-Out, Immunoprecipitation, Staining, Flow Cytometry

    anti igg  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc anti igg
    Anti Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti igg/product/Cell Signaling Technology Inc
    Average 97 stars, based on 1 article reviews
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    igg  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc igg
    Primary antibodies used for immunohistochemistry.
    Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg/product/Cell Signaling Technology Inc
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    1) Product Images from "Selfish Spermatogonial Selection: Evidence from an Immunohistochemical Screen in Testes of Elderly Men"

    Article Title: Selfish Spermatogonial Selection: Evidence from an Immunohistochemical Screen in Testes of Elderly Men

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0042382

    Primary antibodies used for immunohistochemistry.
    Figure Legend Snippet: Primary antibodies used for immunohistochemistry.

    Techniques Used: Immunohistochemistry, Staining

    igg  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc igg
    Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg/product/Cell Signaling Technology Inc
    Average 97 stars, based on 1 article reviews
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    igg  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc igg
    Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg/product/Cell Signaling Technology Inc
    Average 97 stars, based on 1 article reviews
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    igg  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc igg
    List of primary antibodies.
    Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/igg/product/Cell Signaling Technology Inc
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    1) Product Images from "Molecular Markers for Granulovacuolar Degeneration Are Present in Rimmed Vacuoles"

    Article Title: Molecular Markers for Granulovacuolar Degeneration Are Present in Rimmed Vacuoles

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0080995

    List of primary antibodies.
    Figure Legend Snippet: List of primary antibodies.

    Techniques Used:

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    Cell Signaling Technology Inc igg
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    Cell Signaling Technology Inc anti igg
    GFPT1 is critical for endogenous glutamate to determine ferroptosis sensitivity. (A) Schematic overview of the HBP signaling from glucose to O-GlcNAcylation. (B) Endogenous glutamate affects HBP metabolites. Indicated metabolites were measured in H1975-based GGG cells under the same treatment as that in Figure D. (C) Relative GFPT1 activity was measured in GGG cells under the same treatment as that in Figure D but treated with or without erastin (10 µM) for 8 h. The GFPT1 activity was normalized between those treated with erastin and DMSO. (D) Metabolites downstream of GFPT1 <t>restore</t> <t>YAP</t> alteration by erastin. p-YAP S127 , O-YAP T241 and YAP were measured by IB in H1975 cells treated with or without erastin (10 µM), glucose (25 mM), GlcN (5 mM) or GlcNAc (5 mM) for 8 h. (E-F) GlcN and GlcNAc diminish ferroptosis sensitivity. LUAD cells were treated with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM), in the presence or absence of GlcN (5 mM) or GlcNAc (5 mM). Cell death was measured after treatment for 24 h (E). MDA was measured after treatment for 16 h (F). (G) GFPT1 S205A restores YAP alteration by erastin. FLAG-GFPT1 S205A was ectopically expressed in H1975 cells prior to the treatment with or without erastin (10 µM) for 8 h. Indicated proteins were analyzed by IB. (H) GFPT1 boosts protein stability of YAP. CHX (10 µg/ml) chase experiments were performed in H1975 cells with or without GFPT1 overexpression or knockout. The relative protein levels of YAP were shown as the ratios between YAP and GAPDH, and the “0 h” points were arbitrarily set to 100%. (I) GFPT1 reduces ubiquitination of YAP. The ubiquitination of YAP was measured in control cells and H1975 cells with or without GFPT1 overexpression or knockout using anti-Ub antibodies following immunoprecipitation using anti-YAP antibodies. <t>IgG</t> antibodies were used as negative controls. (J-K) Restore of GFPT1 diminishes ferroptosis sensitivity. FLAG-GFPT1 S205A was ectopically expressed in LUAD cells before treatment with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM). Cell death was measured after treatment for 24 h by SYTOX green staining followed by flow cytometry (J). MDA was measured after treatment for 16 h (K). The data are shown as the mean ± SD from three biological replicates (including IB). **P < 0.01 indicates statistical significance. Data in B were analyzed using a one-way ANOVA test. Data in C, E, F, J, K were analyzed using a two-way ANOVA test.
    Anti Igg, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GFPT1 is critical for endogenous glutamate to determine ferroptosis sensitivity. (A) Schematic overview of the HBP signaling from glucose to O-GlcNAcylation. (B) Endogenous glutamate affects HBP metabolites. Indicated metabolites were measured in H1975-based GGG cells under the same treatment as that in Figure D. (C) Relative GFPT1 activity was measured in GGG cells under the same treatment as that in Figure D but treated with or without erastin (10 µM) for 8 h. The GFPT1 activity was normalized between those treated with erastin and DMSO. (D) Metabolites downstream of GFPT1 restore YAP alteration by erastin. p-YAP S127 , O-YAP T241 and YAP were measured by IB in H1975 cells treated with or without erastin (10 µM), glucose (25 mM), GlcN (5 mM) or GlcNAc (5 mM) for 8 h. (E-F) GlcN and GlcNAc diminish ferroptosis sensitivity. LUAD cells were treated with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM), in the presence or absence of GlcN (5 mM) or GlcNAc (5 mM). Cell death was measured after treatment for 24 h (E). MDA was measured after treatment for 16 h (F). (G) GFPT1 S205A restores YAP alteration by erastin. FLAG-GFPT1 S205A was ectopically expressed in H1975 cells prior to the treatment with or without erastin (10 µM) for 8 h. Indicated proteins were analyzed by IB. (H) GFPT1 boosts protein stability of YAP. CHX (10 µg/ml) chase experiments were performed in H1975 cells with or without GFPT1 overexpression or knockout. The relative protein levels of YAP were shown as the ratios between YAP and GAPDH, and the “0 h” points were arbitrarily set to 100%. (I) GFPT1 reduces ubiquitination of YAP. The ubiquitination of YAP was measured in control cells and H1975 cells with or without GFPT1 overexpression or knockout using anti-Ub antibodies following immunoprecipitation using anti-YAP antibodies. IgG antibodies were used as negative controls. (J-K) Restore of GFPT1 diminishes ferroptosis sensitivity. FLAG-GFPT1 S205A was ectopically expressed in LUAD cells before treatment with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM). Cell death was measured after treatment for 24 h by SYTOX green staining followed by flow cytometry (J). MDA was measured after treatment for 16 h (K). The data are shown as the mean ± SD from three biological replicates (including IB). **P < 0.01 indicates statistical significance. Data in B were analyzed using a one-way ANOVA test. Data in C, E, F, J, K were analyzed using a two-way ANOVA test.

    Journal: Theranostics

    Article Title: Endogenous glutamate determines ferroptosis sensitivity via ADCY10-dependent YAP suppression in lung adenocarcinoma

    doi: 10.7150/thno.55482

    Figure Lengend Snippet: GFPT1 is critical for endogenous glutamate to determine ferroptosis sensitivity. (A) Schematic overview of the HBP signaling from glucose to O-GlcNAcylation. (B) Endogenous glutamate affects HBP metabolites. Indicated metabolites were measured in H1975-based GGG cells under the same treatment as that in Figure D. (C) Relative GFPT1 activity was measured in GGG cells under the same treatment as that in Figure D but treated with or without erastin (10 µM) for 8 h. The GFPT1 activity was normalized between those treated with erastin and DMSO. (D) Metabolites downstream of GFPT1 restore YAP alteration by erastin. p-YAP S127 , O-YAP T241 and YAP were measured by IB in H1975 cells treated with or without erastin (10 µM), glucose (25 mM), GlcN (5 mM) or GlcNAc (5 mM) for 8 h. (E-F) GlcN and GlcNAc diminish ferroptosis sensitivity. LUAD cells were treated with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM), in the presence or absence of GlcN (5 mM) or GlcNAc (5 mM). Cell death was measured after treatment for 24 h (E). MDA was measured after treatment for 16 h (F). (G) GFPT1 S205A restores YAP alteration by erastin. FLAG-GFPT1 S205A was ectopically expressed in H1975 cells prior to the treatment with or without erastin (10 µM) for 8 h. Indicated proteins were analyzed by IB. (H) GFPT1 boosts protein stability of YAP. CHX (10 µg/ml) chase experiments were performed in H1975 cells with or without GFPT1 overexpression or knockout. The relative protein levels of YAP were shown as the ratios between YAP and GAPDH, and the “0 h” points were arbitrarily set to 100%. (I) GFPT1 reduces ubiquitination of YAP. The ubiquitination of YAP was measured in control cells and H1975 cells with or without GFPT1 overexpression or knockout using anti-Ub antibodies following immunoprecipitation using anti-YAP antibodies. IgG antibodies were used as negative controls. (J-K) Restore of GFPT1 diminishes ferroptosis sensitivity. FLAG-GFPT1 S205A was ectopically expressed in LUAD cells before treatment with erastin (10 µM) with or without Fer-1 (1 µM) or DFO (80 µM). Cell death was measured after treatment for 24 h by SYTOX green staining followed by flow cytometry (J). MDA was measured after treatment for 16 h (K). The data are shown as the mean ± SD from three biological replicates (including IB). **P < 0.01 indicates statistical significance. Data in B were analyzed using a one-way ANOVA test. Data in C, E, F, J, K were analyzed using a two-way ANOVA test.

    Article Snippet: The antibodies used for co-IP were: anti-FLAG (CST, #8146 or #2368), anti-YAP (Santa Cruz Biotechnology, #sc-101199), anti-βTrCP (Abcam, #ab233638) and anti-IgG (CST, #3900).

    Techniques: Activity Assay, Over Expression, Knock-Out, Immunoprecipitation, Staining, Flow Cytometry