pi3k activity elisa kit  (Echelon Biosciences)


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    Echelon Biosciences pi3k activity elisa kit
    Expression and activation of IGF-IR and <t>PI3K</t> in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.
    Pi3k Activity Elisa Kit, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pi3k activity elisa kit - by Bioz Stars, 2023-01
    95/100 stars

    Images

    1) Product Images from "Rac1 and Stathmin but Not EB1 Are Required for Invasion of Breast Cancer Cells in Response to IGF-I"

    Article Title: Rac1 and Stathmin but Not EB1 Are Required for Invasion of Breast Cancer Cells in Response to IGF-I

    Journal: International Journal of Cell Biology

    doi: 10.1155/2011/615912

    Expression and activation of IGF-IR and PI3K in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.
    Figure Legend Snippet: Expression and activation of IGF-IR and PI3K in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.

    Techniques Used: Expressing, Activation Assay, Western Blot, Immunoprecipitation, Activity Assay

    pi3k activity elisa kit  (Echelon Biosciences)


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    Echelon Biosciences pi3k activity elisa kit
    Expression and activation of IGF-IR and <t>PI3K</t> in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.
    Pi3k Activity Elisa Kit, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pi3k activity elisa kit/product/Echelon Biosciences
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pi3k activity elisa kit - by Bioz Stars, 2023-01
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    Images

    1) Product Images from "Rac1 and Stathmin but Not EB1 Are Required for Invasion of Breast Cancer Cells in Response to IGF-I"

    Article Title: Rac1 and Stathmin but Not EB1 Are Required for Invasion of Breast Cancer Cells in Response to IGF-I

    Journal: International Journal of Cell Biology

    doi: 10.1155/2011/615912

    Expression and activation of IGF-IR and PI3K in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.
    Figure Legend Snippet: Expression and activation of IGF-IR and PI3K in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.

    Techniques Used: Expressing, Activation Assay, Western Blot, Immunoprecipitation, Activity Assay

    pi3k activity  (Echelon Biosciences)


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    Echelon Biosciences pi3k activity
    Expression and activation of IGF-IR and <t>PI3K</t> in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.
    Pi3k Activity, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
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    Images

    1) Product Images from "Rac1 and Stathmin but Not EB1 Are Required for Invasion of Breast Cancer Cells in Response to IGF-I"

    Article Title: Rac1 and Stathmin but Not EB1 Are Required for Invasion of Breast Cancer Cells in Response to IGF-I

    Journal: International Journal of Cell Biology

    doi: 10.1155/2011/615912

    Expression and activation of IGF-IR and PI3K in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.
    Figure Legend Snippet: Expression and activation of IGF-IR and PI3K in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.

    Techniques Used: Expressing, Activation Assay, Western Blot, Immunoprecipitation, Activity Assay

    pi3k activity  (Echelon Biosciences)


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    Echelon Biosciences pi3k activity
    Slit2N inhibits VEGF-C-enhanced <t>PI3K</t> activity and Akt phosphorylation in L-LECs. (A) Representative Western blot analysis of phosphorylated ERK1/2 in L-LECs, after treatment with control, VEGF-C alone [100 ng/ml]; or after preincubation with various concentrations of Slit2N, then VEGF-C. Total ERK1/2 used as loading control. (B) Quantitative analysis of phosphorylated ERK1/2 in L-LECs, after treatment with control, VEGF-C alone [100 ng/ml]; or after preincubation with various concentrations of Slit2N, then VEGF-C. Band intensity of each lane from Figure 4A was determined by densitometry. The ratio of p-ERK1/2 to total ERK1/2 of L-LECs incubated with VEGF-C alone was set to “1” and values of all other ratios were calculated vs. this control. Data represent the mean ± SD of 3 independent experiments. (C) PI3K activity by ELISA in L-LECs incubated with various concentrations of Slit2N and/or VEGF-C [100 ng/ml]. Data represent the mean ± SD of 3 independent experiments (**p < 0.01, ***p < 0.001). (D) Representative Western blot analysis of phosphorylated Akt in L-LECs incubated with a control, VEGF-C [100 ng/ml]; or preincubated with various concentrations of Slit2N, then VEGF-C. Total Akt used as loading control. (E) Quantitative analysis of phosphorylated Akt in L-LECs, after treatment with control, VEGF-C alone [100 ng/ml]; or after preincubation with various concentrations of Slit2N, then VEGF-C. Band intensity of each lane from Figure 4D was determined by densitometry. The ratio of p-Akt to total Akt of L-LECs incubated with VEGF-C alone was set to “1” and values of all other ratios were calculated vs. this control. Data represent the mean ± SD of 3 independent experiments (**p < 0.01).
    Pi3k Activity, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pi3k activity/product/Echelon Biosciences
    Average 95 stars, based on 1 article reviews
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    pi3k activity - by Bioz Stars, 2023-01
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    1) Product Images from "Slit2N and Robo4 regulate lymphangiogenesis through the VEGF-C/VEGFR-3 pathway"

    Article Title: Slit2N and Robo4 regulate lymphangiogenesis through the VEGF-C/VEGFR-3 pathway

    Journal: Cell Communication and Signaling : CCS

    doi: 10.1186/1478-811X-12-25

    Slit2N inhibits VEGF-C-enhanced PI3K activity and Akt phosphorylation in L-LECs. (A) Representative Western blot analysis of phosphorylated ERK1/2 in L-LECs, after treatment with control, VEGF-C alone [100 ng/ml]; or after preincubation with various concentrations of Slit2N, then VEGF-C. Total ERK1/2 used as loading control. (B) Quantitative analysis of phosphorylated ERK1/2 in L-LECs, after treatment with control, VEGF-C alone [100 ng/ml]; or after preincubation with various concentrations of Slit2N, then VEGF-C. Band intensity of each lane from Figure 4A was determined by densitometry. The ratio of p-ERK1/2 to total ERK1/2 of L-LECs incubated with VEGF-C alone was set to “1” and values of all other ratios were calculated vs. this control. Data represent the mean ± SD of 3 independent experiments. (C) PI3K activity by ELISA in L-LECs incubated with various concentrations of Slit2N and/or VEGF-C [100 ng/ml]. Data represent the mean ± SD of 3 independent experiments (**p < 0.01, ***p < 0.001). (D) Representative Western blot analysis of phosphorylated Akt in L-LECs incubated with a control, VEGF-C [100 ng/ml]; or preincubated with various concentrations of Slit2N, then VEGF-C. Total Akt used as loading control. (E) Quantitative analysis of phosphorylated Akt in L-LECs, after treatment with control, VEGF-C alone [100 ng/ml]; or after preincubation with various concentrations of Slit2N, then VEGF-C. Band intensity of each lane from Figure 4D was determined by densitometry. The ratio of p-Akt to total Akt of L-LECs incubated with VEGF-C alone was set to “1” and values of all other ratios were calculated vs. this control. Data represent the mean ± SD of 3 independent experiments (**p < 0.01).
    Figure Legend Snippet: Slit2N inhibits VEGF-C-enhanced PI3K activity and Akt phosphorylation in L-LECs. (A) Representative Western blot analysis of phosphorylated ERK1/2 in L-LECs, after treatment with control, VEGF-C alone [100 ng/ml]; or after preincubation with various concentrations of Slit2N, then VEGF-C. Total ERK1/2 used as loading control. (B) Quantitative analysis of phosphorylated ERK1/2 in L-LECs, after treatment with control, VEGF-C alone [100 ng/ml]; or after preincubation with various concentrations of Slit2N, then VEGF-C. Band intensity of each lane from Figure 4A was determined by densitometry. The ratio of p-ERK1/2 to total ERK1/2 of L-LECs incubated with VEGF-C alone was set to “1” and values of all other ratios were calculated vs. this control. Data represent the mean ± SD of 3 independent experiments. (C) PI3K activity by ELISA in L-LECs incubated with various concentrations of Slit2N and/or VEGF-C [100 ng/ml]. Data represent the mean ± SD of 3 independent experiments (**p < 0.01, ***p < 0.001). (D) Representative Western blot analysis of phosphorylated Akt in L-LECs incubated with a control, VEGF-C [100 ng/ml]; or preincubated with various concentrations of Slit2N, then VEGF-C. Total Akt used as loading control. (E) Quantitative analysis of phosphorylated Akt in L-LECs, after treatment with control, VEGF-C alone [100 ng/ml]; or after preincubation with various concentrations of Slit2N, then VEGF-C. Band intensity of each lane from Figure 4D was determined by densitometry. The ratio of p-Akt to total Akt of L-LECs incubated with VEGF-C alone was set to “1” and values of all other ratios were calculated vs. this control. Data represent the mean ± SD of 3 independent experiments (**p < 0.01).

    Techniques Used: Activity Assay, Western Blot, Incubation, Enzyme-linked Immunosorbent Assay

    Slit2N inhibition of VEGF-C-enhanced PI3K activity and Akt phosphorylation in L-LECs is Robo4-dependent. (A) PI3K activity by ELISA in L-LECs transfected with control siRNAs or Robo4-specific siRNAs, incubated with control or VEGF-C [100 ng/ml]; or after pretreatment with 10 nM Slit2N, then VEGF-C. Data represent the mean ± SD of 3 independent experiments (*p < 0.05; NS: not statistically significant). (B) Representative Western blot analysis of Akt phosphorylation in L-LECs transiently transfected with control siRNAs or Robo4-specific siRNAs, and subsequently incubated with 10 nM Slit2N, VEGF-C [100 ng/ml]; or after pretreatment with Slit2N, then VEGF-C. Total Akt used as loading control. (C) Quantitative analysis of Akt phosphorylation in L-LECs transiently transfected with control siRNAs or Robo4-specific siRNAs, and subsequently incubated with 10 nM Slit2N, VEGF-C [100 ng/ml]; or after pretreatment with Slit2N, then VEGF-C. Band intensity of each lane from Figure 6B was determined by densitometry. The ratio of p-Akt/total Akt in control siRNA-transfected L-LECs incubated with VEGF-C alone was set to “1” and all other ratios were determined vs. this control. Data represent the mean ± SD of 3 independent experiments (***p < 0.001).
    Figure Legend Snippet: Slit2N inhibition of VEGF-C-enhanced PI3K activity and Akt phosphorylation in L-LECs is Robo4-dependent. (A) PI3K activity by ELISA in L-LECs transfected with control siRNAs or Robo4-specific siRNAs, incubated with control or VEGF-C [100 ng/ml]; or after pretreatment with 10 nM Slit2N, then VEGF-C. Data represent the mean ± SD of 3 independent experiments (*p < 0.05; NS: not statistically significant). (B) Representative Western blot analysis of Akt phosphorylation in L-LECs transiently transfected with control siRNAs or Robo4-specific siRNAs, and subsequently incubated with 10 nM Slit2N, VEGF-C [100 ng/ml]; or after pretreatment with Slit2N, then VEGF-C. Total Akt used as loading control. (C) Quantitative analysis of Akt phosphorylation in L-LECs transiently transfected with control siRNAs or Robo4-specific siRNAs, and subsequently incubated with 10 nM Slit2N, VEGF-C [100 ng/ml]; or after pretreatment with Slit2N, then VEGF-C. Band intensity of each lane from Figure 6B was determined by densitometry. The ratio of p-Akt/total Akt in control siRNA-transfected L-LECs incubated with VEGF-C alone was set to “1” and all other ratios were determined vs. this control. Data represent the mean ± SD of 3 independent experiments (***p < 0.001).

    Techniques Used: Inhibition, Activity Assay, Enzyme-linked Immunosorbent Assay, Transfection, Incubation, Western Blot

    pi3k activity assay  (Echelon Biosciences)


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    Echelon Biosciences pi3k activity assay
    (A) The cellular content of PIP3 was quantified using a competitive PIP3 ELISA; *p<0.05; **p<0.01 versus 2008-scb control; # p<0.05 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (B) Activity of <t>PI3K</t> immunoprecipitated with antibody to p85 regulatory subunit of class IA PI3K; *p<0.01; **p<0.01 versus 2008-scb control; # p<0.01 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (C) PIP3 levels; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. (D) Activity of PI3K; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. Values are mean ± SEM, n = 3. PI3K activity is expressed as percent of control.
    Pi3k Activity Assay, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pi3k activity assay/product/Echelon Biosciences
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
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    1) Product Images from "Regulation of the Epithelial-Mesenchymal Transition by Claudin-3 and Claudin-4"

    Article Title: Regulation of the Epithelial-Mesenchymal Transition by Claudin-3 and Claudin-4

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0067496

    (A) The cellular content of PIP3 was quantified using a competitive PIP3 ELISA; *p<0.05; **p<0.01 versus 2008-scb control; # p<0.05 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (B) Activity of PI3K immunoprecipitated with antibody to p85 regulatory subunit of class IA PI3K; *p<0.01; **p<0.01 versus 2008-scb control; # p<0.01 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (C) PIP3 levels; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. (D) Activity of PI3K; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. Values are mean ± SEM, n = 3. PI3K activity is expressed as percent of control.
    Figure Legend Snippet: (A) The cellular content of PIP3 was quantified using a competitive PIP3 ELISA; *p<0.05; **p<0.01 versus 2008-scb control; # p<0.05 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (B) Activity of PI3K immunoprecipitated with antibody to p85 regulatory subunit of class IA PI3K; *p<0.01; **p<0.01 versus 2008-scb control; # p<0.01 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (C) PIP3 levels; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. (D) Activity of PI3K; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. Values are mean ± SEM, n = 3. PI3K activity is expressed as percent of control.

    Techniques Used: Enzyme-linked Immunosorbent Assay, Activity Assay, Immunoprecipitation

    pi3k activity elisa kit  (Echelon Biosciences)


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    Echelon Biosciences pi3k activity elisa kit
    RGS20 regulates <t>PI3K/AKT</t> signaling activation in PC cells. ( a) RNA-Seq analysis identified differentially expressed genes in Penl1 cells with or without RGS20 knockdown. (b) Top 5 enriched pathways for downregulated genes ( n = 118) following RGS20 knockdown in Penl1 cells. (c) GSEA analysis revealed that RGS20-high PC cases exhibited highly enriched PI3K/AKT signaling in GSE57955 dataset. (d) Coimmunoprecitation analysis on RGS20 interaction with PI3K p85 α subunit in PC cells. IgG was used as coimmunoprecitation control. (e) PI3K activity of RGS20-depleted 149RM and Penl1 cells. The PI3K activity in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (f) PI3K activity of 149RCa and LM156 cells with or without the RGS20 overexpression. The PI3K activity in EV control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (g) PI3K activity of RGS20-depleted Penl1 cells after EGF stimulation. After serum deprivation for 36 hours, Penl1 cells (Scr or shRGS20) were stimulated with EGF (50 ng/mL) for 30 min, and then PI3K activity was analyzed. n = 4, ∗∗∗ P < 0.001. (h) Western blotting analysis on PI3K/AKT signaling proteins following RGS20 depletion in 149RM and Penl1 cells. (i) Western blotting analysis on PI3K/AKT signaling proteins following the overexpression of RGS20 in 149RCa and LM156 cells. (j) Western blotting analysis on PI3K/AKT signaling proteins in RGS20-depleted Penl1 cells after EGF stimulation. After serum deprivation for 36 hours, Penl1 cells (Scr or shRGS20) were stimulated with EGF (50 ng/mL) for 30 min. Western blotting was conducted to analyze PI3K/AKT signaling proteins.
    Pi3k Activity Elisa Kit, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pi3k activity elisa kit/product/Echelon Biosciences
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pi3k activity elisa kit - by Bioz Stars, 2023-01
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    1) Product Images from "RGS20 Promotes Tumor Progression through Modulating PI3K/AKT Signaling Activation in Penile Cancer"

    Article Title: RGS20 Promotes Tumor Progression through Modulating PI3K/AKT Signaling Activation in Penile Cancer

    Journal: Journal of Oncology

    doi: 10.1155/2022/1293622

    RGS20 regulates PI3K/AKT signaling activation in PC cells. ( a) RNA-Seq analysis identified differentially expressed genes in Penl1 cells with or without RGS20 knockdown. (b) Top 5 enriched pathways for downregulated genes ( n = 118) following RGS20 knockdown in Penl1 cells. (c) GSEA analysis revealed that RGS20-high PC cases exhibited highly enriched PI3K/AKT signaling in GSE57955 dataset. (d) Coimmunoprecitation analysis on RGS20 interaction with PI3K p85 α subunit in PC cells. IgG was used as coimmunoprecitation control. (e) PI3K activity of RGS20-depleted 149RM and Penl1 cells. The PI3K activity in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (f) PI3K activity of 149RCa and LM156 cells with or without the RGS20 overexpression. The PI3K activity in EV control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (g) PI3K activity of RGS20-depleted Penl1 cells after EGF stimulation. After serum deprivation for 36 hours, Penl1 cells (Scr or shRGS20) were stimulated with EGF (50 ng/mL) for 30 min, and then PI3K activity was analyzed. n = 4, ∗∗∗ P < 0.001. (h) Western blotting analysis on PI3K/AKT signaling proteins following RGS20 depletion in 149RM and Penl1 cells. (i) Western blotting analysis on PI3K/AKT signaling proteins following the overexpression of RGS20 in 149RCa and LM156 cells. (j) Western blotting analysis on PI3K/AKT signaling proteins in RGS20-depleted Penl1 cells after EGF stimulation. After serum deprivation for 36 hours, Penl1 cells (Scr or shRGS20) were stimulated with EGF (50 ng/mL) for 30 min. Western blotting was conducted to analyze PI3K/AKT signaling proteins.
    Figure Legend Snippet: RGS20 regulates PI3K/AKT signaling activation in PC cells. ( a) RNA-Seq analysis identified differentially expressed genes in Penl1 cells with or without RGS20 knockdown. (b) Top 5 enriched pathways for downregulated genes ( n = 118) following RGS20 knockdown in Penl1 cells. (c) GSEA analysis revealed that RGS20-high PC cases exhibited highly enriched PI3K/AKT signaling in GSE57955 dataset. (d) Coimmunoprecitation analysis on RGS20 interaction with PI3K p85 α subunit in PC cells. IgG was used as coimmunoprecitation control. (e) PI3K activity of RGS20-depleted 149RM and Penl1 cells. The PI3K activity in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (f) PI3K activity of 149RCa and LM156 cells with or without the RGS20 overexpression. The PI3K activity in EV control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (g) PI3K activity of RGS20-depleted Penl1 cells after EGF stimulation. After serum deprivation for 36 hours, Penl1 cells (Scr or shRGS20) were stimulated with EGF (50 ng/mL) for 30 min, and then PI3K activity was analyzed. n = 4, ∗∗∗ P < 0.001. (h) Western blotting analysis on PI3K/AKT signaling proteins following RGS20 depletion in 149RM and Penl1 cells. (i) Western blotting analysis on PI3K/AKT signaling proteins following the overexpression of RGS20 in 149RCa and LM156 cells. (j) Western blotting analysis on PI3K/AKT signaling proteins in RGS20-depleted Penl1 cells after EGF stimulation. After serum deprivation for 36 hours, Penl1 cells (Scr or shRGS20) were stimulated with EGF (50 ng/mL) for 30 min. Western blotting was conducted to analyze PI3K/AKT signaling proteins.

    Techniques Used: Activation Assay, RNA Sequencing Assay, Activity Assay, Over Expression, Western Blot

    Knockdown of PI3K p85 α or p110 α suppresses cell proliferation, soft agar clonogenesis, and migration/invasion in PC cell lines. (a) Western blotting on PI3K p85 α or p110 α expression in 149RM and Penl1 cells transfected with scramble (Scr) control or shRNAs targeting p85 α or p110 α . β -Actin was used as loading control. (b) CCK-8 analysis on cell viability after p85 α or p110 α knockdown in 149RM and Penl1 cells. The cell viability in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (c) BrdU incorporation analysis on cell proliferation following p85 α or p110 α knockdown in 149RM and Penl1 cells. The BrdU incorporation in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (d) Soft agar clonogenesis of PC cells following p85 α or p110 α knockdown. The soft agar clonogenesis in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. Bars: 100 μ m. (e) Caspase-3 activity of PC cells following p85 α or p110 α knockdown. The caspase-3 activity in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (f) Wound healing assay on PC cells following p85 α or p110 α knockdown. Micrographs showed the results of Penl1 cells. Bars: 100 μ m. The cell migration in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (g) Transwell invasion assay on PC cells following p85 α or p110 α knockdown. Bars: 50 μ m. The cell invasion in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001.
    Figure Legend Snippet: Knockdown of PI3K p85 α or p110 α suppresses cell proliferation, soft agar clonogenesis, and migration/invasion in PC cell lines. (a) Western blotting on PI3K p85 α or p110 α expression in 149RM and Penl1 cells transfected with scramble (Scr) control or shRNAs targeting p85 α or p110 α . β -Actin was used as loading control. (b) CCK-8 analysis on cell viability after p85 α or p110 α knockdown in 149RM and Penl1 cells. The cell viability in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (c) BrdU incorporation analysis on cell proliferation following p85 α or p110 α knockdown in 149RM and Penl1 cells. The BrdU incorporation in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (d) Soft agar clonogenesis of PC cells following p85 α or p110 α knockdown. The soft agar clonogenesis in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. Bars: 100 μ m. (e) Caspase-3 activity of PC cells following p85 α or p110 α knockdown. The caspase-3 activity in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (f) Wound healing assay on PC cells following p85 α or p110 α knockdown. Micrographs showed the results of Penl1 cells. Bars: 100 μ m. The cell migration in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001. (g) Transwell invasion assay on PC cells following p85 α or p110 α knockdown. Bars: 50 μ m. The cell invasion in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001.

    Techniques Used: Migration, Western Blot, Expressing, Transfection, CCK-8 Assay, BrdU Incorporation Assay, Activity Assay, Wound Healing Assay, Transwell Invasion Assay

    The overexpression of constitutively active PI3K p110 α rescued malignant phenotypes impaired by RGS20 depletion. (a) Western blotting analysis on the expression of PI3K/AKT signaling pathway proteins after transfection with empty vector (EV), PI3K p110 α Myr, or PI3K p110 α KD plasmids in RGS20-depleted 149RM and Penl1cells. (b) CCK-8 analysis on cell viability after PI3K p110 α Myr and KD plasmids transfection in RGS20-depleted 149RM and Penl1 cells. The cell viability in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001, as compared with EV. (c) Soft agar assay on clonogenesis after PI3K p110 α Myr and KD plasmids transfection in RGS20-depleted 149RM and Penl1 cells. The soft agar clonogenesis in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001, as compared with EV. (d) Caspase-3 activity after PI3K p110 α Myr and KD plasmid transfection in RGS20-depleted 149RM and Penl1 cells. The caspase-3 activity in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001, as compared with EV. (e) Wound healing assay on PC cells following PI3K p110 α Myr and KD plasmid transfection in RGS20-depleted 149RM and Penl1 cells. Bars: 100 μ m. The cell migration in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001, as compared with EV. (f) Transwell invasion assay on PC cells following PI3K p110 α Myr and KD plasmid transfection in RGS20-depleted 149RM and Penl1 cells. Bars: 50 μ m. The cell invasion in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001, as compared with EV.
    Figure Legend Snippet: The overexpression of constitutively active PI3K p110 α rescued malignant phenotypes impaired by RGS20 depletion. (a) Western blotting analysis on the expression of PI3K/AKT signaling pathway proteins after transfection with empty vector (EV), PI3K p110 α Myr, or PI3K p110 α KD plasmids in RGS20-depleted 149RM and Penl1cells. (b) CCK-8 analysis on cell viability after PI3K p110 α Myr and KD plasmids transfection in RGS20-depleted 149RM and Penl1 cells. The cell viability in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001, as compared with EV. (c) Soft agar assay on clonogenesis after PI3K p110 α Myr and KD plasmids transfection in RGS20-depleted 149RM and Penl1 cells. The soft agar clonogenesis in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001, as compared with EV. (d) Caspase-3 activity after PI3K p110 α Myr and KD plasmid transfection in RGS20-depleted 149RM and Penl1 cells. The caspase-3 activity in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001, as compared with EV. (e) Wound healing assay on PC cells following PI3K p110 α Myr and KD plasmid transfection in RGS20-depleted 149RM and Penl1 cells. Bars: 100 μ m. The cell migration in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001, as compared with EV. (f) Transwell invasion assay on PC cells following PI3K p110 α Myr and KD plasmid transfection in RGS20-depleted 149RM and Penl1 cells. Bars: 50 μ m. The cell invasion in Scr control was regards as 100%. n = 4, ∗∗∗ P < 0.001, as compared with EV.

    Techniques Used: Over Expression, Western Blot, Expressing, Transfection, Plasmid Preparation, CCK-8 Assay, Soft Agar Assay, Activity Assay, Wound Healing Assay, Migration, Transwell Invasion Assay

    RGS20 knockdown suppressed in vivo tumor growth and disrupted PI3K/AKT signaling activation in the Penl1 xenograft model . ( a). Nude mice xenograft study showed that RGS20 depletion attenuated subcutaneous tumor growth in nude mice. Tumor volume was measured every two days after Penl1 inoculation. n = 6, ∗ P < 0.05, shRGS20 vs. Scr control. (b) Tumor weight of Penl1 xenografts with or without RGS20 knockdown. ∗ P < 0.001. (c) Western blotting analysis on protein lysates extracted from Penl1 xenografts with or without RGS20 depletion ( n = 3). (d) IHC staining on Penl1 xenografts with or without RGS20 depletion at day 26 after inoculation. The tissue sections were incubated with antibodies against indicated antibodies (RGS20, p-AKT (T308), p-AKT (S473), Ki-67, and cleaved caspase-3). Bar, 50 μ m.
    Figure Legend Snippet: RGS20 knockdown suppressed in vivo tumor growth and disrupted PI3K/AKT signaling activation in the Penl1 xenograft model . ( a). Nude mice xenograft study showed that RGS20 depletion attenuated subcutaneous tumor growth in nude mice. Tumor volume was measured every two days after Penl1 inoculation. n = 6, ∗ P < 0.05, shRGS20 vs. Scr control. (b) Tumor weight of Penl1 xenografts with or without RGS20 knockdown. ∗ P < 0.001. (c) Western blotting analysis on protein lysates extracted from Penl1 xenografts with or without RGS20 depletion ( n = 3). (d) IHC staining on Penl1 xenografts with or without RGS20 depletion at day 26 after inoculation. The tissue sections were incubated with antibodies against indicated antibodies (RGS20, p-AKT (T308), p-AKT (S473), Ki-67, and cleaved caspase-3). Bar, 50 μ m.

    Techniques Used: In Vivo, Activation Assay, Western Blot, Immunohistochemistry, Incubation

    The high RGS20 expression is associated with PI3K/AKT signaling activation and unfavorable patient survival in PC. (a) IHC micrographs showed consistent expression of RGS20 and p-AKT (T308, S473) in RGS20-high or RGS20-low PCs. Bars: 100 μ m. (b) The RGS20 expression was significantly correlated with p-AKT (T308) and p-AKT (S473) in our PC cohort ( n = 94). (c, d) Log-rank survival analysis showed that the high RGS20 expression was associated with progression-free survival and overall survival in our PC cohort ( n = 94).
    Figure Legend Snippet: The high RGS20 expression is associated with PI3K/AKT signaling activation and unfavorable patient survival in PC. (a) IHC micrographs showed consistent expression of RGS20 and p-AKT (T308, S473) in RGS20-high or RGS20-low PCs. Bars: 100 μ m. (b) The RGS20 expression was significantly correlated with p-AKT (T308) and p-AKT (S473) in our PC cohort ( n = 94). (c, d) Log-rank survival analysis showed that the high RGS20 expression was associated with progression-free survival and overall survival in our PC cohort ( n = 94).

    Techniques Used: Expressing, Activation Assay

    pi3k activity  (Echelon Biosciences)


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    Echelon Biosciences pi3k activity
    A–B), S9 depresses EGF-triggered activation of <t>PI3K-Akt-mTOR</t> signaling pathway. Serum-deprived Rh30 cells (A) and SK-OV-3 cells (B) were treated with indicated concentrations of S9 for 1 h followed by EGF (50 ng/ml) stimulation for 10 min. Cells were harvested for Western blot analysis with antibodies specific for p-PDK1 (S241), PDK, p-Akt (T308), p-Akt (S473), Akt, p-mTOR (Ser2448), mTOR, p-p70S6K (T389), p70S6K, p-4E-BP1(T37/64), p-4E-BP1 (T70), 4E-BP1 and actin. Arrows indicate p70 isoform of S6 kinase protein. C) S9 blocks Akt membrane translocation and membrane ruffling. CHO (pCORON1000-EGFP-Akt) cells seeded on chamber were starved for 2 h then treated with 5 or 10 µM S9 for 1 h followed by IGF stimulation for 5 min. Fluorescent pictures were captured with confocal fluorescent microscopy. White arrows indicate cell membrane ruffling. Blue arrows indicate fluorescent foci. D) Total Akt granule intensity in each CHO (pCORON1000-EGFP-Akt) cell was counted with statistic module by IN Cell Analyzer 1000. Data shown are representative from two independent experiments.
    Pi3k Activity, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "S9, a Novel Anticancer Agent, Exerts Its Anti-Proliferative Activity by Interfering with Both PI3K-Akt-mTOR Signaling and Microtubule Cytoskeleton"

    Article Title: S9, a Novel Anticancer Agent, Exerts Its Anti-Proliferative Activity by Interfering with Both PI3K-Akt-mTOR Signaling and Microtubule Cytoskeleton

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0004881

    A–B), S9 depresses EGF-triggered activation of PI3K-Akt-mTOR signaling pathway. Serum-deprived Rh30 cells (A) and SK-OV-3 cells (B) were treated with indicated concentrations of S9 for 1 h followed by EGF (50 ng/ml) stimulation for 10 min. Cells were harvested for Western blot analysis with antibodies specific for p-PDK1 (S241), PDK, p-Akt (T308), p-Akt (S473), Akt, p-mTOR (Ser2448), mTOR, p-p70S6K (T389), p70S6K, p-4E-BP1(T37/64), p-4E-BP1 (T70), 4E-BP1 and actin. Arrows indicate p70 isoform of S6 kinase protein. C) S9 blocks Akt membrane translocation and membrane ruffling. CHO (pCORON1000-EGFP-Akt) cells seeded on chamber were starved for 2 h then treated with 5 or 10 µM S9 for 1 h followed by IGF stimulation for 5 min. Fluorescent pictures were captured with confocal fluorescent microscopy. White arrows indicate cell membrane ruffling. Blue arrows indicate fluorescent foci. D) Total Akt granule intensity in each CHO (pCORON1000-EGFP-Akt) cell was counted with statistic module by IN Cell Analyzer 1000. Data shown are representative from two independent experiments.
    Figure Legend Snippet: A–B), S9 depresses EGF-triggered activation of PI3K-Akt-mTOR signaling pathway. Serum-deprived Rh30 cells (A) and SK-OV-3 cells (B) were treated with indicated concentrations of S9 for 1 h followed by EGF (50 ng/ml) stimulation for 10 min. Cells were harvested for Western blot analysis with antibodies specific for p-PDK1 (S241), PDK, p-Akt (T308), p-Akt (S473), Akt, p-mTOR (Ser2448), mTOR, p-p70S6K (T389), p70S6K, p-4E-BP1(T37/64), p-4E-BP1 (T70), 4E-BP1 and actin. Arrows indicate p70 isoform of S6 kinase protein. C) S9 blocks Akt membrane translocation and membrane ruffling. CHO (pCORON1000-EGFP-Akt) cells seeded on chamber were starved for 2 h then treated with 5 or 10 µM S9 for 1 h followed by IGF stimulation for 5 min. Fluorescent pictures were captured with confocal fluorescent microscopy. White arrows indicate cell membrane ruffling. Blue arrows indicate fluorescent foci. D) Total Akt granule intensity in each CHO (pCORON1000-EGFP-Akt) cell was counted with statistic module by IN Cell Analyzer 1000. Data shown are representative from two independent experiments.

    Techniques Used: Activation Assay, Western Blot, Translocation Assay, Microscopy

    A–B) Active kinases were immunoprecipitated from exponentially growing Rh30 cells and kinase assays were performed. In the presence of indicated concentrations of S9, phosphorylation PIP2 to PIP3 were measured by ELISA (A). Phosphorylation of 4E-BP1 (B) was detected by Western blot. Band intensity was quantitated by optical densitometric analysis and normalized to vehicle control. Representative images were presented and relative kinase activity was plotted as mean±SD of three independent experiments. C) S9 abrogates rapamycin-induced hyperphosphorylation of Akt (Ser 473). Rh30 cells were starved overnight and treated with 10 µM of S9 and/or rapamycin (Rapa) for 1.5 h on the next day, followed by stimulation with 50 ng/ml EGF for 10 min. Phosphorylated Akt at Serine 473 and Threonine 308 and total Akt were detected by Western blot. D) Rh30 cells were exposed to 10 µM camptothecin (CPT) for 1 h after pre-incubation with indicated concentrations of S9 or wortmannin (Wort) for 30 min. Cells were collected and γ-H2AX levels were detected with Western blot analysis. E–F) The binding mode of S9 within PI3K (E), mTOR (F). The protein is represented by cartoon. The residues interacting with the compounds are shown in sticks. All of the structural diagrams were prepared using PyMOL ( http://pymol.sourceforge.net ).
    Figure Legend Snippet: A–B) Active kinases were immunoprecipitated from exponentially growing Rh30 cells and kinase assays were performed. In the presence of indicated concentrations of S9, phosphorylation PIP2 to PIP3 were measured by ELISA (A). Phosphorylation of 4E-BP1 (B) was detected by Western blot. Band intensity was quantitated by optical densitometric analysis and normalized to vehicle control. Representative images were presented and relative kinase activity was plotted as mean±SD of three independent experiments. C) S9 abrogates rapamycin-induced hyperphosphorylation of Akt (Ser 473). Rh30 cells were starved overnight and treated with 10 µM of S9 and/or rapamycin (Rapa) for 1.5 h on the next day, followed by stimulation with 50 ng/ml EGF for 10 min. Phosphorylated Akt at Serine 473 and Threonine 308 and total Akt were detected by Western blot. D) Rh30 cells were exposed to 10 µM camptothecin (CPT) for 1 h after pre-incubation with indicated concentrations of S9 or wortmannin (Wort) for 30 min. Cells were collected and γ-H2AX levels were detected with Western blot analysis. E–F) The binding mode of S9 within PI3K (E), mTOR (F). The protein is represented by cartoon. The residues interacting with the compounds are shown in sticks. All of the structural diagrams were prepared using PyMOL ( http://pymol.sourceforge.net ).

    Techniques Used: Immunoprecipitation, Enzyme-linked Immunosorbent Assay, Western Blot, Activity Assay, Incubation, Binding Assay

    pi3k activity elisa kit  (Echelon Biosciences)


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    Echelon Biosciences pi3k activity elisa kit
    In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase <t>(PI3K)</t> inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Pi3k Activity Elisa Kit, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells"

    Article Title: Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0093576

    In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: In Vitro, Invasion Assay, shRNA, Positive Control, Fluorescence

    MMP-2 mRNA levels were determined by real-time RT-PCR in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 mRNA level was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: MMP-2 mRNA levels were determined by real-time RT-PCR in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 mRNA level was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Quantitative RT-PCR, shRNA, Positive Control

    MMP-2 protein levels were determined by Western blot analyses in in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the MMP-2 blot was normalized against that of GAPDH to obtain a relative blot density, which was expressed as fold changes to the relative MMP-2 blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: MMP-2 protein levels were determined by Western blot analyses in in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the MMP-2 blot was normalized against that of GAPDH to obtain a relative blot density, which was expressed as fold changes to the relative MMP-2 blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Western Blot, Stable Transfection, shRNA

    MMP-2 activities were determined with a SensoLyte 520 MMP-2 Assay Kit (AnaSpec) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: MMP-2 activities were determined with a SensoLyte 520 MMP-2 Assay Kit (AnaSpec) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: shRNA, Positive Control, Activity Assay

    Methlythiazoletetrazolium (MTT) cell viability assays were performed in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 24 or 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Viability of the control cells was designated as 100%. The inhibition rate of cell viability was calculated and shown as a percentage of the control cell viability. Each experiment was repeated for three times in triplicates. Data values were expressed as Mean+SD.
    Figure Legend Snippet: Methlythiazoletetrazolium (MTT) cell viability assays were performed in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 24 or 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Viability of the control cells was designated as 100%. The inhibition rate of cell viability was calculated and shown as a percentage of the control cell viability. Each experiment was repeated for three times in triplicates. Data values were expressed as Mean+SD.

    Techniques Used: shRNA, Positive Control, Inhibition

    PI3K activities were determined with a PI3K Activity ELISA kit (Echelon Biosciences) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The PI3K activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: PI3K activities were determined with a PI3K Activity ELISA kit (Echelon Biosciences) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The PI3K activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Activity Assay, Enzyme-linked Immunosorbent Assay, shRNA, Positive Control

    Levels of total Akt and P-Akt at serine 473 (ser473) were determined by Western blot analyses in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Density of the P-Akt (ser473) blot was normalized against that of total Akt to obtain a relative blot density, which was expressed as fold changes to the relative P-Akt (ser473) blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: Levels of total Akt and P-Akt at serine 473 (ser473) were determined by Western blot analyses in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Density of the P-Akt (ser473) blot was normalized against that of total Akt to obtain a relative blot density, which was expressed as fold changes to the relative P-Akt (ser473) blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Western Blot, Stable Transfection, shRNA

    pi3k activity elisa kit k 1000s  (Echelon Biosciences)


    Bioz Verified Symbol Echelon Biosciences is a verified supplier
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    Echelon Biosciences pi3k activity elisa kit k 1000s
    In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase <t>(PI3K)</t> inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Pi3k Activity Elisa Kit K 1000s, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pi3k activity elisa kit k 1000s/product/Echelon Biosciences
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pi3k activity elisa kit k 1000s - by Bioz Stars, 2023-01
    95/100 stars

    Images

    1) Product Images from "Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells"

    Article Title: Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0093576

    In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: In Vitro, Invasion Assay, shRNA, Positive Control, Fluorescence

    MMP-2 mRNA levels were determined by real-time RT-PCR in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 mRNA level was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: MMP-2 mRNA levels were determined by real-time RT-PCR in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 mRNA level was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Quantitative RT-PCR, shRNA, Positive Control

    MMP-2 protein levels were determined by Western blot analyses in in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the MMP-2 blot was normalized against that of GAPDH to obtain a relative blot density, which was expressed as fold changes to the relative MMP-2 blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: MMP-2 protein levels were determined by Western blot analyses in in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the MMP-2 blot was normalized against that of GAPDH to obtain a relative blot density, which was expressed as fold changes to the relative MMP-2 blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Western Blot, Stable Transfection, shRNA

    MMP-2 activities were determined with a SensoLyte 520 MMP-2 Assay Kit (AnaSpec) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: MMP-2 activities were determined with a SensoLyte 520 MMP-2 Assay Kit (AnaSpec) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: shRNA, Positive Control, Activity Assay

    Methlythiazoletetrazolium (MTT) cell viability assays were performed in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 24 or 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Viability of the control cells was designated as 100%. The inhibition rate of cell viability was calculated and shown as a percentage of the control cell viability. Each experiment was repeated for three times in triplicates. Data values were expressed as Mean+SD.
    Figure Legend Snippet: Methlythiazoletetrazolium (MTT) cell viability assays were performed in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 24 or 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Viability of the control cells was designated as 100%. The inhibition rate of cell viability was calculated and shown as a percentage of the control cell viability. Each experiment was repeated for three times in triplicates. Data values were expressed as Mean+SD.

    Techniques Used: shRNA, Positive Control, Inhibition

    PI3K activities were determined with a PI3K Activity ELISA kit (Echelon Biosciences) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The PI3K activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: PI3K activities were determined with a PI3K Activity ELISA kit (Echelon Biosciences) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The PI3K activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Activity Assay, Enzyme-linked Immunosorbent Assay, shRNA, Positive Control

    Levels of total Akt and P-Akt at serine 473 (ser473) were determined by Western blot analyses in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Density of the P-Akt (ser473) blot was normalized against that of total Akt to obtain a relative blot density, which was expressed as fold changes to the relative P-Akt (ser473) blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: Levels of total Akt and P-Akt at serine 473 (ser473) were determined by Western blot analyses in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Density of the P-Akt (ser473) blot was normalized against that of total Akt to obtain a relative blot density, which was expressed as fold changes to the relative P-Akt (ser473) blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Western Blot, Stable Transfection, shRNA

    pi3k activity  (Echelon Biosciences)


    Bioz Verified Symbol Echelon Biosciences is a verified supplier
    Bioz Manufacturer Symbol Echelon Biosciences manufactures this product  
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  • 94

    Structured Review

    Echelon Biosciences pi3k activity
    In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase <t>(PI3K)</t> inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Pi3k Activity, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pi3k activity/product/Echelon Biosciences
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pi3k activity - by Bioz Stars, 2023-01
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    1) Product Images from "Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells"

    Article Title: Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0093576

    In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: In Vitro, Invasion Assay, shRNA, Positive Control, Fluorescence

    MMP-2 mRNA levels were determined by real-time RT-PCR in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 mRNA level was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: MMP-2 mRNA levels were determined by real-time RT-PCR in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 mRNA level was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Quantitative RT-PCR, shRNA, Positive Control

    MMP-2 protein levels were determined by Western blot analyses in in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the MMP-2 blot was normalized against that of GAPDH to obtain a relative blot density, which was expressed as fold changes to the relative MMP-2 blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: MMP-2 protein levels were determined by Western blot analyses in in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the MMP-2 blot was normalized against that of GAPDH to obtain a relative blot density, which was expressed as fold changes to the relative MMP-2 blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Western Blot, Stable Transfection, shRNA

    MMP-2 activities were determined with a SensoLyte 520 MMP-2 Assay Kit (AnaSpec) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: MMP-2 activities were determined with a SensoLyte 520 MMP-2 Assay Kit (AnaSpec) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: shRNA, Positive Control, Activity Assay

    Methlythiazoletetrazolium (MTT) cell viability assays were performed in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 24 or 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Viability of the control cells was designated as 100%. The inhibition rate of cell viability was calculated and shown as a percentage of the control cell viability. Each experiment was repeated for three times in triplicates. Data values were expressed as Mean+SD.
    Figure Legend Snippet: Methlythiazoletetrazolium (MTT) cell viability assays were performed in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 24 or 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Viability of the control cells was designated as 100%. The inhibition rate of cell viability was calculated and shown as a percentage of the control cell viability. Each experiment was repeated for three times in triplicates. Data values were expressed as Mean+SD.

    Techniques Used: shRNA, Positive Control, Inhibition

    PI3K activities were determined with a PI3K Activity ELISA kit (Echelon Biosciences) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The PI3K activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: PI3K activities were determined with a PI3K Activity ELISA kit (Echelon Biosciences) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The PI3K activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Activity Assay, Enzyme-linked Immunosorbent Assay, shRNA, Positive Control

    Levels of total Akt and P-Akt at serine 473 (ser473) were determined by Western blot analyses in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Density of the P-Akt (ser473) blot was normalized against that of total Akt to obtain a relative blot density, which was expressed as fold changes to the relative P-Akt (ser473) blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Figure Legend Snippet: Levels of total Akt and P-Akt at serine 473 (ser473) were determined by Western blot analyses in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Density of the P-Akt (ser473) blot was normalized against that of total Akt to obtain a relative blot density, which was expressed as fold changes to the relative P-Akt (ser473) blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Techniques Used: Western Blot, Stable Transfection, shRNA

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    Echelon Biosciences pi3k activity
    Effects of CD26 on signalling pathway of keloid fibroblasts (KFs) and screening of the related pathways. (a) The relationship between phosphatidylinositol 3-kinase <t>(PI3K)</t> activity and CD26 expression was in CD26 + KFs and CD 26 − KFs tested using a PI3K activity detection kit. (b) Protein kinase B ( AKT) phosphorylation levels in CD26 + KFs and CD26 − KFs were detected and the samples were imaged under a fluorescent microscope; bar = 100 μm. (c) Cells were subjected to western blotting to determine the effects of CD26 expression on mammalian target of rapamycin and downstream molecular phosphorylation. (d) The expression of keloid-related cytokines in CD26 −/+ KFs was detected by ELISA assay. (e) CD26 −/+ KFs were cultured for 24 hours in the presence of insulin-like growth factor-1 receptor inhibitor AZD3463, transforming growth factor-β1 receptor inhibitor SB505124, vascular endothelial growth factor receptor inhibitor Axitinib, platelet-derived growth factor receptor inhibitor Crenolanib (CP-868596), interleukin-1β (IL-1β) inhibitor AS101, interleukin-6 receptor antagonist Tocilizumab and tumor necrosis factor-α inhibitor Bay 11-7085, respectively. The culture medium was then changed every 24 hours. Cell proliferation differences were analysed by cell counting kit-8 assay at 96 hours after seeding. (f) The invasive abilities of CD26 −/+ KFs were detected by transwell assay; bar = 100 μm. Data are expressed as the mean ± standard error of the mean. * p < 0.05, ** p < 0.01, *** p < 0.001. mTOR mammalian target of rapamycin, DAPI 4′,6-diamidino-2-phenylindole, OD optical density
    Pi3k Activity, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "CD26 upregulates proliferation and invasion in keloid fibroblasts through an IGF-1-induced PI3K/AKT/mTOR pathway"

    Article Title: CD26 upregulates proliferation and invasion in keloid fibroblasts through an IGF-1-induced PI3K/AKT/mTOR pathway

    Journal: Burns & Trauma

    doi: 10.1093/burnst/tkaa025

    Effects of CD26 on signalling pathway of keloid fibroblasts (KFs) and screening of the related pathways. (a) The relationship between phosphatidylinositol 3-kinase (PI3K) activity and CD26 expression was in CD26 + KFs and CD 26 − KFs tested using a PI3K activity detection kit. (b) Protein kinase B ( AKT) phosphorylation levels in CD26 + KFs and CD26 − KFs were detected and the samples were imaged under a fluorescent microscope; bar = 100 μm. (c) Cells were subjected to western blotting to determine the effects of CD26 expression on mammalian target of rapamycin and downstream molecular phosphorylation. (d) The expression of keloid-related cytokines in CD26 −/+ KFs was detected by ELISA assay. (e) CD26 −/+ KFs were cultured for 24 hours in the presence of insulin-like growth factor-1 receptor inhibitor AZD3463, transforming growth factor-β1 receptor inhibitor SB505124, vascular endothelial growth factor receptor inhibitor Axitinib, platelet-derived growth factor receptor inhibitor Crenolanib (CP-868596), interleukin-1β (IL-1β) inhibitor AS101, interleukin-6 receptor antagonist Tocilizumab and tumor necrosis factor-α inhibitor Bay 11-7085, respectively. The culture medium was then changed every 24 hours. Cell proliferation differences were analysed by cell counting kit-8 assay at 96 hours after seeding. (f) The invasive abilities of CD26 −/+ KFs were detected by transwell assay; bar = 100 μm. Data are expressed as the mean ± standard error of the mean. * p < 0.05, ** p < 0.01, *** p < 0.001. mTOR mammalian target of rapamycin, DAPI 4′,6-diamidino-2-phenylindole, OD optical density
    Figure Legend Snippet: Effects of CD26 on signalling pathway of keloid fibroblasts (KFs) and screening of the related pathways. (a) The relationship between phosphatidylinositol 3-kinase (PI3K) activity and CD26 expression was in CD26 + KFs and CD 26 − KFs tested using a PI3K activity detection kit. (b) Protein kinase B ( AKT) phosphorylation levels in CD26 + KFs and CD26 − KFs were detected and the samples were imaged under a fluorescent microscope; bar = 100 μm. (c) Cells were subjected to western blotting to determine the effects of CD26 expression on mammalian target of rapamycin and downstream molecular phosphorylation. (d) The expression of keloid-related cytokines in CD26 −/+ KFs was detected by ELISA assay. (e) CD26 −/+ KFs were cultured for 24 hours in the presence of insulin-like growth factor-1 receptor inhibitor AZD3463, transforming growth factor-β1 receptor inhibitor SB505124, vascular endothelial growth factor receptor inhibitor Axitinib, platelet-derived growth factor receptor inhibitor Crenolanib (CP-868596), interleukin-1β (IL-1β) inhibitor AS101, interleukin-6 receptor antagonist Tocilizumab and tumor necrosis factor-α inhibitor Bay 11-7085, respectively. The culture medium was then changed every 24 hours. Cell proliferation differences were analysed by cell counting kit-8 assay at 96 hours after seeding. (f) The invasive abilities of CD26 −/+ KFs were detected by transwell assay; bar = 100 μm. Data are expressed as the mean ± standard error of the mean. * p < 0.05, ** p < 0.01, *** p < 0.001. mTOR mammalian target of rapamycin, DAPI 4′,6-diamidino-2-phenylindole, OD optical density

    Techniques Used: Activity Assay, Expressing, Microscopy, Western Blot, Enzyme-linked Immunosorbent Assay, Cell Culture, Derivative Assay, Cell Counting, Transwell Assay

    CD26 regulates the proliferation and invasion-related proteins of keloid fibroblasts (KFs) by insulin-like growth factor-1 receptor (IGF-1R). Verification of IGF-1R silencing in CD26 +/− KFs using lentiviral-mediated transduction of shRNA specific for IGF-1R by RT-qPCR (a) and western blotting (b) . (c) Cell proliferative abilities were measured with cell counting kit-8 assay in CD26 +/− KFs and IGF-1R-silenced CD26 +/− KFs—the difference between CD26 + KFs and CD26 − KFs disappeared after IGF-1R was knocked down. (d) The EdU-positive rate was counted to evaluate cell growth activity: the difference between CD26 + KFs and CD26 − KFs was not observed after IGF-1R was knocked down. (e) The changes in CD26 +/− KF migration after IGF-1R were knocked down were observed 24 hours after scratching and the percentage of the migrated area was measured. (f) The effects of IGF-1R knock down on CD26 +/− KFs invasive abilities were observed. After IGF-1R was knocked down, (g) the phosphatidylinositol 3-kinase (PI3K) activities in CD26 +/− KFs were detected. (h) The phosphorylation levels of PI3K were determined by western blotting. Data are expressed as the mean ± standard error of the mean. * p < 0.05, ** p < 0.01. CD26 − /sh-IGF1R stably silenced IGF-1Rs in CD26 − KFs, CD26 + /sh-IGF1R stably silenced IGF-1Rs in CD26 + KFs, CD26 + /sh-control , scrambled shRNA control group of CD26 + KFs, P13K phosphatidylinositol 3-kinase, OD optical density, ns no statistical difference
    Figure Legend Snippet: CD26 regulates the proliferation and invasion-related proteins of keloid fibroblasts (KFs) by insulin-like growth factor-1 receptor (IGF-1R). Verification of IGF-1R silencing in CD26 +/− KFs using lentiviral-mediated transduction of shRNA specific for IGF-1R by RT-qPCR (a) and western blotting (b) . (c) Cell proliferative abilities were measured with cell counting kit-8 assay in CD26 +/− KFs and IGF-1R-silenced CD26 +/− KFs—the difference between CD26 + KFs and CD26 − KFs disappeared after IGF-1R was knocked down. (d) The EdU-positive rate was counted to evaluate cell growth activity: the difference between CD26 + KFs and CD26 − KFs was not observed after IGF-1R was knocked down. (e) The changes in CD26 +/− KF migration after IGF-1R were knocked down were observed 24 hours after scratching and the percentage of the migrated area was measured. (f) The effects of IGF-1R knock down on CD26 +/− KFs invasive abilities were observed. After IGF-1R was knocked down, (g) the phosphatidylinositol 3-kinase (PI3K) activities in CD26 +/− KFs were detected. (h) The phosphorylation levels of PI3K were determined by western blotting. Data are expressed as the mean ± standard error of the mean. * p < 0.05, ** p < 0.01. CD26 − /sh-IGF1R stably silenced IGF-1Rs in CD26 − KFs, CD26 + /sh-IGF1R stably silenced IGF-1Rs in CD26 + KFs, CD26 + /sh-control , scrambled shRNA control group of CD26 + KFs, P13K phosphatidylinositol 3-kinase, OD optical density, ns no statistical difference

    Techniques Used: Transduction, shRNA, Quantitative RT-PCR, Western Blot, Cell Counting, Activity Assay, Migration, Stable Transfection

    CD26 regulates the proliferative and invasive behaviours of keloid fibroblasts (KFs) by phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) activation in the insulin growth factor-1 (IGF-1)/IGF-1 receptor-mediated signalling pathway. The cell proliferation of CD26 +/− KFs with or without PI3K inhibitor LY294002 (10 µM) treated for 24 hours, were then measured with cell counting kit-8 assay (a) , as well as EdU staining, the ratio of EdU-positive cells to Hoechst-labelled cells in each group was determined (b) . (c) The effect of PI3K on cell migration of CD26 +/− KFs was evaluated after treatment with LY294002 (10 µM) for 24 h. (d) The changes in the invasive abilities of CD26 +/− KFs after LY294002 treatment for 24 hours was observed. (e) The expression of molecules in the AKT pathway was tested by western blotting after treatment with LY294002. P70S6K: 70kDa ribosomal proteinS6 kinase. Data are expressed as the mean ± standard error of the mean. * p < 0.05, ** p < 0.01, *** p < 0.001. AKT protein kinase B, mTOR mammalian target of rapamycin. OD optical density, ns no statistical difference
    Figure Legend Snippet: CD26 regulates the proliferative and invasive behaviours of keloid fibroblasts (KFs) by phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) activation in the insulin growth factor-1 (IGF-1)/IGF-1 receptor-mediated signalling pathway. The cell proliferation of CD26 +/− KFs with or without PI3K inhibitor LY294002 (10 µM) treated for 24 hours, were then measured with cell counting kit-8 assay (a) , as well as EdU staining, the ratio of EdU-positive cells to Hoechst-labelled cells in each group was determined (b) . (c) The effect of PI3K on cell migration of CD26 +/− KFs was evaluated after treatment with LY294002 (10 µM) for 24 h. (d) The changes in the invasive abilities of CD26 +/− KFs after LY294002 treatment for 24 hours was observed. (e) The expression of molecules in the AKT pathway was tested by western blotting after treatment with LY294002. P70S6K: 70kDa ribosomal proteinS6 kinase. Data are expressed as the mean ± standard error of the mean. * p < 0.05, ** p < 0.01, *** p < 0.001. AKT protein kinase B, mTOR mammalian target of rapamycin. OD optical density, ns no statistical difference

    Techniques Used: Activation Assay, Cell Counting, Staining, Migration, Expressing, Western Blot

    CD26 regulates the proliferative and invasive behaviours of keloid fibroblasts (KFs) by the mammalian target of rapamycin (mTOR) in the insulin-like growth factor-1 (IGF-1)–IGF-1 receptor (IGF-1R)/ phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)-mediated signalling pathway. The role of mTOR in the IGF-1–IGF-1R/PI3K/AKT activation of CD26 + KFs was detected by treatment with mTOR inhibitor KU-0063794 (10 µM) for 24 h. (a) Cell counting kit-8 assay and (b) EdU staining showed that proliferation advantage in CD26 + KFs did not remain after blockage of mTOR. (c) To evaluate the mTOR activation in cell migration regulation, a scratch wound was measured after 24 hours and there was no significant difference between the CD26 + KFs treated with KU-0063794 (CD26 + / KU-0063794) and CD26 − KFs treated with KU-0063794 (CD26 − /KU-0063794). (d) The invasive abilities of CD26 +/− KFs were observed in transwell assay. (e) CD26 regulates the phosphorylation levels of S6 kinase and 4E-binding protein through IGF1R/PI3K/AKT/mTOR. Data are expressed as the mean ± the standard error of the mean. * p < 0.05, ** p < 0.01, *** p < 0.001. S6K S6 kinase, 4EBP1 4E-binding protein, OD optical density, ns no statistical difference
    Figure Legend Snippet: CD26 regulates the proliferative and invasive behaviours of keloid fibroblasts (KFs) by the mammalian target of rapamycin (mTOR) in the insulin-like growth factor-1 (IGF-1)–IGF-1 receptor (IGF-1R)/ phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)-mediated signalling pathway. The role of mTOR in the IGF-1–IGF-1R/PI3K/AKT activation of CD26 + KFs was detected by treatment with mTOR inhibitor KU-0063794 (10 µM) for 24 h. (a) Cell counting kit-8 assay and (b) EdU staining showed that proliferation advantage in CD26 + KFs did not remain after blockage of mTOR. (c) To evaluate the mTOR activation in cell migration regulation, a scratch wound was measured after 24 hours and there was no significant difference between the CD26 + KFs treated with KU-0063794 (CD26 + / KU-0063794) and CD26 − KFs treated with KU-0063794 (CD26 − /KU-0063794). (d) The invasive abilities of CD26 +/− KFs were observed in transwell assay. (e) CD26 regulates the phosphorylation levels of S6 kinase and 4E-binding protein through IGF1R/PI3K/AKT/mTOR. Data are expressed as the mean ± the standard error of the mean. * p < 0.05, ** p < 0.01, *** p < 0.001. S6K S6 kinase, 4EBP1 4E-binding protein, OD optical density, ns no statistical difference

    Techniques Used: Activation Assay, Cell Counting, Staining, Migration, Transwell Assay, Binding Assay

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  • 95
    Echelon Biosciences pi3k activity elisa kit
    Expression and activation of IGF-IR and <t>PI3K</t> in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.
    Pi3k Activity Elisa Kit, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Echelon Biosciences pi3k activity
    Expression and activation of IGF-IR and <t>PI3K</t> in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.
    Pi3k Activity, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pi3k activity/product/Echelon Biosciences
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pi3k activity - by Bioz Stars, 2023-01
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    95
    Echelon Biosciences pi3k activity assay
    (A) The cellular content of PIP3 was quantified using a competitive PIP3 ELISA; *p<0.05; **p<0.01 versus 2008-scb control; # p<0.05 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (B) Activity of <t>PI3K</t> immunoprecipitated with antibody to p85 regulatory subunit of class IA PI3K; *p<0.01; **p<0.01 versus 2008-scb control; # p<0.01 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (C) PIP3 levels; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. (D) Activity of PI3K; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. Values are mean ± SEM, n = 3. PI3K activity is expressed as percent of control.
    Pi3k Activity Assay, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pi3k activity assay/product/Echelon Biosciences
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pi3k activity assay - by Bioz Stars, 2023-01
    95/100 stars
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    95
    Echelon Biosciences pi3k activity elisa kit k 1000s
    In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase <t>(PI3K)</t> inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.
    Pi3k Activity Elisa Kit K 1000s, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pi3k activity elisa kit k 1000s/product/Echelon Biosciences
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pi3k activity elisa kit k 1000s - by Bioz Stars, 2023-01
    95/100 stars
      Buy from Supplier

    Image Search Results


    Expression and activation of IGF-IR and PI3K in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.

    Journal: International Journal of Cell Biology

    Article Title: Rac1 and Stathmin but Not EB1 Are Required for Invasion of Breast Cancer Cells in Response to IGF-I

    doi: 10.1155/2011/615912

    Figure Lengend Snippet: Expression and activation of IGF-IR and PI3K in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.

    Article Snippet: PI3K activity was measured by immunoprecipitation of PI3K p85 with anti-p85 antibody, followed by the kinase reaction of the immunoprecipitates in a PI3K activity ELISA kit (Echelon Bioscience, Salt Lake City, UT), according to the manufacturer's instructions.

    Techniques: Expressing, Activation Assay, Western Blot, Immunoprecipitation, Activity Assay

    Expression and activation of IGF-IR and PI3K in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.

    Journal: International Journal of Cell Biology

    Article Title: Rac1 and Stathmin but Not EB1 Are Required for Invasion of Breast Cancer Cells in Response to IGF-I

    doi: 10.1155/2011/615912

    Figure Lengend Snippet: Expression and activation of IGF-IR and PI3K in MDA-MB-231 and MCF7 cells. (a) After stimulation with IGF-I for 0, 0.5, or 6 h, cells were lysed and processed for immunoblotting with anti-IGF-IR antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (b) After stimulation with IGF-I, cells were immunostained with antibody to phospho-IGF-IR (Tyr1135/1135). Scale Bars 20 μ m. (c) After stimulation with IGF-I, cells were lysed and processed for immunoblotting with anti-PI3K (p85) antibody. Values are given as band intensity relative to that in unstimulated control MDA-MB-231 cells. (d) Cells after stimulation with IGF-I were lysed and immunoprecipitated with anti-PI3K (p85) antibody. PI3K activity was assayed for the immunoprecipitates as described in Materials and Methods. The mean (SD) values of triplicate assays are given as activity relative to that in unstimulated control MDA-MB-231 cells.

    Article Snippet: PI3K activity was measured by immunoprecipitation of PI3K p85 with anti-p85 antibody, followed by the kinase reaction of the immunoprecipitates in a PI3K activity ELISA kit (Echelon Bioscience, Salt Lake City, UT), according to the manufacturer's instructions.

    Techniques: Expressing, Activation Assay, Western Blot, Immunoprecipitation, Activity Assay

    (A) The cellular content of PIP3 was quantified using a competitive PIP3 ELISA; *p<0.05; **p<0.01 versus 2008-scb control; # p<0.05 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (B) Activity of PI3K immunoprecipitated with antibody to p85 regulatory subunit of class IA PI3K; *p<0.01; **p<0.01 versus 2008-scb control; # p<0.01 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (C) PIP3 levels; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. (D) Activity of PI3K; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. Values are mean ± SEM, n = 3. PI3K activity is expressed as percent of control.

    Journal: PLoS ONE

    Article Title: Regulation of the Epithelial-Mesenchymal Transition by Claudin-3 and Claudin-4

    doi: 10.1371/journal.pone.0067496

    Figure Lengend Snippet: (A) The cellular content of PIP3 was quantified using a competitive PIP3 ELISA; *p<0.05; **p<0.01 versus 2008-scb control; # p<0.05 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (B) Activity of PI3K immunoprecipitated with antibody to p85 regulatory subunit of class IA PI3K; *p<0.01; **p<0.01 versus 2008-scb control; # p<0.01 versus CLDN3KD; ## p<0.01 versus CLDN4KD. (C) PIP3 levels; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. (D) Activity of PI3K; *p<0.001 versus CLDN3KD; **p<0.001 versus CLDN4KD. Values are mean ± SEM, n = 3. PI3K activity is expressed as percent of control.

    Article Snippet: PI3K activity assay was performed with an enzyme-linked immunosorbent assay (ELISA) K-1000s-PI3-kinase activity (ELISA:Pico, Echelon Biosciences Inc., Salt Lake City, UT,) according the manufacturer's instructions.

    Techniques: Enzyme-linked Immunosorbent Assay, Activity Assay, Immunoprecipitation

    In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Journal: PLoS ONE

    Article Title: Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells

    doi: 10.1371/journal.pone.0093576

    Figure Lengend Snippet: In vitro cell invasion assays were performed with a Fluorimetric Cell Invasion Assay kit (Chemicon; Millipore) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Cell invasion was determined by fluorescence and shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Article Snippet: The PI3K Activity ELISA kit (K-1000s) was purchased from Echelon Biosciences (Salt Lake City, UT, USA).

    Techniques: In Vitro, Invasion Assay, shRNA, Positive Control, Fluorescence

    MMP-2 mRNA levels were determined by real-time RT-PCR in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 mRNA level was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Journal: PLoS ONE

    Article Title: Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells

    doi: 10.1371/journal.pone.0093576

    Figure Lengend Snippet: MMP-2 mRNA levels were determined by real-time RT-PCR in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 mRNA level was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Article Snippet: The PI3K Activity ELISA kit (K-1000s) was purchased from Echelon Biosciences (Salt Lake City, UT, USA).

    Techniques: Quantitative RT-PCR, shRNA, Positive Control

    MMP-2 protein levels were determined by Western blot analyses in in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the MMP-2 blot was normalized against that of GAPDH to obtain a relative blot density, which was expressed as fold changes to the relative MMP-2 blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Journal: PLoS ONE

    Article Title: Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells

    doi: 10.1371/journal.pone.0093576

    Figure Lengend Snippet: MMP-2 protein levels were determined by Western blot analyses in in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the MMP-2 blot was normalized against that of GAPDH to obtain a relative blot density, which was expressed as fold changes to the relative MMP-2 blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Article Snippet: The PI3K Activity ELISA kit (K-1000s) was purchased from Echelon Biosciences (Salt Lake City, UT, USA).

    Techniques: Western Blot, Stable Transfection, shRNA

    MMP-2 activities were determined with a SensoLyte 520 MMP-2 Assay Kit (AnaSpec) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Journal: PLoS ONE

    Article Title: Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells

    doi: 10.1371/journal.pone.0093576

    Figure Lengend Snippet: MMP-2 activities were determined with a SensoLyte 520 MMP-2 Assay Kit (AnaSpec) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The MMP-2 activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Article Snippet: The PI3K Activity ELISA kit (K-1000s) was purchased from Echelon Biosciences (Salt Lake City, UT, USA).

    Techniques: shRNA, Positive Control, Activity Assay

    Methlythiazoletetrazolium (MTT) cell viability assays were performed in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 24 or 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Viability of the control cells was designated as 100%. The inhibition rate of cell viability was calculated and shown as a percentage of the control cell viability. Each experiment was repeated for three times in triplicates. Data values were expressed as Mean+SD.

    Journal: PLoS ONE

    Article Title: Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells

    doi: 10.1371/journal.pone.0093576

    Figure Lengend Snippet: Methlythiazoletetrazolium (MTT) cell viability assays were performed in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 24 or 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. Viability of the control cells was designated as 100%. The inhibition rate of cell viability was calculated and shown as a percentage of the control cell viability. Each experiment was repeated for three times in triplicates. Data values were expressed as Mean+SD.

    Article Snippet: The PI3K Activity ELISA kit (K-1000s) was purchased from Echelon Biosciences (Salt Lake City, UT, USA).

    Techniques: shRNA, Positive Control, Inhibition

    PI3K activities were determined with a PI3K Activity ELISA kit (Echelon Biosciences) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The PI3K activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Journal: PLoS ONE

    Article Title: Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells

    doi: 10.1371/journal.pone.0093576

    Figure Lengend Snippet: PI3K activities were determined with a PI3K Activity ELISA kit (Echelon Biosciences) in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) OS cells treated with control IgG (PK136 mAb, 50 µg/mL), 14G2a mAb (50 µg/mL), selective ETAR antagonist BQ123 (5 µM), and 14G2a (50 µg/mL)+BQ123 (5 µM) for 48 hours. Cells with knockdown of ETAR (ETAR-shRNA) with or without 14G2a mAb treatment were also tested. Cells treated with selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM) was used as a positive control. The PI3K activity was shown as fold changes to that of the untreated control cells (designated as 1). Each experiment was repeated for three times in duplicates. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Article Snippet: The PI3K Activity ELISA kit (K-1000s) was purchased from Echelon Biosciences (Salt Lake City, UT, USA).

    Techniques: Activity Assay, Enzyme-linked Immunosorbent Assay, shRNA, Positive Control

    Levels of total Akt and P-Akt at serine 473 (ser473) were determined by Western blot analyses in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Density of the P-Akt (ser473) blot was normalized against that of total Akt to obtain a relative blot density, which was expressed as fold changes to the relative P-Akt (ser473) blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Journal: PLoS ONE

    Article Title: Endothelin A Receptor Antagonism Enhances Inhibitory Effects of Anti-Ganglioside GD2 Monoclonal Antibody on Invasiveness and Viability of Human Osteosarcoma Cells

    doi: 10.1371/journal.pone.0093576

    Figure Lengend Snippet: Levels of total Akt and P-Akt at serine 473 (ser473) were determined by Western blot analyses in Saos-2 ( A ), MG-63 ( B ) and SJSA-1 ( C ) cells treated with control IgG (50 µg/mL, lane 2), selective ETAR antagonist BQ123 (5 µM, lane 3), stably-transduced ETAR-shRNA (lane 4), 14G2a mAb (50 µg/mL, lane 5), 14G2a+BQ123 (lane 6), 14G2a+ETAR-shRNA (lane 7), and selective phosphatidylinositide 3-kinase (PI3K) inhibitor BKM120 (50 µM, lane 8). The untreated control was in lane 1 . Density of the P-Akt (ser473) blot was normalized against that of total Akt to obtain a relative blot density, which was expressed as fold changes to the relative P-Akt (ser473) blot density of the untreated control cells (designated as 1). Three independent experiments were performed for each Western blot analysis. Data values were expressed as Mean+SD. a p <0.05 vs. control or control IgG; b p <0.05 vs. BQ123; c p <0.05 vs. ETAR-shRNA; d p <0.05 vs. 14G2a; e p <0.05 vs. 14G2a+BQ123; f p <0.05 vs. 14G2a+ETAR-shRNA.

    Article Snippet: The PI3K Activity ELISA kit (K-1000s) was purchased from Echelon Biosciences (Salt Lake City, UT, USA).

    Techniques: Western Blot, Stable Transfection, shRNA