β actin  (Millipore)

 
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    Name:
    PSF CBA FLUC
    Description:
    In PSF CBA FLUC CHICKEN BETA ACTIN PROMOTER PLASMID expression of Firefly luciferase is regulated by the chicken beta actin CBA mammalian promoterPromoter Expression Level Plasmid vector contains the chicken beta actin CBA promoter to drive protein expression The CBA promoter contains a GC rich region CpG island that helps to maintain expression of the promoter in long term culture
    Catalog Number:
    OGS598
    Price:
    None
    Applications:
    Cloning in a gene: PSF-CBA-FLUC - CHICKEN BETA ACTIN PROMOTER PLASMID contains a gene within the main multiple cloning site (NotI-ClaI). Any plasmid that we sell where the gene is this configuration will be located in the exact same position in relation to the start and stop codon of the gene. The only exceptions to this rule are fusions proteins where the fusion gene may be positioned at the front or end of the MCS to allow gene fusion.By positioning all of our genes in the same location it allows them to be transferred between plasmids using the same cloning method and restriction sites regardless of the plasmid being used from our product range. Inserting a new gene into this plasmid should be easily possible using a range of standard restriction enzyme sites that flank the gene currently in the vector.Multiple cloning site notes: In the multiple cloning site there are two important restriction sites called BsgI and BseRI sites. These sites both cut the DNA at the same position and cleave the stop codon of the gene in the multiple cloning site in this plasmid thereby producing a TA overhang. This overhang is compatible with any of our peptide or reporter fusion tag plasmids also cut with either of these enzymes. This allows seamless C-terminal fusions to be made with the gene in this multiple cloning site using a single cloning step from our C-terminal peptide and reporter tag product range. Normally the easiest method is to clone the C-terminal tag from our other plasmid products into this plasmid using BsgI or BseRI and the downstream ClaI restriction site.BseRI and BsgI sites are non-palindromic and cleave a defined number of bases away from their binding sites. This allows them to cut the upstream stop codon in the gene in this plasmid regardless of the gene sequence.
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    Structured Review

    Millipore β actin
    PSF CBA FLUC
    In PSF CBA FLUC CHICKEN BETA ACTIN PROMOTER PLASMID expression of Firefly luciferase is regulated by the chicken beta actin CBA mammalian promoterPromoter Expression Level Plasmid vector contains the chicken beta actin CBA promoter to drive protein expression The CBA promoter contains a GC rich region CpG island that helps to maintain expression of the promoter in long term culture
    https://www.bioz.com/result/β actin/product/Millipore
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    β actin - by Bioz Stars, 2021-04
    86/100 stars

    Images

    1) Product Images from "A Locus Encompassing the Epstein-Barr Virus bglf4 Kinase Regulates Expression of Genes Encoding Viral Structural Proteins"

    Article Title: A Locus Encompassing the Epstein-Barr Virus bglf4 Kinase Regulates Expression of Genes Encoding Viral Structural Proteins

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1004307

    BGLF3 regulates late gene expression. A) Western blot analysis of EBV lytic proteins to study the effect of knocking down expression of BGLF3 and BGLF3.5. 2089 cells were transfected with two concentrations of the ZEBRA expression plasmid and siRNAs to BGLF3 (siG3) and BGLF3.5 (siG3.5). Cells were harvested after 48 hours of transfection. The membrane was blotted with antibodies specific to BGLF4, ZEBRA and the late FR3 protein. The level of β-actin used as a loading control. B) Total DNA was prepared from the same samples and was analyzed by quantitative PCR for the level of the upstream region of oriLyt as a marker for EBV replication. The relative DNA concentration was calculated using the standard curve method. The experiment is a representation of two biological replicates.
    Figure Legend Snippet: BGLF3 regulates late gene expression. A) Western blot analysis of EBV lytic proteins to study the effect of knocking down expression of BGLF3 and BGLF3.5. 2089 cells were transfected with two concentrations of the ZEBRA expression plasmid and siRNAs to BGLF3 (siG3) and BGLF3.5 (siG3.5). Cells were harvested after 48 hours of transfection. The membrane was blotted with antibodies specific to BGLF4, ZEBRA and the late FR3 protein. The level of β-actin used as a loading control. B) Total DNA was prepared from the same samples and was analyzed by quantitative PCR for the level of the upstream region of oriLyt as a marker for EBV replication. The relative DNA concentration was calculated using the standard curve method. The experiment is a representation of two biological replicates.

    Techniques Used: Expressing, Western Blot, Transfection, Plasmid Preparation, Real-time Polymerase Chain Reaction, Marker, Concentration Assay

    2) Product Images from "Autophagy limits the cytotoxic effects of the AKT inhibitor AZ7328 in human bladder cancer cells"

    Article Title: Autophagy limits the cytotoxic effects of the AKT inhibitor AZ7328 in human bladder cancer cells

    Journal: Cancer Biology & Therapy

    doi: 10.4161/cbt.21793

    Figure 5. Potential predictors of response to AKT inhibition. (A) Western blot analysis of baseline PTEN status among the panel of 12 cell lines. The corresponding relative density indicates the PTEN band intensity relative to β-actin.
    Figure Legend Snippet: Figure 5. Potential predictors of response to AKT inhibition. (A) Western blot analysis of baseline PTEN status among the panel of 12 cell lines. The corresponding relative density indicates the PTEN band intensity relative to β-actin.

    Techniques Used: Inhibition, Western Blot

    3) Product Images from "An Activin A/BMP2 chimera, AB215, blocks estrogen signaling via induction of ID proteins in breast cancer cells"

    Article Title: An Activin A/BMP2 chimera, AB215, blocks estrogen signaling via induction of ID proteins in breast cancer cells

    Journal: BMC Cancer

    doi: 10.1186/1471-2407-14-549

    Enhanced signaling capacity of AB215 compared to BMP2 in breast cancer cells. A) Time course western blot analysis of MCF7 cells was performed after exposing BMP2 and AB215 for indicated time. The arrow points to the ligand-induced upper band. B - C) ID1-driven Luciferase reporter assay of BMP2 and AB215 for MCF7, T47D and SK-BR-3 cells in the B) absence and C) presence of 10nM E2. Cells were reverse transfected with ID1-Luciferase reporter plasmid in triplicate. Transfected cells were exposed to ligands for 18 hours. Relative luciferase value has been normalized with β-gal. The data is shown in means ± SD, (n: number of independent experiments = 3). D) mRNA level of IDs in MCF7 cells. BMP2 and AB215 induced mRNA expression level of ID1/2/3/4 has been measured by qRT-PCR in MCF7 cells. Cells were exposed to 500 ng/ml of BMP2 and AB215 for 18 hours. Relative mRNA level was calculated in quadruplicate using ΔΔCτ method with the control and β-actin as a reference. All data are shown in means + SD, n = 3. ( * = P ≤ 0.05, ** = P ≤ 0.01, *** = P ≤ 0.001 ).
    Figure Legend Snippet: Enhanced signaling capacity of AB215 compared to BMP2 in breast cancer cells. A) Time course western blot analysis of MCF7 cells was performed after exposing BMP2 and AB215 for indicated time. The arrow points to the ligand-induced upper band. B - C) ID1-driven Luciferase reporter assay of BMP2 and AB215 for MCF7, T47D and SK-BR-3 cells in the B) absence and C) presence of 10nM E2. Cells were reverse transfected with ID1-Luciferase reporter plasmid in triplicate. Transfected cells were exposed to ligands for 18 hours. Relative luciferase value has been normalized with β-gal. The data is shown in means ± SD, (n: number of independent experiments = 3). D) mRNA level of IDs in MCF7 cells. BMP2 and AB215 induced mRNA expression level of ID1/2/3/4 has been measured by qRT-PCR in MCF7 cells. Cells were exposed to 500 ng/ml of BMP2 and AB215 for 18 hours. Relative mRNA level was calculated in quadruplicate using ΔΔCτ method with the control and β-actin as a reference. All data are shown in means + SD, n = 3. ( * = P ≤ 0.05, ** = P ≤ 0.01, *** = P ≤ 0.001 ).

    Techniques Used: Western Blot, Luciferase, Reporter Assay, Transfection, Plasmid Preparation, Expressing, Quantitative RT-PCR

    4) Product Images from "Therapeutic potential of combined BRAF/MEK blockade in BRAF-wild type preclinical tumor models"

    Article Title: Therapeutic potential of combined BRAF/MEK blockade in BRAF-wild type preclinical tumor models

    Journal: Journal of Experimental & Clinical Cancer Research : CR

    doi: 10.1186/s13046-018-0820-5

    Effects of single and combined MEK and RAF inhibition in patient-derived pancreatic organoids. a . Phase-contrast images showing an organoid formation assay. Organoids were seeded as singles cells and treated after 48 h with different doses of trametinib (10 nM,), dabrafenib (10 μM) and combination ratio 1:1000. Images were detected with EVOS Cell Imaging System ( b ). Patient-derived pancreatic organoid T1 ( KRAS -mut) was incubated with dabrafenib and trametinib alone or in combination for 24 h. Protein lysate were analyzed by Western Blotting using p-ERK antibody. β-Actin is used as protein loading at blotting control. c . Patient-derived pancreatic organoid T1 was treated with increasing amounts of the dabrafenib (0.1–10 μM) alone or in combination with trametinib (01–10 nM) ratio 1:1000, for 72 h. Cell viability was evaluated by Cell Titer Glo 2.0 assay. The table shows the CI of trametinib and dabrafenib in a normal pancreatic organoid N1 and in 4 different PDAC organoids KRAS -mut
    Figure Legend Snippet: Effects of single and combined MEK and RAF inhibition in patient-derived pancreatic organoids. a . Phase-contrast images showing an organoid formation assay. Organoids were seeded as singles cells and treated after 48 h with different doses of trametinib (10 nM,), dabrafenib (10 μM) and combination ratio 1:1000. Images were detected with EVOS Cell Imaging System ( b ). Patient-derived pancreatic organoid T1 ( KRAS -mut) was incubated with dabrafenib and trametinib alone or in combination for 24 h. Protein lysate were analyzed by Western Blotting using p-ERK antibody. β-Actin is used as protein loading at blotting control. c . Patient-derived pancreatic organoid T1 was treated with increasing amounts of the dabrafenib (0.1–10 μM) alone or in combination with trametinib (01–10 nM) ratio 1:1000, for 72 h. Cell viability was evaluated by Cell Titer Glo 2.0 assay. The table shows the CI of trametinib and dabrafenib in a normal pancreatic organoid N1 and in 4 different PDAC organoids KRAS -mut

    Techniques Used: Inhibition, Derivative Assay, Tube Formation Assay, Imaging, Incubation, Western Blot

    5) Product Images from "The Antioxidant N-Acetylcysteine Prevents HIF-1 Stabilization under Hypoxia In Vitro but Does Not Affect Tumorigenesis in Multiple Breast Cancer Models In Vivo"

    Article Title: The Antioxidant N-Acetylcysteine Prevents HIF-1 Stabilization under Hypoxia In Vitro but Does Not Affect Tumorigenesis in Multiple Breast Cancer Models In Vivo

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0066388

    NAC treatment does not alter the hypoxic phenotype of primary tumors. Tumors from NAC-treated or control mice were stained with Hif-1α (red), PIM (green) and DAPI (blue). Five sections per tumor were stained and imaged to obtain an average percentage threshold area of Hif-1α staining determined from 5 random images per section. A) Representative images of PyMT endstage tumors (from Figure 3G ) with Hif-1α staining values calculated for individual tumors ( B) and averaged (C) (control n = 5 tumors; NAC n = 7 tumors). D) Representative images of EO771 endstage tumors (from Figure 3C–D ) with Hif-1α staining values calculated for individual tumors ( E) and averaged (F) (n = 6 tumors for all groups). Mean ± SEM; n.s. indicates not significant. G) EO771 tumor lysates (from Figure 3C–D ) were probed for Hif-1α and β-actin (loading control) by Western blot (control n = 3 tumors; NAC n = 4 tumors).
    Figure Legend Snippet: NAC treatment does not alter the hypoxic phenotype of primary tumors. Tumors from NAC-treated or control mice were stained with Hif-1α (red), PIM (green) and DAPI (blue). Five sections per tumor were stained and imaged to obtain an average percentage threshold area of Hif-1α staining determined from 5 random images per section. A) Representative images of PyMT endstage tumors (from Figure 3G ) with Hif-1α staining values calculated for individual tumors ( B) and averaged (C) (control n = 5 tumors; NAC n = 7 tumors). D) Representative images of EO771 endstage tumors (from Figure 3C–D ) with Hif-1α staining values calculated for individual tumors ( E) and averaged (F) (n = 6 tumors for all groups). Mean ± SEM; n.s. indicates not significant. G) EO771 tumor lysates (from Figure 3C–D ) were probed for Hif-1α and β-actin (loading control) by Western blot (control n = 3 tumors; NAC n = 4 tumors).

    Techniques Used: Mouse Assay, Staining, Western Blot

    6) Product Images from "Convergence of Notch and ?-catenin signaling induces arterial fate in vascular progenitors"

    Article Title: Convergence of Notch and ?-catenin signaling induces arterial fate in vascular progenitors

    Journal: The Journal of Cell Biology

    doi: 10.1083/jcb.200904114

    Enhancement of arterial gene expression through dual activation of Notch and β-catenin during in vivo angiogenesis. Matrigel containing VEGF (100 ng/ml), heparin (10 units/ml), and adenoviral vectors (vehicle [control], HA-tagged N1ICD [NICD], and/or CA-β-catenin) were injected subcutaneously in mice. After 7 d, the mice were sacrificed and plugs were excised. (A) Western blot for HA-tagged N1ICD and CA-β-catenin in recovered cells from Matrigel plugs. (B) Hematoxylin and eosin staining of Matrigel sections. Overall appearances were not different. Invasion of blood vessels with vascular lumen and blood cells were observed. Bars: 200 µm. (C) Representative result of Western blot for VE-cadherin, ephrinB2, and Neuropilin1 in recovered cells from Matrigel plugs. (D) Quantitative evaluation of VE-cadherin (left graph), ephrinB2 (middle graph), and Neuropilin1 (right graph) protein expression in Matrigels. Relative expression normalized with β-actin expression is shown. ( n = 3; *, P
    Figure Legend Snippet: Enhancement of arterial gene expression through dual activation of Notch and β-catenin during in vivo angiogenesis. Matrigel containing VEGF (100 ng/ml), heparin (10 units/ml), and adenoviral vectors (vehicle [control], HA-tagged N1ICD [NICD], and/or CA-β-catenin) were injected subcutaneously in mice. After 7 d, the mice were sacrificed and plugs were excised. (A) Western blot for HA-tagged N1ICD and CA-β-catenin in recovered cells from Matrigel plugs. (B) Hematoxylin and eosin staining of Matrigel sections. Overall appearances were not different. Invasion of blood vessels with vascular lumen and blood cells were observed. Bars: 200 µm. (C) Representative result of Western blot for VE-cadherin, ephrinB2, and Neuropilin1 in recovered cells from Matrigel plugs. (D) Quantitative evaluation of VE-cadherin (left graph), ephrinB2 (middle graph), and Neuropilin1 (right graph) protein expression in Matrigels. Relative expression normalized with β-actin expression is shown. ( n = 3; *, P

    Techniques Used: Expressing, Activation Assay, In Vivo, Injection, Mouse Assay, Western Blot, Staining

    7) Product Images from "Mutations in the IGF-II pathway that confer resistance to lytic reovirus infection"

    Article Title: Mutations in the IGF-II pathway that confer resistance to lytic reovirus infection

    Journal: BMC Cell Biology

    doi: 10.1186/1471-2121-5-32

    CTCF and Igf2 expression in RIE-1 and 6B72 cells. Levels of CTCF protein were assessed by Western blot analysis (A), normalized to a β-actin control. Levels of Igf2 transcripts were assessed by Northern blot analysis (B) normalized to GAPDH control. Protein content, as assayed by western blot analysis and standardized to β-actin was decreased in the 6B72 cell clone to 30% of control. Reverse transcriptase PCR analysis of Igf2 transcripts (C). The RT-PCR products (arrows) were separated on a 2% agarose gel, revealing an additional transcript in the 6B72 cells. The DNA sequence of the larger RT-PCR product (D) revealed an alternatively spliced transcript ( Igf2 sv ) generated by splicing of exon 2 to a cryptic 3' splice site located 14 nucleotides upstream of exon 3.
    Figure Legend Snippet: CTCF and Igf2 expression in RIE-1 and 6B72 cells. Levels of CTCF protein were assessed by Western blot analysis (A), normalized to a β-actin control. Levels of Igf2 transcripts were assessed by Northern blot analysis (B) normalized to GAPDH control. Protein content, as assayed by western blot analysis and standardized to β-actin was decreased in the 6B72 cell clone to 30% of control. Reverse transcriptase PCR analysis of Igf2 transcripts (C). The RT-PCR products (arrows) were separated on a 2% agarose gel, revealing an additional transcript in the 6B72 cells. The DNA sequence of the larger RT-PCR product (D) revealed an alternatively spliced transcript ( Igf2 sv ) generated by splicing of exon 2 to a cryptic 3' splice site located 14 nucleotides upstream of exon 3.

    Techniques Used: Expressing, Western Blot, Northern Blot, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Agarose Gel Electrophoresis, Sequencing, Generated

    8) Product Images from "The Role of Protein Kinase A Regulation of the E6 PDZ-Binding Domain during the Differentiation-Dependent Life Cycle of Human Papillomavirus Type 18"

    Article Title: The Role of Protein Kinase A Regulation of the E6 PDZ-Binding Domain during the Differentiation-Dependent Life Cycle of Human Papillomavirus Type 18

    Journal: Journal of Virology

    doi: 10.1128/JVI.01234-13

    Details of E6 C-terminal mutations in HPV18 genomes and analysis of the activity of these mutations in an in vivo degradation assay. (A) Two mutant HPV18 genomes were constructed. E6ΔPDZ contains base substitutions at nucleotides 567, 570, and 571, thereby altering the codons for glutamic acid 155 and threonine 156 to translation termination codons. This mutant genome supports expression of an E6 protein in which the PBM, ETQV, is deleted. E6153PKA contains a single base substitution at nucleotide 562 that alters the codon for arginine 153 to leucine; the mutant protein expressed from this genome is not a substrate for PKA and targets cellular PDZ proteins in a constitutive, PKA-independent manner ( 17 ). The position of the initiator methionine codon for the E7 protein, which is separated from the translation termination codon of E6 by 8 nucleotides, is indicated. (B) In vivo E6-mediated degradation assays of epitope-tagged PDZ domain-containing targets (DLG1, MAGI-1, and SCRIB) and p53. Urea-soluble cell extracts were harvested at 24 and 48 h posttransfection, and the level of E6 targets was assessed by Western blotting. Each antibody data set (all four transfections) was taken from the same exposure. Shown below each degradation experiment are the amounts of PDZ proteins or p53 remaining relative to the transfections with empty vector (following normalization to the loading control [GAPDH or β-actin]). The efficiency of transfections of each set of expression plasmids was monitored by inclusion of a GFP-expressing plasmid, and the efficiencies were shown to be equivalent (WT, 32%; E6153PKA, 33%; E6ΔPDZ, 32%) between the cotransfections.
    Figure Legend Snippet: Details of E6 C-terminal mutations in HPV18 genomes and analysis of the activity of these mutations in an in vivo degradation assay. (A) Two mutant HPV18 genomes were constructed. E6ΔPDZ contains base substitutions at nucleotides 567, 570, and 571, thereby altering the codons for glutamic acid 155 and threonine 156 to translation termination codons. This mutant genome supports expression of an E6 protein in which the PBM, ETQV, is deleted. E6153PKA contains a single base substitution at nucleotide 562 that alters the codon for arginine 153 to leucine; the mutant protein expressed from this genome is not a substrate for PKA and targets cellular PDZ proteins in a constitutive, PKA-independent manner ( 17 ). The position of the initiator methionine codon for the E7 protein, which is separated from the translation termination codon of E6 by 8 nucleotides, is indicated. (B) In vivo E6-mediated degradation assays of epitope-tagged PDZ domain-containing targets (DLG1, MAGI-1, and SCRIB) and p53. Urea-soluble cell extracts were harvested at 24 and 48 h posttransfection, and the level of E6 targets was assessed by Western blotting. Each antibody data set (all four transfections) was taken from the same exposure. Shown below each degradation experiment are the amounts of PDZ proteins or p53 remaining relative to the transfections with empty vector (following normalization to the loading control [GAPDH or β-actin]). The efficiency of transfections of each set of expression plasmids was monitored by inclusion of a GFP-expressing plasmid, and the efficiencies were shown to be equivalent (WT, 32%; E6153PKA, 33%; E6ΔPDZ, 32%) between the cotransfections.

    Techniques Used: Activity Assay, In Vivo, Degradation Assay, Mutagenesis, Construct, Expressing, Western Blot, Transfection, Plasmid Preparation

    9) Product Images from "Neuroprotective Effects of a Smoothened Receptor Agonist against Early Brain Injury after Experimental Subarachnoid Hemorrhage in Rats"

    Article Title: Neuroprotective Effects of a Smoothened Receptor Agonist against Early Brain Injury after Experimental Subarachnoid Hemorrhage in Rats

    Journal: Frontiers in Cellular Neuroscience

    doi: 10.3389/fncel.2016.00306

    Effects of treatment with PUR on Bax, Bcl-2 in mRNA and protein levels. (A) The relative expression levels of Bax and Bcl-2 mRNA in the PFC were analyzed by RT-PCR. The densities of the protein bands were analyzed and normalized to β-actin, n = 3. (B) Representative western blots showing levels of Bax and Bcl-2 in the PFC, Bar graphs showing quantification of the protein levels of Bax and Bcl-2, n = 3. Both the determination of the two signs in mRNA and protein levels were obtained from three separate experiments. Values represent the mean ± SD. *** p
    Figure Legend Snippet: Effects of treatment with PUR on Bax, Bcl-2 in mRNA and protein levels. (A) The relative expression levels of Bax and Bcl-2 mRNA in the PFC were analyzed by RT-PCR. The densities of the protein bands were analyzed and normalized to β-actin, n = 3. (B) Representative western blots showing levels of Bax and Bcl-2 in the PFC, Bar graphs showing quantification of the protein levels of Bax and Bcl-2, n = 3. Both the determination of the two signs in mRNA and protein levels were obtained from three separate experiments. Values represent the mean ± SD. *** p

    Techniques Used: Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot

    Effects of PUR on Shh pathway . The quantification of Shh, Gli-1, and Ptch was measured by RT-PCR (A) and western blot (B) 48 h post-SAH. Each value was normalized to β-actin. Bar graphs showing quantification of mRNA and protein levels of Shh, Gli-1, and Ptch were determined by Image-Pro Plus 6.0, n = 3. Values represent the mean ± SD. ** p
    Figure Legend Snippet: Effects of PUR on Shh pathway . The quantification of Shh, Gli-1, and Ptch was measured by RT-PCR (A) and western blot (B) 48 h post-SAH. Each value was normalized to β-actin. Bar graphs showing quantification of mRNA and protein levels of Shh, Gli-1, and Ptch were determined by Image-Pro Plus 6.0, n = 3. Values represent the mean ± SD. ** p

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Western Blot

    10) Product Images from "Ribavirin Enhances IFN-? Signalling and MxA Expression: A Novel Immune Modulation Mechanism during Treatment of HCV"

    Article Title: Ribavirin Enhances IFN-? Signalling and MxA Expression: A Novel Immune Modulation Mechanism during Treatment of HCV

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0027866

    HCV core co-localises with MxA. (A) Immunoblot of lysates from Huh7 cells transfected with HCV-DNA construct for 0, 6, 12 and 24 h, probed with HCV core and β-Actin (n = 3). (B) Confocal micrograph of Huh7s transfected with EV or HCV-DNA construct (n = 3). (C) Huh7s transfected with HCV-DNA construct and treated with IFN-α for 2 h, Ribavirin for 2 h or both for 2 h (n = 4). Confocal micrographs show MxA, HCV core and the nucleus, labelled with Alexa 488, Alexa 568 and DAPI, respectively. Antibody staining is indicated along the side of images Bar, 10 µm (n = 3).
    Figure Legend Snippet: HCV core co-localises with MxA. (A) Immunoblot of lysates from Huh7 cells transfected with HCV-DNA construct for 0, 6, 12 and 24 h, probed with HCV core and β-Actin (n = 3). (B) Confocal micrograph of Huh7s transfected with EV or HCV-DNA construct (n = 3). (C) Huh7s transfected with HCV-DNA construct and treated with IFN-α for 2 h, Ribavirin for 2 h or both for 2 h (n = 4). Confocal micrographs show MxA, HCV core and the nucleus, labelled with Alexa 488, Alexa 568 and DAPI, respectively. Antibody staining is indicated along the side of images Bar, 10 µm (n = 3).

    Techniques Used: Transfection, Construct, Staining

    IFN-α induced STAT1 and STAT3 phosphorylation is enhanced in the presence of Ribavirin. Lysates from Huh7 hepatocytes treated with IFN-α for 2 h, Ribavirin for 2 h or both for 2 h were immunoblotted for pSTAT1 and 3 and reprobed for their protein counterparts and β-actin loading control (data representative of three independent experiments).
    Figure Legend Snippet: IFN-α induced STAT1 and STAT3 phosphorylation is enhanced in the presence of Ribavirin. Lysates from Huh7 hepatocytes treated with IFN-α for 2 h, Ribavirin for 2 h or both for 2 h were immunoblotted for pSTAT1 and 3 and reprobed for their protein counterparts and β-actin loading control (data representative of three independent experiments).

    Techniques Used:

    11) Product Images from "Lactate Activates HIF-1 in Oxidative but Not in Warburg-Phenotype Human Tumor Cells"

    Article Title: Lactate Activates HIF-1 in Oxidative but Not in Warburg-Phenotype Human Tumor Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0046571

    MCT1 and CD147 interact at the plasma membrane of oxidative tumor cells. (A) The left graph shows basal SLC16A1 /MCT1 mRNA expression normalized to RPL19 mRNA expression detected in SiHa and WiDr TCs using RT-qPCR. ** p = 0.0062; n = 5–8. On the right, MCT1 and β-actin were detected using Western blotting (WB) in cell lysates. The upper panels show a representative experiment and the graph HIF-1α protein expression normalized to β-actin levels. *** p = 0.0004; n = 5. (B) as in (A) but detecting CD147 instead of MCT1. RT-qPCR: *** p
    Figure Legend Snippet: MCT1 and CD147 interact at the plasma membrane of oxidative tumor cells. (A) The left graph shows basal SLC16A1 /MCT1 mRNA expression normalized to RPL19 mRNA expression detected in SiHa and WiDr TCs using RT-qPCR. ** p = 0.0062; n = 5–8. On the right, MCT1 and β-actin were detected using Western blotting (WB) in cell lysates. The upper panels show a representative experiment and the graph HIF-1α protein expression normalized to β-actin levels. *** p = 0.0004; n = 5. (B) as in (A) but detecting CD147 instead of MCT1. RT-qPCR: *** p

    Techniques Used: Expressing, Quantitative RT-PCR, Western Blot

    Targeting MCT1 inhibits lactate-induced but not basal HIF-1 activity in tumor cells. (A) SiHa TCs were cultured during 24-h in fresh medium containing 10 mM lactate, lactate +5 mM α-cyano-4-hydroxycinnamate (CHC), or none of the drugs. HIF-1α and β-actin were detected using Western blotting. The upper panels show a representative experiment and the graph shows HIF-1α protein expression normalized to β-actin. *** p
    Figure Legend Snippet: Targeting MCT1 inhibits lactate-induced but not basal HIF-1 activity in tumor cells. (A) SiHa TCs were cultured during 24-h in fresh medium containing 10 mM lactate, lactate +5 mM α-cyano-4-hydroxycinnamate (CHC), or none of the drugs. HIF-1α and β-actin were detected using Western blotting. The upper panels show a representative experiment and the graph shows HIF-1α protein expression normalized to β-actin. *** p

    Techniques Used: Activity Assay, Cell Culture, Western Blot, Expressing

    Lactate induces normoxic HIF-1α protein stabilization in oxidative tumor cells, not in Warburg tumor cells. (A) Lactate release in the supernatant of SiHa and WiDr TCs was measured using a CMA600 enzymatic analyzer after 24-h of culture in fresh medium. *** p = 0.0002; n = 4. (B–G) HIF-1α and β-actin protein expression was detected using Western blotting in the lysates of oxidative SiHa or Warburg WiDr TCs. The upper panels show representative experiments and the graphs HIF-1α protein expression normalized to β-actin levels. (B) SiHa and WiDr cells were untreated to detect basal HIF-1α protein expression. * p = 0.0414; n = 3–4. (C) SiHa TCs were cultured during 24-h under hypoxia (1% O 2 ) or not. * p = 0.011; n = 3. (D) As in (C) but with WiDr TCs. ** p = 0.0057; n = 5. (E) SiHa TCs were cultured during 24-h in the presence of 10 mM lactate or not. ** p = 0.0029; n = 4. (F) As in (E) but with WiDr TCs. ns , p = 0.1449; n = 8. (G) SiHa TCs were exposed to increasing doses of lactate during 24-h. * p
    Figure Legend Snippet: Lactate induces normoxic HIF-1α protein stabilization in oxidative tumor cells, not in Warburg tumor cells. (A) Lactate release in the supernatant of SiHa and WiDr TCs was measured using a CMA600 enzymatic analyzer after 24-h of culture in fresh medium. *** p = 0.0002; n = 4. (B–G) HIF-1α and β-actin protein expression was detected using Western blotting in the lysates of oxidative SiHa or Warburg WiDr TCs. The upper panels show representative experiments and the graphs HIF-1α protein expression normalized to β-actin levels. (B) SiHa and WiDr cells were untreated to detect basal HIF-1α protein expression. * p = 0.0414; n = 3–4. (C) SiHa TCs were cultured during 24-h under hypoxia (1% O 2 ) or not. * p = 0.011; n = 3. (D) As in (C) but with WiDr TCs. ** p = 0.0057; n = 5. (E) SiHa TCs were cultured during 24-h in the presence of 10 mM lactate or not. ** p = 0.0029; n = 4. (F) As in (E) but with WiDr TCs. ns , p = 0.1449; n = 8. (G) SiHa TCs were exposed to increasing doses of lactate during 24-h. * p

    Techniques Used: Expressing, Western Blot, Cell Culture

    Lactate inhibits PHD2 activity in oxidative tumor cells. (A) HIF-1α and β-actin protein expression was detected using Western blotting in the lysates of SiHa TCs incubated during 24-h with 10 mM lactate or not and increasing doses of 2-oxoglutarate. The upper panels show representative experiments and the graph HIF-1α protein expression normalized to β-actin levels. Data are expressed as % of lactate induction. ** p
    Figure Legend Snippet: Lactate inhibits PHD2 activity in oxidative tumor cells. (A) HIF-1α and β-actin protein expression was detected using Western blotting in the lysates of SiHa TCs incubated during 24-h with 10 mM lactate or not and increasing doses of 2-oxoglutarate. The upper panels show representative experiments and the graph HIF-1α protein expression normalized to β-actin levels. Data are expressed as % of lactate induction. ** p

    Techniques Used: Activity Assay, Expressing, Western Blot, Incubation

    12) Product Images from "Cyclin-dependent kinase 9 (CDK9) is a novel prognostic marker and therapeutic target in osteosarcoma"

    Article Title: Cyclin-dependent kinase 9 (CDK9) is a novel prognostic marker and therapeutic target in osteosarcoma

    Journal: EBioMedicine

    doi: 10.1016/j.ebiom.2018.12.022

    CDK9 inhibition by siRNA transfection decreases osteosarcoma cell proliferation and migration . CDK9 expression and osteosarcoma cell proliferation of (A) U2OS and (B) KHOS were determined by immunofluorescence after an incubation with a CDK9 antibody (green) and β-Actin (red). Representative images of (C) U2OS, and (D) KHOS cell migration after CDK9 siRNA and negative control (NC) siRNA transfection for 0 h, 24 h, 48 h, and 72 h. The distance of cell migration of the osteosarcoma cell lines (E) U2OS, and (F) KHOS were measured after siRNA transfection. (Scale bar, 50 μm).
    Figure Legend Snippet: CDK9 inhibition by siRNA transfection decreases osteosarcoma cell proliferation and migration . CDK9 expression and osteosarcoma cell proliferation of (A) U2OS and (B) KHOS were determined by immunofluorescence after an incubation with a CDK9 antibody (green) and β-Actin (red). Representative images of (C) U2OS, and (D) KHOS cell migration after CDK9 siRNA and negative control (NC) siRNA transfection for 0 h, 24 h, 48 h, and 72 h. The distance of cell migration of the osteosarcoma cell lines (E) U2OS, and (F) KHOS were measured after siRNA transfection. (Scale bar, 50 μm).

    Techniques Used: Inhibition, Transfection, Migration, Expressing, Immunofluorescence, Incubation, Negative Control

    13) Product Images from "Combination treatment with leptin and pioglitazone in a mouse model of Alzheimer's disease"

    Article Title: Combination treatment with leptin and pioglitazone in a mouse model of Alzheimer's disease

    Journal: Alzheimer's & Dementia : Translational Research & Clinical Interventions

    doi: 10.1016/j.trci.2016.11.002

    Effect of leptin and pioglitazone treatment on synaptic protein levels. (A and B) Representative β-actin–normalized immunoblot images and quantitation of SYN (A.1 and B.1), PSD-95 (A.2 and B.2), vGLUT1 (A.3 and B.3), and GAD65/67 (A.4
    Figure Legend Snippet: Effect of leptin and pioglitazone treatment on synaptic protein levels. (A and B) Representative β-actin–normalized immunoblot images and quantitation of SYN (A.1 and B.1), PSD-95 (A.2 and B.2), vGLUT1 (A.3 and B.3), and GAD65/67 (A.4

    Techniques Used: Quantitation Assay

    14) Product Images from "Expression and cyclic variations of catechol-O-methyl transferase in human endometrial stroma"

    Article Title: Expression and cyclic variations of catechol-O-methyl transferase in human endometrial stroma

    Journal: Fertility and sterility

    doi: 10.1016/j.fertnstert.2007.01.042

    Effects of estrogen and progesterone on COMT protein expression in pHES cells. Soluble-COMT protein expression was up-regulated with estrogen treatment and down-regulated with progesterone treatment. β-actin is the internal loading control. Lane
    Figure Legend Snippet: Effects of estrogen and progesterone on COMT protein expression in pHES cells. Soluble-COMT protein expression was up-regulated with estrogen treatment and down-regulated with progesterone treatment. β-actin is the internal loading control. Lane

    Techniques Used: Expressing

    Effect of 2-ME2 on Bcl-2 and VEGF protein expression in pHES. 2-Methoxy estradiol markedly down-regulated Bcl-2 and VEGF protein expression at 1 µM. β-actin is the internal loading control. +C indicates positive control, C indicates vehicle
    Figure Legend Snippet: Effect of 2-ME2 on Bcl-2 and VEGF protein expression in pHES. 2-Methoxy estradiol markedly down-regulated Bcl-2 and VEGF protein expression at 1 µM. β-actin is the internal loading control. +C indicates positive control, C indicates vehicle

    Techniques Used: Expressing, Positive Control

    Related Articles

    Software:

    Article Title: Anthocyanins Derived from Vitis coignetiae Pulliat Contributes Anti-Cancer Effects by Suppressing NF-κB Pathways in Hep3B Human Hepatocellular Carcinoma Cells and In Vivo
    Article Snippet: .. ImageJ software 1.51K ( http://rsb.info.nih.gov) was used to quantify each protein band followed by densitometry reading undertaken after normalization with β-Actin expression. .. Animal Experiment Athymic (nu/nu) male nude mice, obtained from SLC (Sankyo Lab Service, Tokyo, Japan), were housed under pathogen-free conditions with a 12 h light/12 h dark schedule, and fed with an autoclaved diet and water.

    Article Title: Zi Shen Huo Luo Formula Prevents Aldosterone-Induced Cardiomyocyte Hypertrophy and Cardiac Fibroblast Proliferation by Regulating the Striatin-Mediated MR/EGFR/ERK Signaling Pathway
    Article Snippet: Finally, immunoreactivity was visualized with an ECL kit (Millipore, MA, USA). .. The relative expression levels (ratio compared to Na-K ATPase or β-actin expression) of the target proteins were quantified by ImageJ software. ..

    Expressing:

    Article Title: Anthocyanins Derived from Vitis coignetiae Pulliat Contributes Anti-Cancer Effects by Suppressing NF-κB Pathways in Hep3B Human Hepatocellular Carcinoma Cells and In Vivo
    Article Snippet: .. ImageJ software 1.51K ( http://rsb.info.nih.gov) was used to quantify each protein band followed by densitometry reading undertaken after normalization with β-Actin expression. .. Animal Experiment Athymic (nu/nu) male nude mice, obtained from SLC (Sankyo Lab Service, Tokyo, Japan), were housed under pathogen-free conditions with a 12 h light/12 h dark schedule, and fed with an autoclaved diet and water.

    Article Title: Rhus verniciflua Stokes extract suppresses migration and invasion in human gastric adenocarcinoma AGS cells
    Article Snippet: The relative intensity of each protein band was quantified using an Image Quant™ LAS 500 imaging system (GE Healthcare Bio-Sciences AB). .. The expression levels were normalized to β-actin expression. ..

    Article Title: Zi Shen Huo Luo Formula Prevents Aldosterone-Induced Cardiomyocyte Hypertrophy and Cardiac Fibroblast Proliferation by Regulating the Striatin-Mediated MR/EGFR/ERK Signaling Pathway
    Article Snippet: Finally, immunoreactivity was visualized with an ECL kit (Millipore, MA, USA). .. The relative expression levels (ratio compared to Na-K ATPase or β-actin expression) of the target proteins were quantified by ImageJ software. ..

    Article Title: Dimethyl Fumarate Attenuates Lung Inflammation and Oxidative Stress Induced by Chronic Exposure to Diesel Exhaust Particles in Mice
    Article Snippet: The antigen–antibody complexes were detected using enhanced chemiluminescence (Santa Cruz Biotechnology, Dallas, TX, USA). .. The β-actin expression was used as a loading control for all immunoblotting and data were expressed as arbitrary units. ..

    Article Title: Hypoxia induced exosomal circRNA promotes metastasis of Colorectal Cancer via targeting GEF-H1/RhoA axis
    Article Snippet: .. Western blot analysisThe expression of protein was assessed by western blot analysis, and its expression in the samples was normalized to β-actin expression. .. Cells and tissues were lysed in RIPA buffer with freshly added protease inhibitor cocktail.

    Plasmid Preparation:

    Article Title: Stable High-Producer Cell Clone Expressing Virus-Like Particles of the Japanese Encephalitis Virus E Protein for a Second-Generation Subunit Vaccine
    Article Snippet: A JEV cDNA clone, J12, encoding the viral signal peptide of the carboxyl terminus of C, prM, and E proteins (amino acid positions 105 to 794) of the Beijing-1 strain was recovered from a vaccinia virus transfer plasmid, pS13J12Gal (unpublished data), by digestion with Sal I and Xho I and then inserted into the Xho I site of a pCAGGS expression vector ( ) to generate pCAGJ12. .. The fragment containing the CMV enhancer, the β-actin promoter and intron, and J12 cDNA was recovered from the pCAGJ12 plasmid and replaced with the regulatory region of a pEFBOSbsr plasmid (a kind gift from M. Tatsumi) to confer resistance to blasticidin S (BS) (Calbiochem, Inc., Darmstadt, Germany). .. The resulting plasmid, pCAGJ12bsr (Fig. ), was used to establish stable cell lines.

    Western Blot:

    Article Title: Hypoxia induced exosomal circRNA promotes metastasis of Colorectal Cancer via targeting GEF-H1/RhoA axis
    Article Snippet: .. Western blot analysisThe expression of protein was assessed by western blot analysis, and its expression in the samples was normalized to β-actin expression. .. Cells and tissues were lysed in RIPA buffer with freshly added protease inhibitor cocktail.

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    • β actin antibodies  (Millipore)
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    Millipore β actin antibodies
    Knockout of EZH2 gene by CRISPR/Cas9 and stable infection of EBV. (A) Loss of EZH2 expression in the EZH2-KO Akata(−) cells. Two independent clones were used (named cl1 and cl2) for both WT and KO. Levels of EZH2, H3K27me3, histone H3, and <t>β-actin</t> in the samples from WT and EZH2-KO cells were examined by Western blotting. (B) Growth kinetics of EZH2-KO Akata(−) cells. A quantity of 1.0 × 10 5 cells/ml was seeded and cultured. Cell numbers were counted after the indicated number of days. (C) Reduced growth speed in the EZH2-KO cells. The WT and EZH2-KO Akata cells were infected with EBV and cultured in the presence of G418. At 0, 4, 8, and 12 weeks postinfection, cells were seeded at 1.0 × 10 5 cells/ml, and cell numbers were counted after 3 days. The average and SD from 2 independent replicates are shown as a ratio relative to the value of the starting point (0 week). Student’s t test was performed, and asterisks indicate statistical significance (*, P
    β Actin Antibodies, supplied by Millipore, 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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    mouse monoclonal anti β actin antibody  (Millipore)
    97
    Millipore mouse monoclonal anti β actin antibody
    Expression of tumor necrosis factor (TNF)-α in lipopolysaccharide (LPS)-stimulated normal human dermal fibroblasts (NHDFs) and expression of heat shock protein 47 (HSP47) in transforming growth factor-β (TGF-β)-stimulated NHDFs. ( a ) TNF-α secreted from the NHDFs in the culture media was detected by Western blotting. ( b ) Expression of HSP47 in TGF-β-stimulated NHDFs. These graphs show TNF-α and HSP47 expression levels determined by densitometry. TNF-α and HSP47 expression levels were normalized against <t>β-actin</t> expression levels.
    Mouse Monoclonal Anti β Actin Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    mouse monoclonal anti β actin antibody - by Bioz Stars, 2021-04
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    anti β actin antibody  (Millipore)
    97
    Millipore anti β actin antibody
    Characterization of phenotypic and functional features of SP and non-SP cell fractions from metastatic and AI PC3 cells and cytotoxic effects induced by GDC-0449 and docetaxel on SP PC3 cells (A) Representative data of the Hoechst dye efflux profile obtained after staining of parental PC3 cell line with fluorescent Hoechst dye showing the SP cell subpopulation (green) and non-SP fraction (blue) detected in the total mass of PC3 cells. (B) Western blot analyses of expression levels of prostatic stem cell-like markers (CD133 and CD44, multidrug transporter ABCG2), SHH, GLI-1, mesenchymal associated molecules (N-cadherin and vimentin) and <t>β-actin</t> detected in SP cells versus non-SP cell fraction isolated from parental PC3 cell line. (C) and (D) Representative pictures of the immunofluorescence analyses of the expression levels and cellular localization of markers in SP and non-SP PC3 cells shown at the original magnification of × 630. (E) Representative pictures of the dense prostaspheres formed by SP PC3 cells without or after a treatment with rhSHH as compared to diffuse, abortive and very small aggregates formed by SHH-stimulated non-SP PC3 cells. Moreover, the prostaspheres derived from PC3 cells untreated or treated with 5 uM GDC0449 and 5 nM docetaxel, alone or in combination, are shown at a similar magnification of × 200. (F) The self-renewal capability of SP cells versus the non-SP cell fraction from PC3 cells was estimated based on their ability to form the non-adherent aggregates in serum-free culture conditions under ultra-low attachment plate (Corning, Invitrogen). The quantitative data of the number of prostaspheres formed by the SP PC3 cell fraction without (basal level) or after a treatment with exogenous rhSHH in the absence (control) or presence of 5 μM GDC-0449 and 5 nM docetaxel, alone or in combination, are also shown. * p
    Anti β Actin Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Knockout of EZH2 gene by CRISPR/Cas9 and stable infection of EBV. (A) Loss of EZH2 expression in the EZH2-KO Akata(−) cells. Two independent clones were used (named cl1 and cl2) for both WT and KO. Levels of EZH2, H3K27me3, histone H3, and β-actin in the samples from WT and EZH2-KO cells were examined by Western blotting. (B) Growth kinetics of EZH2-KO Akata(−) cells. A quantity of 1.0 × 10 5 cells/ml was seeded and cultured. Cell numbers were counted after the indicated number of days. (C) Reduced growth speed in the EZH2-KO cells. The WT and EZH2-KO Akata cells were infected with EBV and cultured in the presence of G418. At 0, 4, 8, and 12 weeks postinfection, cells were seeded at 1.0 × 10 5 cells/ml, and cell numbers were counted after 3 days. The average and SD from 2 independent replicates are shown as a ratio relative to the value of the starting point (0 week). Student’s t test was performed, and asterisks indicate statistical significance (*, P

    Journal: mSphere

    Article Title: Regulation of Epstein-Barr Virus Life Cycle and Cell Proliferation by Histone H3K27 Methyltransferase EZH2 in Akata Cells

    doi: 10.1128/mSphere.00478-18

    Figure Lengend Snippet: Knockout of EZH2 gene by CRISPR/Cas9 and stable infection of EBV. (A) Loss of EZH2 expression in the EZH2-KO Akata(−) cells. Two independent clones were used (named cl1 and cl2) for both WT and KO. Levels of EZH2, H3K27me3, histone H3, and β-actin in the samples from WT and EZH2-KO cells were examined by Western blotting. (B) Growth kinetics of EZH2-KO Akata(−) cells. A quantity of 1.0 × 10 5 cells/ml was seeded and cultured. Cell numbers were counted after the indicated number of days. (C) Reduced growth speed in the EZH2-KO cells. The WT and EZH2-KO Akata cells were infected with EBV and cultured in the presence of G418. At 0, 4, 8, and 12 weeks postinfection, cells were seeded at 1.0 × 10 5 cells/ml, and cell numbers were counted after 3 days. The average and SD from 2 independent replicates are shown as a ratio relative to the value of the starting point (0 week). Student’s t test was performed, and asterisks indicate statistical significance (*, P

    Article Snippet: Anti-histone H3K4me3 and β-actin antibodies were from Millipore and Sigma-Aldrich, respectively.

    Techniques: Knock-Out, CRISPR, Infection, Expressing, Clone Assay, Western Blot, Cell Culture

    Increased viral gene expression in WT and EZH2-KO Akata cells during lytic phase. (A to H) WT and EZH2-KO cells latently infected with EBV were treated with anti-IgG. RNA was harvested from the cells at 0 (latency), 1, and 2 days postinduction (dpi) and subjected to qRT-PCR analysis. The data are normalized and shown as fold change for the average and SD from three samples. (I) WT and EZH2-KO cells latently infected with EBV were treated with anti-IgG as in panels A to H. Protein samples were collected on days 0 and 2, and LMP1, BZLF1, BRLF1, BALF2, BMRF1, gB, and β-actin levels were assessed by Western blotting. Student’s t test was performed, and asterisks indicate statistical significance (*, P

    Journal: mSphere

    Article Title: Regulation of Epstein-Barr Virus Life Cycle and Cell Proliferation by Histone H3K27 Methyltransferase EZH2 in Akata Cells

    doi: 10.1128/mSphere.00478-18

    Figure Lengend Snippet: Increased viral gene expression in WT and EZH2-KO Akata cells during lytic phase. (A to H) WT and EZH2-KO cells latently infected with EBV were treated with anti-IgG. RNA was harvested from the cells at 0 (latency), 1, and 2 days postinduction (dpi) and subjected to qRT-PCR analysis. The data are normalized and shown as fold change for the average and SD from three samples. (I) WT and EZH2-KO cells latently infected with EBV were treated with anti-IgG as in panels A to H. Protein samples were collected on days 0 and 2, and LMP1, BZLF1, BRLF1, BALF2, BMRF1, gB, and β-actin levels were assessed by Western blotting. Student’s t test was performed, and asterisks indicate statistical significance (*, P

    Article Snippet: Anti-histone H3K4me3 and β-actin antibodies were from Millipore and Sigma-Aldrich, respectively.

    Techniques: Expressing, Infection, Quantitative RT-PCR, Western Blot

    Expression of tumor necrosis factor (TNF)-α in lipopolysaccharide (LPS)-stimulated normal human dermal fibroblasts (NHDFs) and expression of heat shock protein 47 (HSP47) in transforming growth factor-β (TGF-β)-stimulated NHDFs. ( a ) TNF-α secreted from the NHDFs in the culture media was detected by Western blotting. ( b ) Expression of HSP47 in TGF-β-stimulated NHDFs. These graphs show TNF-α and HSP47 expression levels determined by densitometry. TNF-α and HSP47 expression levels were normalized against β-actin expression levels.

    Journal: Polymers

    Article Title: Development of a Gene Delivery System of Oligonucleotides for Fibroses by Targeting Cell-Surface Vimentin-Expressing Cells with N-Acetylglucosamine-Bearing Polymer-Conjugated Polyethyleneimine

    doi: 10.3390/polym12071508

    Figure Lengend Snippet: Expression of tumor necrosis factor (TNF)-α in lipopolysaccharide (LPS)-stimulated normal human dermal fibroblasts (NHDFs) and expression of heat shock protein 47 (HSP47) in transforming growth factor-β (TGF-β)-stimulated NHDFs. ( a ) TNF-α secreted from the NHDFs in the culture media was detected by Western blotting. ( b ) Expression of HSP47 in TGF-β-stimulated NHDFs. These graphs show TNF-α and HSP47 expression levels determined by densitometry. TNF-α and HSP47 expression levels were normalized against β-actin expression levels.

    Article Snippet: Mouse monoclonal anti-β-actin antibody was purchased from Sigma-Aldrich.

    Techniques: Expressing, Western Blot

    Characterization of phenotypic and functional features of SP and non-SP cell fractions from metastatic and AI PC3 cells and cytotoxic effects induced by GDC-0449 and docetaxel on SP PC3 cells (A) Representative data of the Hoechst dye efflux profile obtained after staining of parental PC3 cell line with fluorescent Hoechst dye showing the SP cell subpopulation (green) and non-SP fraction (blue) detected in the total mass of PC3 cells. (B) Western blot analyses of expression levels of prostatic stem cell-like markers (CD133 and CD44, multidrug transporter ABCG2), SHH, GLI-1, mesenchymal associated molecules (N-cadherin and vimentin) and β-actin detected in SP cells versus non-SP cell fraction isolated from parental PC3 cell line. (C) and (D) Representative pictures of the immunofluorescence analyses of the expression levels and cellular localization of markers in SP and non-SP PC3 cells shown at the original magnification of × 630. (E) Representative pictures of the dense prostaspheres formed by SP PC3 cells without or after a treatment with rhSHH as compared to diffuse, abortive and very small aggregates formed by SHH-stimulated non-SP PC3 cells. Moreover, the prostaspheres derived from PC3 cells untreated or treated with 5 uM GDC0449 and 5 nM docetaxel, alone or in combination, are shown at a similar magnification of × 200. (F) The self-renewal capability of SP cells versus the non-SP cell fraction from PC3 cells was estimated based on their ability to form the non-adherent aggregates in serum-free culture conditions under ultra-low attachment plate (Corning, Invitrogen). The quantitative data of the number of prostaspheres formed by the SP PC3 cell fraction without (basal level) or after a treatment with exogenous rhSHH in the absence (control) or presence of 5 μM GDC-0449 and 5 nM docetaxel, alone or in combination, are also shown. * p

    Journal: Oncotarget

    Article Title: Inhibition of hedgehog signaling improves the anti-carcinogenic effects of docetaxel in prostate cancer

    doi:

    Figure Lengend Snippet: Characterization of phenotypic and functional features of SP and non-SP cell fractions from metastatic and AI PC3 cells and cytotoxic effects induced by GDC-0449 and docetaxel on SP PC3 cells (A) Representative data of the Hoechst dye efflux profile obtained after staining of parental PC3 cell line with fluorescent Hoechst dye showing the SP cell subpopulation (green) and non-SP fraction (blue) detected in the total mass of PC3 cells. (B) Western blot analyses of expression levels of prostatic stem cell-like markers (CD133 and CD44, multidrug transporter ABCG2), SHH, GLI-1, mesenchymal associated molecules (N-cadherin and vimentin) and β-actin detected in SP cells versus non-SP cell fraction isolated from parental PC3 cell line. (C) and (D) Representative pictures of the immunofluorescence analyses of the expression levels and cellular localization of markers in SP and non-SP PC3 cells shown at the original magnification of × 630. (E) Representative pictures of the dense prostaspheres formed by SP PC3 cells without or after a treatment with rhSHH as compared to diffuse, abortive and very small aggregates formed by SHH-stimulated non-SP PC3 cells. Moreover, the prostaspheres derived from PC3 cells untreated or treated with 5 uM GDC0449 and 5 nM docetaxel, alone or in combination, are shown at a similar magnification of × 200. (F) The self-renewal capability of SP cells versus the non-SP cell fraction from PC3 cells was estimated based on their ability to form the non-adherent aggregates in serum-free culture conditions under ultra-low attachment plate (Corning, Invitrogen). The quantitative data of the number of prostaspheres formed by the SP PC3 cell fraction without (basal level) or after a treatment with exogenous rhSHH in the absence (control) or presence of 5 μM GDC-0449 and 5 nM docetaxel, alone or in combination, are also shown. * p

    Article Snippet: Fetal bovine serum (FBS), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), 3′,3′-dihexyloxacarbocyanine iodide (DiOC6 (3)) 2′,7′-dichlorofluorescein diacetate (DCFH-DA), mouse monoclonal anti-vimentin (V6630) and anti-β-actin antibody (clone AC-15) were obtained from Sigma-Aldrich (St. Louis, MO).

    Techniques: Functional Assay, Staining, Western Blot, Expressing, Isolation, Immunofluorescence, Derivative Assay

    Disruption of the Foxo1 gene results in embryonic lethality. ( A ) PCR analysis with genomic DNA from E9.5 yolk sacs obtained from a mating of Foxo1 +/- mice. PCR amplification from the wild-type and Foxo1- targeted loci resulted in fragment sizes of 301 bp and 815 bp, respectively. Samples were independently amplified with each primer set. ( B ) Western blot analysis of lysates from E9.5 Foxo1 +/+ and Foxo1 -/- embryos by using antibodies against Foxo1 and β-actin. ( C ) At E9.5, both Foxo1 +/+ and Foxo1 +/- yolk sacs had well developed blood vessels, whereas Foxo1 -/- yolk sacs lacked them. ( D ) At E9.5, Foxo1 +/+ and Foxo1 +/- embryos were phenotypically indistinguishable in appearance, whereas Foxo1 -/- embryos were approximately half their size. In addition, cardiac looping was retarded and the pericardium was distended (arrowhead). (Scale bar, 500 μm.)

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Disruption of forkhead transcription factor (FOXO) family members in mice reveals their functional diversification

    doi: 10.1073/pnas.0400093101

    Figure Lengend Snippet: Disruption of the Foxo1 gene results in embryonic lethality. ( A ) PCR analysis with genomic DNA from E9.5 yolk sacs obtained from a mating of Foxo1 +/- mice. PCR amplification from the wild-type and Foxo1- targeted loci resulted in fragment sizes of 301 bp and 815 bp, respectively. Samples were independently amplified with each primer set. ( B ) Western blot analysis of lysates from E9.5 Foxo1 +/+ and Foxo1 -/- embryos by using antibodies against Foxo1 and β-actin. ( C ) At E9.5, both Foxo1 +/+ and Foxo1 +/- yolk sacs had well developed blood vessels, whereas Foxo1 -/- yolk sacs lacked them. ( D ) At E9.5, Foxo1 +/+ and Foxo1 +/- embryos were phenotypically indistinguishable in appearance, whereas Foxo1 -/- embryos were approximately half their size. In addition, cardiac looping was retarded and the pericardium was distended (arrowhead). (Scale bar, 500 μm.)

    Article Snippet: Antibodies used for Western analysis were polyclonal anti-FKHR antibody (C-20, Santa Cruz Biotechnology), polyclonal anti-FKHRL1 antibody (Upstate Biotechnology, Waltham, MA), monoclonal anti-β-actin antibody (clone AC-15, Sigma), and horseradish peroxidase (HRP)-conjugated secondary antibodies (Dako).

    Techniques: Polymerase Chain Reaction, Mouse Assay, Amplification, Western Blot

    Targeted disruption of the Foxo3a gene and Foxo4 gene. ( A ) Targeting strategy for Foxo3a . A retroviral promoter trap vector was inserted in intron 1 of Foxo3a . SA, splice acceptor site; SD, splice donor site; LTR, long terminal repeat sequence; pA, polyadenylation signal; puro, puromycin. ( B ) Southern analysis of Sac I-digested genomic DNA obtained from a mating of Foxo3a +/- mice by using the Neo - and Csk -specific probes. Different intensities demonstrated by hybridization of the Neo-specific probe discriminate zero, one, or two Foxo3a gene disruptions, representing +/+, +/-, and -/- mice, respectively. ( C ) Western analysis of lung tissue lysates from Foxo3a +/+ and Foxo3a -/- mice by using antibodies against Foxo3a and β-actin. ( D ) Restriction maps of the wild-type Foxo4 locus, the targeting vector, and the targeted locus. The targeting vector contained a PGK-Neo cassette instead of the Foxo4 exon1. A targeting probe (3′ region of Foxo4 cDNA) was also indicated. Restriction enzymes were indicated as B, Bam HI; C, Cla I; E, Eco RI; H, Hin dIII; N, Nco I; and Sp, Spe I. ( E ) Southern analysis of Bam HI-digested genomic DNA obtained from Foxo4 female (+/+, +/-, and -/-) and male (+/y and -/y) mice by using a probe from the 3′ region of Foxo4 cDNA. The Foxo4 wild-type allele (10 kb) and targeted allele (16 kb) were indicated. ( F ) Northern analysis of total RNA from skeletal muscle of Foxo4 female (+/+ and -/-) and male (+/y and -/y) mice by using the same probe used for Southern blot analysis. The ethidium bromide-staining 28S and 18S bands are also indicated.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Disruption of forkhead transcription factor (FOXO) family members in mice reveals their functional diversification

    doi: 10.1073/pnas.0400093101

    Figure Lengend Snippet: Targeted disruption of the Foxo3a gene and Foxo4 gene. ( A ) Targeting strategy for Foxo3a . A retroviral promoter trap vector was inserted in intron 1 of Foxo3a . SA, splice acceptor site; SD, splice donor site; LTR, long terminal repeat sequence; pA, polyadenylation signal; puro, puromycin. ( B ) Southern analysis of Sac I-digested genomic DNA obtained from a mating of Foxo3a +/- mice by using the Neo - and Csk -specific probes. Different intensities demonstrated by hybridization of the Neo-specific probe discriminate zero, one, or two Foxo3a gene disruptions, representing +/+, +/-, and -/- mice, respectively. ( C ) Western analysis of lung tissue lysates from Foxo3a +/+ and Foxo3a -/- mice by using antibodies against Foxo3a and β-actin. ( D ) Restriction maps of the wild-type Foxo4 locus, the targeting vector, and the targeted locus. The targeting vector contained a PGK-Neo cassette instead of the Foxo4 exon1. A targeting probe (3′ region of Foxo4 cDNA) was also indicated. Restriction enzymes were indicated as B, Bam HI; C, Cla I; E, Eco RI; H, Hin dIII; N, Nco I; and Sp, Spe I. ( E ) Southern analysis of Bam HI-digested genomic DNA obtained from Foxo4 female (+/+, +/-, and -/-) and male (+/y and -/y) mice by using a probe from the 3′ region of Foxo4 cDNA. The Foxo4 wild-type allele (10 kb) and targeted allele (16 kb) were indicated. ( F ) Northern analysis of total RNA from skeletal muscle of Foxo4 female (+/+ and -/-) and male (+/y and -/y) mice by using the same probe used for Southern blot analysis. The ethidium bromide-staining 28S and 18S bands are also indicated.

    Article Snippet: Antibodies used for Western analysis were polyclonal anti-FKHR antibody (C-20, Santa Cruz Biotechnology), polyclonal anti-FKHRL1 antibody (Upstate Biotechnology, Waltham, MA), monoclonal anti-β-actin antibody (clone AC-15, Sigma), and horseradish peroxidase (HRP)-conjugated secondary antibodies (Dako).

    Techniques: Plasmid Preparation, Sequencing, Mouse Assay, Hybridization, Western Blot, Hi-C, Northern Blot, Southern Blot, Staining