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    Addgene inc aktar2
    All-optical control of endogenous RAS in steady state. a RAS regulation by light in cells expressing EKAR2G2, in serum. Epifluorescence microscopy; scale bar, 20 µm. Pseudocolor scale: black = 1.0 and white = 2.05. b Normalized whole-cell average FRET/donor ratio in HeLa cells expressing the optogenetically enhanced iB(RAS) and EKAR2G2 FRET biosensor. n = 3, error bars represent SEM. Source data are provided as a Source Data file. c Illumination scheme used for the experiments shown in Fig. 9. d RAS-Akt signaling regulation in cells expressing optogenetically controlled iB(RAS). Representative time-lapse panels of <t>AktAR2</t> FRET/donor ratio, imaged in HeLa cells in serum. Epifluorescence microscopy; scale bar, 20 µm. Pseudocolor scale: black = 1.0; white = 2.88. e Normalized whole-cell average FRET/donor ratio as a function of time, in HeLa cells expressing the AktAR2 FRET biosensor. Error bars represent SEM, n = 3 independent experiments. Source data are provided as a Source Data file. b , e Black: control cells without light-activation. Green: cells with 740 nm illumination starting at t = 300 s time point. Magenta: cells were irradiated with 740 nm light for 2400 s prior to imaging. f Schematic representation of all-optical control of RAS signaling using iB(RAS). Signaling nodes are shown in gray, optogenetic tools and biosensors are shown in light gray. Fast negative feedback loops acting via phosphorylation are shown as red dotted lines. Slow feedback loops acting via transcription inhibition are shown in gray.
    Aktar2, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/aktar2/product/Addgene inc
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    aktar2 - by Bioz Stars, 2021-09
    93/100 stars
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    80
    Addgene inc fret based akt activity reporter aktar2
    Effect of BCR KO on proliferation of GCB-DLBCL lines correlates with reduction in <t>AKT</t> activity. (A) Correlation between relative proliferation after BCR KO in GCB-DLBCL lines, as calculated by the ratio (BCR-KO/BCR replete) of their absolute growth curve slopes, and the BCR KO–induced log2 change in HRK expression. For each cell line, the average value of 2 to 4 biological replicates (each with a different IgH-targeting gRNA) is displayed. (B) Correlation for GCB-DLBCL lines between BCR KO–induced reductions in proliferation and AKT activity, measured by <t>FRET</t> efficiency with the <t>Lyn-AktAR2</t> reporter. AKT activity reduction value is the average of 3 biological replicates, each with a different IgH-targeting gRNA. (C) Relative decline of KO cells in GCB-DLBCL lines after IgH-targeting KO of BCR (upper panel) or combined KO of all 3 AKT genes (lower panel). Values shown are the mean ± SD from 2 (BCR KO) or 3 (AKT KO) biological replicates. (D) Relative decline of BCR-KO cells in DLBCL lines that are either unmodified or have previously undergone PTEN KO. Values shown are the mean ± SD from 3 biological replicates.
    Fret Based Akt Activity Reporter Aktar2, supplied by Addgene inc, used in various techniques. Bioz Stars score: 80/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/fret based akt activity reporter aktar2/product/Addgene inc
    Average 80 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    fret based akt activity reporter aktar2 - by Bioz Stars, 2021-09
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    Image Search Results


    All-optical control of endogenous RAS in steady state. a RAS regulation by light in cells expressing EKAR2G2, in serum. Epifluorescence microscopy; scale bar, 20 µm. Pseudocolor scale: black = 1.0 and white = 2.05. b Normalized whole-cell average FRET/donor ratio in HeLa cells expressing the optogenetically enhanced iB(RAS) and EKAR2G2 FRET biosensor. n = 3, error bars represent SEM. Source data are provided as a Source Data file. c Illumination scheme used for the experiments shown in Fig. 9. d RAS-Akt signaling regulation in cells expressing optogenetically controlled iB(RAS). Representative time-lapse panels of AktAR2 FRET/donor ratio, imaged in HeLa cells in serum. Epifluorescence microscopy; scale bar, 20 µm. Pseudocolor scale: black = 1.0; white = 2.88. e Normalized whole-cell average FRET/donor ratio as a function of time, in HeLa cells expressing the AktAR2 FRET biosensor. Error bars represent SEM, n = 3 independent experiments. Source data are provided as a Source Data file. b , e Black: control cells without light-activation. Green: cells with 740 nm illumination starting at t = 300 s time point. Magenta: cells were irradiated with 740 nm light for 2400 s prior to imaging. f Schematic representation of all-optical control of RAS signaling using iB(RAS). Signaling nodes are shown in gray, optogenetic tools and biosensors are shown in light gray. Fast negative feedback loops acting via phosphorylation are shown as red dotted lines. Slow feedback loops acting via transcription inhibition are shown in gray.

    Journal: Nature Communications

    Article Title: Optogenetic regulation of endogenous proteins

    doi: 10.1038/s41467-020-14460-4

    Figure Lengend Snippet: All-optical control of endogenous RAS in steady state. a RAS regulation by light in cells expressing EKAR2G2, in serum. Epifluorescence microscopy; scale bar, 20 µm. Pseudocolor scale: black = 1.0 and white = 2.05. b Normalized whole-cell average FRET/donor ratio in HeLa cells expressing the optogenetically enhanced iB(RAS) and EKAR2G2 FRET biosensor. n = 3, error bars represent SEM. Source data are provided as a Source Data file. c Illumination scheme used for the experiments shown in Fig. 9. d RAS-Akt signaling regulation in cells expressing optogenetically controlled iB(RAS). Representative time-lapse panels of AktAR2 FRET/donor ratio, imaged in HeLa cells in serum. Epifluorescence microscopy; scale bar, 20 µm. Pseudocolor scale: black = 1.0; white = 2.88. e Normalized whole-cell average FRET/donor ratio as a function of time, in HeLa cells expressing the AktAR2 FRET biosensor. Error bars represent SEM, n = 3 independent experiments. Source data are provided as a Source Data file. b , e Black: control cells without light-activation. Green: cells with 740 nm illumination starting at t = 300 s time point. Magenta: cells were irradiated with 740 nm light for 2400 s prior to imaging. f Schematic representation of all-optical control of RAS signaling using iB(RAS). Signaling nodes are shown in gray, optogenetic tools and biosensors are shown in light gray. Fast negative feedback loops acting via phosphorylation are shown as red dotted lines. Slow feedback loops acting via transcription inhibition are shown in gray.

    Article Snippet: For EKAR2G2 (Addgene #40178) and AKTAR2 (Addgene #64932) biosensor experiments, HeLa cells were transiently transfected together with pQP-AR10 (Supplementary Table ) construct at DNA ratio of 1: 1.

    Techniques: Expressing, Epifluorescence Microscopy, Activation Assay, Irradiation, Imaging, Inhibition

    Effect of BCR KO on proliferation of GCB-DLBCL lines correlates with reduction in AKT activity. (A) Correlation between relative proliferation after BCR KO in GCB-DLBCL lines, as calculated by the ratio (BCR-KO/BCR replete) of their absolute growth curve slopes, and the BCR KO–induced log2 change in HRK expression. For each cell line, the average value of 2 to 4 biological replicates (each with a different IgH-targeting gRNA) is displayed. (B) Correlation for GCB-DLBCL lines between BCR KO–induced reductions in proliferation and AKT activity, measured by FRET efficiency with the Lyn-AktAR2 reporter. AKT activity reduction value is the average of 3 biological replicates, each with a different IgH-targeting gRNA. (C) Relative decline of KO cells in GCB-DLBCL lines after IgH-targeting KO of BCR (upper panel) or combined KO of all 3 AKT genes (lower panel). Values shown are the mean ± SD from 2 (BCR KO) or 3 (AKT KO) biological replicates. (D) Relative decline of BCR-KO cells in DLBCL lines that are either unmodified or have previously undergone PTEN KO. Values shown are the mean ± SD from 3 biological replicates.

    Journal: Blood

    Article Title: Tonic B-cell receptor signaling in diffuse large B-cell lymphoma

    doi: 10.1182/blood-2016-10-747303

    Figure Lengend Snippet: Effect of BCR KO on proliferation of GCB-DLBCL lines correlates with reduction in AKT activity. (A) Correlation between relative proliferation after BCR KO in GCB-DLBCL lines, as calculated by the ratio (BCR-KO/BCR replete) of their absolute growth curve slopes, and the BCR KO–induced log2 change in HRK expression. For each cell line, the average value of 2 to 4 biological replicates (each with a different IgH-targeting gRNA) is displayed. (B) Correlation for GCB-DLBCL lines between BCR KO–induced reductions in proliferation and AKT activity, measured by FRET efficiency with the Lyn-AktAR2 reporter. AKT activity reduction value is the average of 3 biological replicates, each with a different IgH-targeting gRNA. (C) Relative decline of KO cells in GCB-DLBCL lines after IgH-targeting KO of BCR (upper panel) or combined KO of all 3 AKT genes (lower panel). Values shown are the mean ± SD from 2 (BCR KO) or 3 (AKT KO) biological replicates. (D) Relative decline of BCR-KO cells in DLBCL lines that are either unmodified or have previously undergone PTEN KO. Values shown are the mean ± SD from 3 biological replicates.

    Article Snippet: The FRET-based AKT activity reporter AktAR2 (Addgene) was modified by N-terminal addition of the 10 N-terminal amino acids (AA) from Lyn kinase.

    Techniques: Activity Assay, Expressing

    Effect of BCR surface density on tonic BCR signaling across GCB-DLBCL lines. (A) Correlation between BCR surface density based on CD79A-GFP fusion and reduction in proliferation caused by BCR KO in GCB-DLBCL lines. For reduction in proliferation, averages of 3 biological replicates (each with a different IgH-targeting gRNA) are displayed. (B) Correlation between BCR surface density based on CD79A-GFP fusion and BCR KO–induced reduction in AKT activity, as measured by percentage decrease of AKT activity reporter FRET efficiency, E , determined by FCM. For decrease of E , averages of 3 biological replicates (each with a different IgH-targeting gRNA) are displayed.

    Journal: Blood

    Article Title: Tonic B-cell receptor signaling in diffuse large B-cell lymphoma

    doi: 10.1182/blood-2016-10-747303

    Figure Lengend Snippet: Effect of BCR surface density on tonic BCR signaling across GCB-DLBCL lines. (A) Correlation between BCR surface density based on CD79A-GFP fusion and reduction in proliferation caused by BCR KO in GCB-DLBCL lines. For reduction in proliferation, averages of 3 biological replicates (each with a different IgH-targeting gRNA) are displayed. (B) Correlation between BCR surface density based on CD79A-GFP fusion and BCR KO–induced reduction in AKT activity, as measured by percentage decrease of AKT activity reporter FRET efficiency, E , determined by FCM. For decrease of E , averages of 3 biological replicates (each with a different IgH-targeting gRNA) are displayed.

    Article Snippet: The FRET-based AKT activity reporter AktAR2 (Addgene) was modified by N-terminal addition of the 10 N-terminal amino acids (AA) from Lyn kinase.

    Techniques: Activity Assay

    Effect of BCR surface density on tonic BCR signaling. (A) BCR surface density, measured by anti-IgH isotype antibody staining and FCM, increases with doxycycline (Dox) concentration in cells of OCI-Ly19 and SUDHL-4 GCB-DLBCL lines engineered with both KO of the endogenous IgH locus and expression of its same-isotype IgH by a Dox-inducible expression vector. Red and blue arrows on the y-axis mark the similarly determined BCR surface density of unmodified (WT) cells of the lines. For each line, BCR surface density is based on flow cytometric measurement of fluorescence intensity with an antibody specific for its IgH isotype; therefore, values are not directly comparable between isotypes or to BCR surface density determined with a different antibody (eg, to IgL) or by CD79A-GFP fluorescence. (B) Absolute growth curves for GCB-DLBCL lines with inducible IgH expression, relative to the starting point at the indicated Dox concentration (3 days for OCI-Ly19, 5 days for SUDHL-4), showing that growth rates increase with Dox concentration. (C) Correlation between BCR surface density and relative proliferation, defined here as the ratio of slopes of absolute growth curves at different Dox concentrations to that of the curve for maximal Dox concentration (100 ng/mL), for GCB-DLBCL lines at different levels of induced IgH expression. (D) Correlation between BCR surface density and cell size, estimated by the forward scatter value, for GCB-DLBCL lines at different levels of induced IgH expression. (E) Correlation between BCR surface density and AKT activity, measured by FRET efficiency of an AKT activity reporter, for OCI-Ly19 cells with different levels of induced IgH expression, achieved by different durations (up to 7 days) of Dox withdrawal. Data from 2 independent experiments are displayed.

    Journal: Blood

    Article Title: Tonic B-cell receptor signaling in diffuse large B-cell lymphoma

    doi: 10.1182/blood-2016-10-747303

    Figure Lengend Snippet: Effect of BCR surface density on tonic BCR signaling. (A) BCR surface density, measured by anti-IgH isotype antibody staining and FCM, increases with doxycycline (Dox) concentration in cells of OCI-Ly19 and SUDHL-4 GCB-DLBCL lines engineered with both KO of the endogenous IgH locus and expression of its same-isotype IgH by a Dox-inducible expression vector. Red and blue arrows on the y-axis mark the similarly determined BCR surface density of unmodified (WT) cells of the lines. For each line, BCR surface density is based on flow cytometric measurement of fluorescence intensity with an antibody specific for its IgH isotype; therefore, values are not directly comparable between isotypes or to BCR surface density determined with a different antibody (eg, to IgL) or by CD79A-GFP fluorescence. (B) Absolute growth curves for GCB-DLBCL lines with inducible IgH expression, relative to the starting point at the indicated Dox concentration (3 days for OCI-Ly19, 5 days for SUDHL-4), showing that growth rates increase with Dox concentration. (C) Correlation between BCR surface density and relative proliferation, defined here as the ratio of slopes of absolute growth curves at different Dox concentrations to that of the curve for maximal Dox concentration (100 ng/mL), for GCB-DLBCL lines at different levels of induced IgH expression. (D) Correlation between BCR surface density and cell size, estimated by the forward scatter value, for GCB-DLBCL lines at different levels of induced IgH expression. (E) Correlation between BCR surface density and AKT activity, measured by FRET efficiency of an AKT activity reporter, for OCI-Ly19 cells with different levels of induced IgH expression, achieved by different durations (up to 7 days) of Dox withdrawal. Data from 2 independent experiments are displayed.

    Article Snippet: The FRET-based AKT activity reporter AktAR2 (Addgene) was modified by N-terminal addition of the 10 N-terminal amino acids (AA) from Lyn kinase.

    Techniques: Staining, Concentration Assay, Expressing, Plasmid Preparation, Flow Cytometry, Fluorescence, Activity Assay