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  • 86
    Thermo Fisher qt01658692 gapdh
    Qt01658692 Gapdh, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore gapdh
    Detection of CRMP-5 and PKB in retinal proteome through <t>microarray.</t> CRMP-5 was detected significantly higher in the experimental group compared to the control group. Additionally, PKB was detected insignificantly higher in the experimental group. <t>GAPDH</t> served as loading control (n = 6, p
    Gapdh, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 12123 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore g3p dehydrogenase
    A model illustrating glycerol-triggered modulation of root development. The diagram shows the different root patterns in the absence (left) or the presence (right) of glycerol. The middle section shows a condensed schematic of plant glycerol metabolism and three important genes in this study (red). Glycerol is phosphorylated to <t>G3P</t> by GLI1 and can also be generated by GPDHc1 via the reduction of DHAP. G3P is oxidized to DHAP by FAD-GPDH or dephosphorylate to glycerol by GPP. Exogenous glycerol treatment can cause modifications of multiple pathways, including increased G3P and reactive oxygen species (ROS) levels, reduced the phosphate level and expression of PIN1 and PIN7 . It also affected polar auxin transport and the root meristem activity, thus resulting in modified root growth and development. Abbreviations: GLI1, glycerol kinase; GPDHc1, cytosolic glycerol-3-phosphate dehydrogenase; FAD-GPDH, flavin adenine dinucleotide-dependent glycerol-3-phosphate dehydrogenase; GPP, glycerol-3-phosphatase; G3P, glycerol-3-phosphate; DHAP, dihydroxyacetone phosphate; ATP, adenosine triphosphate; FAD, flavin adenine dinucleotide; NADH, the reduced form of nicotinamide adenine dinucleotide. For the sake of clarity, some co-substrates and/or co-products have been omitted in some reactions.
    G3p Dehydrogenase, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 41 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher gapdh
    Effect of G17 on Snail expression in gastric cancer cells . (A) AGSE cells were treated as in 1A or (B) 1B above and subjected to Western Blot analysis utilizing antibodies against Snail and <t>GAPDH</t> (as control). (C) Western Blot analysis of cell extracts with the indicated antibodies, treated with 100 nM G17 for 1 hour, following an overnight pretreatment with 100 nM YM 022. (D) Subconfluent AGSE cells were transiently transfected with Snail-luciferase vector (Snail-luc) along with <t>β-Gal</t> vector (for normalization of transfection). Forty-eight hours after transfection, cells were treated overnight in the presence (+) or absence (-) of 100 nM G17, and luciferase and β-Gal assays were performed. The RLU/β-Gal values were represented as percent control, considering the untreated samples as 100%. Each transfection was performed in triplicate, and the data represent the mean ± SD of at least two independent experiments.
    Gapdh, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 8977 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    AstraZeneca g3p
    Effect of G17 on Snail expression in gastric cancer cells . (A) AGSE cells were treated as in 1A or (B) 1B above and subjected to Western Blot analysis utilizing antibodies against Snail and <t>GAPDH</t> (as control). (C) Western Blot analysis of cell extracts with the indicated antibodies, treated with 100 nM G17 for 1 hour, following an overnight pretreatment with 100 nM YM 022. (D) Subconfluent AGSE cells were transiently transfected with Snail-luciferase vector (Snail-luc) along with <t>β-Gal</t> vector (for normalization of transfection). Forty-eight hours after transfection, cells were treated overnight in the presence (+) or absence (-) of 100 nM G17, and luciferase and β-Gal assays were performed. The RLU/β-Gal values were represented as percent control, considering the untreated samples as 100%. Each transfection was performed in triplicate, and the data represent the mean ± SD of at least two independent experiments.
    G3p, supplied by AstraZeneca, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    American Radiolabeled Chemicals Inc g3p
    Effect of G17 on Snail expression in gastric cancer cells . (A) AGSE cells were treated as in 1A or (B) 1B above and subjected to Western Blot analysis utilizing antibodies against Snail and <t>GAPDH</t> (as control). (C) Western Blot analysis of cell extracts with the indicated antibodies, treated with 100 nM G17 for 1 hour, following an overnight pretreatment with 100 nM YM 022. (D) Subconfluent AGSE cells were transiently transfected with Snail-luciferase vector (Snail-luc) along with <t>β-Gal</t> vector (for normalization of transfection). Forty-eight hours after transfection, cells were treated overnight in the presence (+) or absence (-) of 100 nM G17, and luciferase and β-Gal assays were performed. The RLU/β-Gal values were represented as percent control, considering the untreated samples as 100%. Each transfection was performed in triplicate, and the data represent the mean ± SD of at least two independent experiments.
    G3p, supplied by American Radiolabeled Chemicals Inc, used in various techniques. Bioz Stars score: 89/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    BioVision g3p detection assay buffer
    Effect of G17 on Snail expression in gastric cancer cells . (A) AGSE cells were treated as in 1A or (B) 1B above and subjected to Western Blot analysis utilizing antibodies against Snail and <t>GAPDH</t> (as control). (C) Western Blot analysis of cell extracts with the indicated antibodies, treated with 100 nM G17 for 1 hour, following an overnight pretreatment with 100 nM YM 022. (D) Subconfluent AGSE cells were transiently transfected with Snail-luciferase vector (Snail-luc) along with <t>β-Gal</t> vector (for normalization of transfection). Forty-eight hours after transfection, cells were treated overnight in the presence (+) or absence (-) of 100 nM G17, and luciferase and β-Gal assays were performed. The RLU/β-Gal values were represented as percent control, considering the untreated samples as 100%. Each transfection was performed in triplicate, and the data represent the mean ± SD of at least two independent experiments.
    G3p Detection Assay Buffer, supplied by BioVision, used in various techniques. Bioz Stars score: 86/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore anti gapdh antibody
    FEN1 modulation and DNA damage in combined treated MDA-MB231 cells. (a) Real-time assay: FEN1 RNA expression is detected in cells treated with indicated concentrations of AEs plus and minus PTX (20 nM). 25 μ M AEs vs. 25 μ M AEs+PTX ∗∗ p = 0.0052. (b) FEN1/p-ERK1-2 protein expression: FEN1 protein expression and phosphorylation level of ERK1-2 were detected in total lysate of treated MDA-MB231 cells. Quantification of band intensities was performed using ImageJ software, normalized by <t>β</t> -actin expression level. Relative values are calculated by comparing sample band intensities to control in each setting (±PTX). (c) Effect of p-ERK1-2 inhibitor on FEN1 expression: FEN1 and p-ERK1-2 level were detected in MDA-MB231 treated for 60 minutes with U0126 (20 μ M), a MAPK/ERK inhibitor. Quantification of band intensities was performed using ImageJ software, normalized by <t>GAPDH</t> expression level. Relative values are calculated by comparing sample band intensities to control. (d) DNA damage level: DNA damage marker γ -H 2 AX was detected in cells treated with AEs plus and minus PTX. Quantification of band intensities was performed using ImageJ software, normalized by β -actin expression level. Relative values are calculated by comparing sample band intensities to control in each setting (±PTX).
    Anti Gapdh Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 4811 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher control gapdh
    Seven active chalcones up-regulate the <t>GCLC</t> and HO-1 mRNA levels in PC12 cells. Cells were pretreated with 10 µM chalcones or vehicle control for 6 h. The mRNA levels of antioxidant genes GCLC ( A ) and HO-1 ( B ) were measured by qRT-PCR. The mRNA values for each gene were normalized to those of internal control <t>GAPDH</t> (glyceraldehyde phosphate dehydrogenase) mRNA, and expressed as a ratio to DMSO. Each bar represents the mean ± SD of three separate experiments. Statistical significance relative to DMSO group is indicated as follows: * p
    Control Gapdh, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 213 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    American Radiolabeled Chemicals Inc h g3p
    Seven active chalcones up-regulate the <t>GCLC</t> and HO-1 mRNA levels in PC12 cells. Cells were pretreated with 10 µM chalcones or vehicle control for 6 h. The mRNA levels of antioxidant genes GCLC ( A ) and HO-1 ( B ) were measured by qRT-PCR. The mRNA values for each gene were normalized to those of internal control <t>GAPDH</t> (glyceraldehyde phosphate dehydrogenase) mRNA, and expressed as a ratio to DMSO. Each bar represents the mean ± SD of three separate experiments. Statistical significance relative to DMSO group is indicated as follows: * p
    H G3p, supplied by American Radiolabeled Chemicals Inc, used in various techniques. Bioz Stars score: 85/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Detection of CRMP-5 and PKB in retinal proteome through microarray. CRMP-5 was detected significantly higher in the experimental group compared to the control group. Additionally, PKB was detected insignificantly higher in the experimental group. GAPDH served as loading control (n = 6, p

    Journal: PLoS ONE

    Article Title: Neuroprotective and neuroregenerative effects of CRMP-5 on retinal ganglion cells in an experimental in vivo and in vitro model of glaucoma

    doi: 10.1371/journal.pone.0207190

    Figure Lengend Snippet: Detection of CRMP-5 and PKB in retinal proteome through microarray. CRMP-5 was detected significantly higher in the experimental group compared to the control group. Additionally, PKB was detected insignificantly higher in the experimental group. GAPDH served as loading control (n = 6, p

    Article Snippet: Protein analysis through antibody-microarray CRMP-5-, PKB- and GAPDH- (as a loading control) Abs (GAPDH-AB, Sigma Aldrich, USA) (0.1 mg/mL) were spotted on a glass-nitrocellulose 16 multi-pad slide (Oncyte, Grace Bio-Labs, USA) with nine technical replicates per subarray using a non-contact array spotter (ciFLEXARRAYER 3, Scienion, Germany).

    Techniques: Microarray

    A model illustrating glycerol-triggered modulation of root development. The diagram shows the different root patterns in the absence (left) or the presence (right) of glycerol. The middle section shows a condensed schematic of plant glycerol metabolism and three important genes in this study (red). Glycerol is phosphorylated to G3P by GLI1 and can also be generated by GPDHc1 via the reduction of DHAP. G3P is oxidized to DHAP by FAD-GPDH or dephosphorylate to glycerol by GPP. Exogenous glycerol treatment can cause modifications of multiple pathways, including increased G3P and reactive oxygen species (ROS) levels, reduced the phosphate level and expression of PIN1 and PIN7 . It also affected polar auxin transport and the root meristem activity, thus resulting in modified root growth and development. Abbreviations: GLI1, glycerol kinase; GPDHc1, cytosolic glycerol-3-phosphate dehydrogenase; FAD-GPDH, flavin adenine dinucleotide-dependent glycerol-3-phosphate dehydrogenase; GPP, glycerol-3-phosphatase; G3P, glycerol-3-phosphate; DHAP, dihydroxyacetone phosphate; ATP, adenosine triphosphate; FAD, flavin adenine dinucleotide; NADH, the reduced form of nicotinamide adenine dinucleotide. For the sake of clarity, some co-substrates and/or co-products have been omitted in some reactions.

    Journal: PLoS ONE

    Article Title: Glycerol Affects Root Development through Regulation of Multiple Pathways in Arabidopsis

    doi: 10.1371/journal.pone.0086269

    Figure Lengend Snippet: A model illustrating glycerol-triggered modulation of root development. The diagram shows the different root patterns in the absence (left) or the presence (right) of glycerol. The middle section shows a condensed schematic of plant glycerol metabolism and three important genes in this study (red). Glycerol is phosphorylated to G3P by GLI1 and can also be generated by GPDHc1 via the reduction of DHAP. G3P is oxidized to DHAP by FAD-GPDH or dephosphorylate to glycerol by GPP. Exogenous glycerol treatment can cause modifications of multiple pathways, including increased G3P and reactive oxygen species (ROS) levels, reduced the phosphate level and expression of PIN1 and PIN7 . It also affected polar auxin transport and the root meristem activity, thus resulting in modified root growth and development. Abbreviations: GLI1, glycerol kinase; GPDHc1, cytosolic glycerol-3-phosphate dehydrogenase; FAD-GPDH, flavin adenine dinucleotide-dependent glycerol-3-phosphate dehydrogenase; GPP, glycerol-3-phosphatase; G3P, glycerol-3-phosphate; DHAP, dihydroxyacetone phosphate; ATP, adenosine triphosphate; FAD, flavin adenine dinucleotide; NADH, the reduced form of nicotinamide adenine dinucleotide. For the sake of clarity, some co-substrates and/or co-products have been omitted in some reactions.

    Article Snippet: The DHAP content was assayed in reaction buffer (50 mM HEPES-NaOH, 1 mM MgCl2 and 10 µM NADH) containing 0.75 units of G3P dehydrogenase (Sigma) .

    Techniques: Generated, Expressing, Activity Assay, Modification

    G3P levels in seedlings treated with glycerol (nmol g −1 FW). (A) Wild-type seedlings were grown on agar plates containing 0.5×Murashige and Skoog (MS) medium plus 1% (w/v) sucrose in the absence or presence of 1 mM glycerol from 1–5 days post-germination (dpg). The G3P levels of the seedlings were examined. The data are presented as the mean ± SE (n = 3–4). The asterisks indicate significant differences between the means as determined by Student’s t-test (control versus 1 mM glycerol: *, p

    Journal: PLoS ONE

    Article Title: Glycerol Affects Root Development through Regulation of Multiple Pathways in Arabidopsis

    doi: 10.1371/journal.pone.0086269

    Figure Lengend Snippet: G3P levels in seedlings treated with glycerol (nmol g −1 FW). (A) Wild-type seedlings were grown on agar plates containing 0.5×Murashige and Skoog (MS) medium plus 1% (w/v) sucrose in the absence or presence of 1 mM glycerol from 1–5 days post-germination (dpg). The G3P levels of the seedlings were examined. The data are presented as the mean ± SE (n = 3–4). The asterisks indicate significant differences between the means as determined by Student’s t-test (control versus 1 mM glycerol: *, p

    Article Snippet: The DHAP content was assayed in reaction buffer (50 mM HEPES-NaOH, 1 mM MgCl2 and 10 µM NADH) containing 0.75 units of G3P dehydrogenase (Sigma) .

    Techniques: Mass Spectrometry

    Measurement of G3P binding to GlpT at various temperatures using intrinsic tryptophan fluorescence quenching. G3P was titrated until fluorescence quenching was saturated (about 15 μ M). The dissociation constant of G3P binding was found to be about 0.7 μ M, and substrate binding did not show temperature dependence. Data points represent the mean ± SE of three separate measurements.

    Journal: Biochemistry

    Article Title: Kinetic Evidence Is Consistent with the Rocker-Switch Mechanism of Membrane Transport by GlpT †

    doi: 10.1021/bi701383g

    Figure Lengend Snippet: Measurement of G3P binding to GlpT at various temperatures using intrinsic tryptophan fluorescence quenching. G3P was titrated until fluorescence quenching was saturated (about 15 μ M). The dissociation constant of G3P binding was found to be about 0.7 μ M, and substrate binding did not show temperature dependence. Data points represent the mean ± SE of three separate measurements.

    Article Snippet: For kinetics experiments, after equilibration of the whole cells to the desired temperature, substrate transport was initiated by the addition of [14 C]G3P (Sigma, St. Louis, MO), ranging from 6.25 to 400 μ M in a final reaction volume of 110 μ L (104.5 μ L of cells and 5.5 μ L of [14 C]G3P).

    Techniques: Binding Assay, Fluorescence

    Crystal structure of GlpT and the rocker-switch mechanism. (A) Ribbon diagram of GlpT viewed parallel to the membrane. The molecule consists of 12 transmembrane α-helices. The N-terminal domain is colored green, and the C-terminal domain is colored pink. (B) Schematic diagram of the single-binding site, alternating the access mechanism with a rocker-switch type of movement for the GlpT-mediated G3P–P i exchange reaction. The diagram describes the proposed conformational changes that the transporter undergoes during the reaction cycle. C o represents the protein in the outward-facing conformation, and C i represents the inward-facing one. The G3P substrate is represented by a small disk and triangle, and P i is represented by a small disk. (C) Schematic free-energy diagram illustrating the energy levels of the different conformations of GlpT that occur during the transport reaction cycle under physiological conditions. In the absence of substrate binding, the energy barrier (represented by a dotted line) prevents the conformational interconversion between the C o and C i states of the transporter. Substrate binding lowers the energy barrier sufficiently to allow Brownian motion (kT) to drive the conformational interconversion. S denotes the substrate.

    Journal: Biochemistry

    Article Title: Kinetic Evidence Is Consistent with the Rocker-Switch Mechanism of Membrane Transport by GlpT †

    doi: 10.1021/bi701383g

    Figure Lengend Snippet: Crystal structure of GlpT and the rocker-switch mechanism. (A) Ribbon diagram of GlpT viewed parallel to the membrane. The molecule consists of 12 transmembrane α-helices. The N-terminal domain is colored green, and the C-terminal domain is colored pink. (B) Schematic diagram of the single-binding site, alternating the access mechanism with a rocker-switch type of movement for the GlpT-mediated G3P–P i exchange reaction. The diagram describes the proposed conformational changes that the transporter undergoes during the reaction cycle. C o represents the protein in the outward-facing conformation, and C i represents the inward-facing one. The G3P substrate is represented by a small disk and triangle, and P i is represented by a small disk. (C) Schematic free-energy diagram illustrating the energy levels of the different conformations of GlpT that occur during the transport reaction cycle under physiological conditions. In the absence of substrate binding, the energy barrier (represented by a dotted line) prevents the conformational interconversion between the C o and C i states of the transporter. Substrate binding lowers the energy barrier sufficiently to allow Brownian motion (kT) to drive the conformational interconversion. S denotes the substrate.

    Article Snippet: For kinetics experiments, after equilibration of the whole cells to the desired temperature, substrate transport was initiated by the addition of [14 C]G3P (Sigma, St. Louis, MO), ranging from 6.25 to 400 μ M in a final reaction volume of 110 μ L (104.5 μ L of cells and 5.5 μ L of [14 C]G3P).

    Techniques: Binding Assay

    ( A ) MET transcript levels, as measured by qRT–PCR, after ectopic expression of miR-34c mimic in PC3. Mean of quadruplicate is shown. HPRT and PGK1 were used as endogenous controls. ( B ) Western blot on the protein levels of MET after ectopically expressing miR-34c in PC3, PNT2, and DU145 cells. Both the 170 kDa precursor and the 140 kDa mature form of Met are detected. GAPDH and α -actinin were used as loading controls.

    Journal: British Journal of Cancer

    Article Title: The tumour suppressor miR-34c targets MET in prostate cancer cells

    doi: 10.1038/bjc.2013.449

    Figure Lengend Snippet: ( A ) MET transcript levels, as measured by qRT–PCR, after ectopic expression of miR-34c mimic in PC3. Mean of quadruplicate is shown. HPRT and PGK1 were used as endogenous controls. ( B ) Western blot on the protein levels of MET after ectopically expressing miR-34c in PC3, PNT2, and DU145 cells. Both the 170 kDa precursor and the 140 kDa mature form of Met are detected. GAPDH and α -actinin were used as loading controls.

    Article Snippet: The membrane was incubated with antibodies directed against MET (sc-10, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA), GAPDH 1:20000 (a-GAPDH, MAB374, Chemicon, CA, USA), and α -actinin (sc-17829, Santa Cruz Biotechnology).

    Techniques: Quantitative RT-PCR, Expressing, Western Blot

    rAAV2/1-mediated delivery of miR-Atx3-148 to the cerebellum of SCA3/MJD84.2 mice leads to gene silencing of human mutant Ataxin-3 . ( a ) The U6-miRNA expression cassette was cloned into a recombinant AAV expression vector upstream of a CMV-driven hrGFP expression cassette. rAAV2/1 virus was delivered into the deep cerebellar nuclei (DCN) of SCA3/MJD84.2 transgenic mice. Purkinje neurons in the cerebellar cortex, which project their axons to the DCN, are also transduced by DCN-delivered rAAV2/1 via retrograde transport. ( b – e ) Cerebellar coronal sections were obtained from injected mice 8 weeks post-surgery. Confocal microscopy analysis revealed strong hrGFP expression throughout neurons in the DCN ( b , c ) and in cerebellar Purkinje neurons ( d , e ).Scale bar in ( b ) and ( d ) = 200 µm. Scale bar in ( c ) and ( e ) = 50 µm. ( f ) Mutant human ATXN3 mRNA levels were suppressed following delivery of the rAAV2/1 miR-Atx3-148 virus. In contrast, the steady state levels of mouse Atxn3 and Calbindin-1 mRNAs remained unchanged in the presence of rAAV-miR-Atx3-148 virus. ( g ) Cerebellar mutant Ataxin-3 protein expression was confirmed by western blot using the 1H9 antibody. This antibody recognizes human mutant Ataxin-3 (top band) and endogenous mouse ataxin-3 (lower band). Four control-injected (lanes 1–4) and four rAAV-miR-148-injected (lanes 5–8) SCA3/84.2 mice were compared. As a group, SCA3/84.2 mice injected with rAAV-miR-148 showed a significant reduction (~34%) in mutant Ataxin-3 levels when compared to control injected mice. Gapdh levels were used to normalize for protein loading.* P

    Journal: Molecular Therapy

    Article Title: Silencing Mutant ATXN3 Expression Resolves Molecular Phenotypes in SCA3 Transgenic Mice

    doi: 10.1038/mt.2013.152

    Figure Lengend Snippet: rAAV2/1-mediated delivery of miR-Atx3-148 to the cerebellum of SCA3/MJD84.2 mice leads to gene silencing of human mutant Ataxin-3 . ( a ) The U6-miRNA expression cassette was cloned into a recombinant AAV expression vector upstream of a CMV-driven hrGFP expression cassette. rAAV2/1 virus was delivered into the deep cerebellar nuclei (DCN) of SCA3/MJD84.2 transgenic mice. Purkinje neurons in the cerebellar cortex, which project their axons to the DCN, are also transduced by DCN-delivered rAAV2/1 via retrograde transport. ( b – e ) Cerebellar coronal sections were obtained from injected mice 8 weeks post-surgery. Confocal microscopy analysis revealed strong hrGFP expression throughout neurons in the DCN ( b , c ) and in cerebellar Purkinje neurons ( d , e ).Scale bar in ( b ) and ( d ) = 200 µm. Scale bar in ( c ) and ( e ) = 50 µm. ( f ) Mutant human ATXN3 mRNA levels were suppressed following delivery of the rAAV2/1 miR-Atx3-148 virus. In contrast, the steady state levels of mouse Atxn3 and Calbindin-1 mRNAs remained unchanged in the presence of rAAV-miR-Atx3-148 virus. ( g ) Cerebellar mutant Ataxin-3 protein expression was confirmed by western blot using the 1H9 antibody. This antibody recognizes human mutant Ataxin-3 (top band) and endogenous mouse ataxin-3 (lower band). Four control-injected (lanes 1–4) and four rAAV-miR-148-injected (lanes 5–8) SCA3/84.2 mice were compared. As a group, SCA3/84.2 mice injected with rAAV-miR-148 showed a significant reduction (~34%) in mutant Ataxin-3 levels when compared to control injected mice. Gapdh levels were used to normalize for protein loading.* P

    Article Snippet: Ataxin-3 levels were detected by western blotting using the 1H9 antibody as mentioned above and normalized for Gapdh levels assessed using a monoclonal anti-GAPDH antibody (1:3,000 dilution; Millipore).

    Techniques: Mouse Assay, Mutagenesis, Expressing, Clone Assay, Recombinant, Plasmid Preparation, Transgenic Assay, Injection, Confocal Microscopy, Western Blot

    Lab-on-a-disc scWestern analysis of 14 single cells from a sparse, 24-cell biospecimens. (A) A box-and-whisker plot of cell-size distribution of U251-GFP cells settled in microwells. (B) A false-color overlay of fluorescence micrographs from GFP (green), β-TUB (blue), GADPH (red), STAT3 (black) proteins with fluorescence intensity profile plots. (C) Box-and-whisker plots of STAT3, GFP, β-TUB, and GAPDH distributions are obtained from an area-under-the-curve analysis. Box ends represent 25th and 75th percentiles; IQR is the difference between the 25th and 75th percentiles; median value is the red line at box middle; whiskers spread to 95% confidence limits; and red dots indicate outliers.

    Journal: Lab on a chip

    Article Title: High-selectivity cytology via lab-on-a-disc western blotting of individual cells

    doi: 10.1039/c6lc01333c

    Figure Lengend Snippet: Lab-on-a-disc scWestern analysis of 14 single cells from a sparse, 24-cell biospecimens. (A) A box-and-whisker plot of cell-size distribution of U251-GFP cells settled in microwells. (B) A false-color overlay of fluorescence micrographs from GFP (green), β-TUB (blue), GADPH (red), STAT3 (black) proteins with fluorescence intensity profile plots. (C) Box-and-whisker plots of STAT3, GFP, β-TUB, and GAPDH distributions are obtained from an area-under-the-curve analysis. Box ends represent 25th and 75th percentiles; IQR is the difference between the 25th and 75th percentiles; median value is the red line at box middle; whiskers spread to 95% confidence limits; and red dots indicate outliers.

    Article Snippet: Primary GAPDH, β-Tubulin, turboGFP, STAT3, and H3K79me2 antibodies were purchased from Sigma (SAB2500450), Abcam (ab6046), Pierce (PA5–22688), Cell Signalling (9139), and Abcam (ab3594), respectively.

    Techniques: Whisker Assay, Fluorescence

    Effect of G17 on Snail expression in gastric cancer cells . (A) AGSE cells were treated as in 1A or (B) 1B above and subjected to Western Blot analysis utilizing antibodies against Snail and GAPDH (as control). (C) Western Blot analysis of cell extracts with the indicated antibodies, treated with 100 nM G17 for 1 hour, following an overnight pretreatment with 100 nM YM 022. (D) Subconfluent AGSE cells were transiently transfected with Snail-luciferase vector (Snail-luc) along with β-Gal vector (for normalization of transfection). Forty-eight hours after transfection, cells were treated overnight in the presence (+) or absence (-) of 100 nM G17, and luciferase and β-Gal assays were performed. The RLU/β-Gal values were represented as percent control, considering the untreated samples as 100%. Each transfection was performed in triplicate, and the data represent the mean ± SD of at least two independent experiments.

    Journal: Journal of Molecular Signaling

    Article Title: Glycogen Synthase Kinase-3beta regulates Snail and beta-catenin during gastrin-induced migration of gastric cancer cells

    doi: 10.1186/1750-2187-5-9

    Figure Lengend Snippet: Effect of G17 on Snail expression in gastric cancer cells . (A) AGSE cells were treated as in 1A or (B) 1B above and subjected to Western Blot analysis utilizing antibodies against Snail and GAPDH (as control). (C) Western Blot analysis of cell extracts with the indicated antibodies, treated with 100 nM G17 for 1 hour, following an overnight pretreatment with 100 nM YM 022. (D) Subconfluent AGSE cells were transiently transfected with Snail-luciferase vector (Snail-luc) along with β-Gal vector (for normalization of transfection). Forty-eight hours after transfection, cells were treated overnight in the presence (+) or absence (-) of 100 nM G17, and luciferase and β-Gal assays were performed. The RLU/β-Gal values were represented as percent control, considering the untreated samples as 100%. Each transfection was performed in triplicate, and the data represent the mean ± SD of at least two independent experiments.

    Article Snippet: The antibodies utilized were obtained from the following sources: GSK3β, pGSK3βSer9 , AKT, pAKTSer473 , pc-Jun, c-Jun, Snail and Lamin A/C (Cell Signaling Technology, Danvers, MA), GAPDH (Ambion, Austin, TX), β-catenin (BD Biosciences, San Jose, CA), HA.11 (Covance, Berkeley, CA).

    Techniques: Expressing, Western Blot, Transfection, Luciferase, Plasmid Preparation

    Effect of GSK3β inhibition on G17-induced Snail expression and β-catenin nuclear translocation . (A) AGSE cells were treated with (+) or without (-) 100 nM G17, following an overnight pretreatment with either none (lanes 1, 2), 5 μM (lanes 3, 4) or 10 μM (lanes 5, 6) AR-A014418. Western Blot analysis was then performed with the antibodies indicated. (B) Luciferase (with Snail-luc) and β-Gal assays were performed as in 2D following a 1 hour pretreatment with AR-A014418. (C) AGSE cells were co-transfected with Snail-luc and β-Gal vectors along with either Empty vector (lanes 1, 2), GSK3β-S9A mutant vector (lanes 3, 4) or GSK3β-K/A mutant vector (lanes 5, 6). Luciferase and β-Gal assays were performed after G17 treatment as in 2D. Each transfection (3B, 3C) was performed in triplicate, and the data represent the mean ± SD of at least two independent experiments. (D) Upper Panel : Confluent AGSE cells were treated with G17 for 8 hours after an overnight pretreatment with none (lanes 1, 2), or AR-A014418 (lanes 3, 4) or SP600125 (lanes 5, 6). At the end of treatment, nuclear protein was isolated and subjected to Western Blot analysis with antibodies against β-catenin, GAPDH (cytoplasmic protein) or Lamin A/C (nuclear protein). Lower Panel : Cells were pretreated as in the upper panel, followed by 1 hour G17 treatment and Western Blot analysis.

    Journal: Journal of Molecular Signaling

    Article Title: Glycogen Synthase Kinase-3beta regulates Snail and beta-catenin during gastrin-induced migration of gastric cancer cells

    doi: 10.1186/1750-2187-5-9

    Figure Lengend Snippet: Effect of GSK3β inhibition on G17-induced Snail expression and β-catenin nuclear translocation . (A) AGSE cells were treated with (+) or without (-) 100 nM G17, following an overnight pretreatment with either none (lanes 1, 2), 5 μM (lanes 3, 4) or 10 μM (lanes 5, 6) AR-A014418. Western Blot analysis was then performed with the antibodies indicated. (B) Luciferase (with Snail-luc) and β-Gal assays were performed as in 2D following a 1 hour pretreatment with AR-A014418. (C) AGSE cells were co-transfected with Snail-luc and β-Gal vectors along with either Empty vector (lanes 1, 2), GSK3β-S9A mutant vector (lanes 3, 4) or GSK3β-K/A mutant vector (lanes 5, 6). Luciferase and β-Gal assays were performed after G17 treatment as in 2D. Each transfection (3B, 3C) was performed in triplicate, and the data represent the mean ± SD of at least two independent experiments. (D) Upper Panel : Confluent AGSE cells were treated with G17 for 8 hours after an overnight pretreatment with none (lanes 1, 2), or AR-A014418 (lanes 3, 4) or SP600125 (lanes 5, 6). At the end of treatment, nuclear protein was isolated and subjected to Western Blot analysis with antibodies against β-catenin, GAPDH (cytoplasmic protein) or Lamin A/C (nuclear protein). Lower Panel : Cells were pretreated as in the upper panel, followed by 1 hour G17 treatment and Western Blot analysis.

    Article Snippet: The antibodies utilized were obtained from the following sources: GSK3β, pGSK3βSer9 , AKT, pAKTSer473 , pc-Jun, c-Jun, Snail and Lamin A/C (Cell Signaling Technology, Danvers, MA), GAPDH (Ambion, Austin, TX), β-catenin (BD Biosciences, San Jose, CA), HA.11 (Covance, Berkeley, CA).

    Techniques: Inhibition, Expressing, Translocation Assay, Western Blot, Luciferase, Transfection, Plasmid Preparation, Mutagenesis, Isolation

    Effect of overexpression of GSK3β on G17-induced migration . (A) . Subconfluent AGSE cells were transiently transfected with Empty Vector, GSK3β-WT, GSK3β-KA mutant or GSK3β-S9A mutant vectors. The cells were wounded linearly 48 hours post-transfection and, after an overnight recovery following wounding, they were treated with G17 and pictures obtained at the indicated times. (B) AGSE cells were transfected as in 4A followed by G17 treatment and wound healing assay. The distance of migration of the wounded edges for each time point were measured at several places and the average distance was represented by bar diagrams as

    Journal: Journal of Molecular Signaling

    Article Title: Glycogen Synthase Kinase-3beta regulates Snail and beta-catenin during gastrin-induced migration of gastric cancer cells

    doi: 10.1186/1750-2187-5-9

    Figure Lengend Snippet: Effect of overexpression of GSK3β on G17-induced migration . (A) . Subconfluent AGSE cells were transiently transfected with Empty Vector, GSK3β-WT, GSK3β-KA mutant or GSK3β-S9A mutant vectors. The cells were wounded linearly 48 hours post-transfection and, after an overnight recovery following wounding, they were treated with G17 and pictures obtained at the indicated times. (B) AGSE cells were transfected as in 4A followed by G17 treatment and wound healing assay. The distance of migration of the wounded edges for each time point were measured at several places and the average distance was represented by bar diagrams as "Average Gap". (C) AGSE cells transfected in A and treated with G17 were analyzed for protein expression. Western Blot analysis was performed with an HA.11 antibody to detect ectopic HA-tagged GSK3β proteins and with β-catenin and GAPDH antibodies to detect the corresponding endogenous proteins.

    Article Snippet: The antibodies utilized were obtained from the following sources: GSK3β, pGSK3βSer9 , AKT, pAKTSer473 , pc-Jun, c-Jun, Snail and Lamin A/C (Cell Signaling Technology, Danvers, MA), GAPDH (Ambion, Austin, TX), β-catenin (BD Biosciences, San Jose, CA), HA.11 (Covance, Berkeley, CA).

    Techniques: Over Expression, Migration, Transfection, Plasmid Preparation, Mutagenesis, Wound Healing Assay, Expressing, Western Blot

    FEN1 modulation and DNA damage in combined treated MDA-MB231 cells. (a) Real-time assay: FEN1 RNA expression is detected in cells treated with indicated concentrations of AEs plus and minus PTX (20 nM). 25 μ M AEs vs. 25 μ M AEs+PTX ∗∗ p = 0.0052. (b) FEN1/p-ERK1-2 protein expression: FEN1 protein expression and phosphorylation level of ERK1-2 were detected in total lysate of treated MDA-MB231 cells. Quantification of band intensities was performed using ImageJ software, normalized by β -actin expression level. Relative values are calculated by comparing sample band intensities to control in each setting (±PTX). (c) Effect of p-ERK1-2 inhibitor on FEN1 expression: FEN1 and p-ERK1-2 level were detected in MDA-MB231 treated for 60 minutes with U0126 (20 μ M), a MAPK/ERK inhibitor. Quantification of band intensities was performed using ImageJ software, normalized by GAPDH expression level. Relative values are calculated by comparing sample band intensities to control. (d) DNA damage level: DNA damage marker γ -H 2 AX was detected in cells treated with AEs plus and minus PTX. Quantification of band intensities was performed using ImageJ software, normalized by β -actin expression level. Relative values are calculated by comparing sample band intensities to control in each setting (±PTX).

    Journal: Oxidative Medicine and Cellular Longevity

    Article Title: Artichoke Polyphenols Sensitize Human Breast Cancer Cells to Chemotherapeutic Drugs via a ROS-Mediated Downregulation of Flap Endonuclease 1

    doi: 10.1155/2020/7965435

    Figure Lengend Snippet: FEN1 modulation and DNA damage in combined treated MDA-MB231 cells. (a) Real-time assay: FEN1 RNA expression is detected in cells treated with indicated concentrations of AEs plus and minus PTX (20 nM). 25 μ M AEs vs. 25 μ M AEs+PTX ∗∗ p = 0.0052. (b) FEN1/p-ERK1-2 protein expression: FEN1 protein expression and phosphorylation level of ERK1-2 were detected in total lysate of treated MDA-MB231 cells. Quantification of band intensities was performed using ImageJ software, normalized by β -actin expression level. Relative values are calculated by comparing sample band intensities to control in each setting (±PTX). (c) Effect of p-ERK1-2 inhibitor on FEN1 expression: FEN1 and p-ERK1-2 level were detected in MDA-MB231 treated for 60 minutes with U0126 (20 μ M), a MAPK/ERK inhibitor. Quantification of band intensities was performed using ImageJ software, normalized by GAPDH expression level. Relative values are calculated by comparing sample band intensities to control. (d) DNA damage level: DNA damage marker γ -H 2 AX was detected in cells treated with AEs plus and minus PTX. Quantification of band intensities was performed using ImageJ software, normalized by β -actin expression level. Relative values are calculated by comparing sample band intensities to control in each setting (±PTX).

    Article Snippet: 1 : 1000), anti-LC3 (MBL International, Woburn, MA, USA, PD014 dil.1 : 400), anti-ERK1-2 (Cell Signaling Technology #9102 dil.1 : 1000), anti-pERK1-2 (Cell Signaling Technology # 9101S dil.1 : 1000), anti-FEN1 (Santa Cruz Biotechnology Inc. Dallas, TX, USA, sc-28355 dil.1 : 1000), anti-β -actin (MP # 69100 dil.1 : 10000), anti-GAPDH (Sigma Aldrich G8795 dil.

    Techniques: Multiple Displacement Amplification, RNA Expression, Expressing, Software, Marker

    Seven active chalcones up-regulate the GCLC and HO-1 mRNA levels in PC12 cells. Cells were pretreated with 10 µM chalcones or vehicle control for 6 h. The mRNA levels of antioxidant genes GCLC ( A ) and HO-1 ( B ) were measured by qRT-PCR. The mRNA values for each gene were normalized to those of internal control GAPDH (glyceraldehyde phosphate dehydrogenase) mRNA, and expressed as a ratio to DMSO. Each bar represents the mean ± SD of three separate experiments. Statistical significance relative to DMSO group is indicated as follows: * p

    Journal: International Journal of Molecular Sciences

    Article Title: Synthetic Chalcones with Potent Antioxidant Ability on H2O2-Induced Apoptosis in PC12 Cells

    doi: 10.3390/ijms151018525

    Figure Lengend Snippet: Seven active chalcones up-regulate the GCLC and HO-1 mRNA levels in PC12 cells. Cells were pretreated with 10 µM chalcones or vehicle control for 6 h. The mRNA levels of antioxidant genes GCLC ( A ) and HO-1 ( B ) were measured by qRT-PCR. The mRNA values for each gene were normalized to those of internal control GAPDH (glyceraldehyde phosphate dehydrogenase) mRNA, and expressed as a ratio to DMSO. Each bar represents the mean ± SD of three separate experiments. Statistical significance relative to DMSO group is indicated as follows: * p

    Article Snippet: Reverse transcription and amplification of target genes GCLC and HO-1 , as well as the internal control GAPDH , were performed according to the manufacturer’s instructions (Invitrogen, Carlsbad, CA, USA).

    Techniques: Quantitative RT-PCR