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  • 92
    Millipore pflag cytomegalovirus cmv
    FcRL4 and FcRL5 bind human immunoglobulins HEK293T cells were transiently transfected with human cDNA for FcRL1, 3, 4, 5, 6 and CD200R expressed in <t>pFLAG-CMV-3,</t> or CD32 in pEF6. Binding of heat-aggregated human serum IgG and IgA (Y-axis) to cells expressing human FcRL4 and FcRL5 was easily observable by FACS. Cell surface expression of each protein was confirmed using FITC-conjugated anti-FLAG M2 monoclonal antibody (X-axis), or by staining with anti-CD32 antibody. Binding to CD200R and CD32 are included as negative and positive controls respectively. No binding of human IgM to any of the FcRL proteins was observed (not shown).
    Pflag Cytomegalovirus Cmv, supplied by Millipore, used in various techniques. Bioz Stars score: 92/100, based on 27 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Millipore pflag cmv
    RGR-d protein expression in cultured human fetal RPE cells. Immunoblots of whole cell proteins were probed with the HRGR-DE7 antibody. The analysis included polarized human fetal RPE cells from different samples: ( A ) donor HFRPE-A and ( B ) donors HFRPE-B and HFRPE-C. A specific protein band that corresponds in size to RGR-d was found in each donor RPE cell. Cell extracts with FLAG-RGR and FLAG-RGR-d fusion proteins were used both for size comparison and as protein controls. <t>pFLAG-hRGR,</t> pFLAG-hRGR-d, and <t>pFLAG-CMV-4</t> control expression vectors were transfected into ARPE-19 cells. Untreated ARPE-19 cells did not express detectable levels of endogenous RGR or RGR-d proteins.
    Pflag Cmv, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 159 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Millipore pflag cmv 6
    RGR-d protein expression in cultured human fetal RPE cells. Immunoblots of whole cell proteins were probed with the HRGR-DE7 antibody. The analysis included polarized human fetal RPE cells from different samples: ( A ) donor HFRPE-A and ( B ) donors HFRPE-B and HFRPE-C. A specific protein band that corresponds in size to RGR-d was found in each donor RPE cell. Cell extracts with FLAG-RGR and FLAG-RGR-d fusion proteins were used both for size comparison and as protein controls. <t>pFLAG-hRGR,</t> pFLAG-hRGR-d, and <t>pFLAG-CMV-4</t> control expression vectors were transfected into ARPE-19 cells. Untreated ARPE-19 cells did not express detectable levels of endogenous RGR or RGR-d proteins.
    Pflag Cmv 6, supplied by Millipore, used in various techniques. Bioz Stars score: 85/100, based on 28 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Millipore pflag cmv 14
    RGR-d protein expression in cultured human fetal RPE cells. Immunoblots of whole cell proteins were probed with the HRGR-DE7 antibody. The analysis included polarized human fetal RPE cells from different samples: ( A ) donor HFRPE-A and ( B ) donors HFRPE-B and HFRPE-C. A specific protein band that corresponds in size to RGR-d was found in each donor RPE cell. Cell extracts with FLAG-RGR and FLAG-RGR-d fusion proteins were used both for size comparison and as protein controls. <t>pFLAG-hRGR,</t> pFLAG-hRGR-d, and <t>pFLAG-CMV-4</t> control expression vectors were transfected into ARPE-19 cells. Untreated ARPE-19 cells did not express detectable levels of endogenous RGR or RGR-d proteins.
    Pflag Cmv 14, supplied by Millipore, used in various techniques. Bioz Stars score: 85/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore pflag cmv 6c
    FAT10 and Vpr coimmunoprecipitate and colocalize to mitochondria. (A) HEK293T cells were cotransfected with pcDNA4-FAT10 and either <t>pFLAG-Vpr</t> or <t>pFLAG-CMV-6c</t> (FLAG control). Cell lysate was immunoprecipitated with anti-FLAG and immunoblotted using anti-FLAG and anti-FAT10. FAT10 was detected in immunoprecipitates from cells expressing FLAG-Vpr but not FLAG alone. Results were confirmed in a reciprocal experiment in which pFLAG-FAT10 or p-FLAG-CMV-6c (FLAG control) was cotransfected with pNL4-3:ΔG/P-EGFP. Immunoprecipitation of cell lysate using anti-FLAG and subsequent immunoblotting using anti-FLAG and anti-Vpr demonstrated that Vpr was present in immunoprecipitate from cells transfected FLAG-FAT10 but not the FLAG control. (B) HEK293T cells were cotransfected with pGFP-Vpr or pHR (GFP control) and pFLAG-FAT10. GFP-Vpr and FLAG-FAT10 colocalized with MitotrackerCMXRos, primarily in large aggregates. (C) Western blotting of mitochondrial and cytoplasmic protein from HEK293T cells after cotransfection with pHA-Vpr and pFLAG-FAT10 demonstrated that FAT10 and Vpr in were present mitochondrial and cytoplasmic fractions. Blots were stripped and reprobed with anti-COX4 and anti-β-actin to confirm the purity of mitochondrial fractions.
    Pflag Cmv 6c, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 60 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    98
    Millipore pflag cmv 2
    Mapping of the binding site for LOX-PP on Hsp70. (A) A schematic representation of the Hsp70 mutants used in this study is shown. Full-length (WT) or deletion mutant Hsp70 cDNAs (ΔBglII and ΔSmaI) were inserted into the <t>pFLAG-CMV-2</t> vector.
    Pflag Cmv 2, supplied by Millipore, used in various techniques. Bioz Stars score: 98/100, based on 781 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Millipore pflag cmv 5a
    Mapping of the binding site for LOX-PP on Hsp70. (A) A schematic representation of the Hsp70 mutants used in this study is shown. Full-length (WT) or deletion mutant Hsp70 cDNAs (ΔBglII and ΔSmaI) were inserted into the <t>pFLAG-CMV-2</t> vector.
    Pflag Cmv 5a, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 150 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Millipore pflag cmv 5b
    Mapping of the binding site for LOX-PP on Hsp70. (A) A schematic representation of the Hsp70 mutants used in this study is shown. Full-length (WT) or deletion mutant Hsp70 cDNAs (ΔBglII and ΔSmaI) were inserted into the <t>pFLAG-CMV-2</t> vector.
    Pflag Cmv 5b, supplied by Millipore, used in various techniques. Bioz Stars score: 85/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Millipore pflag cmv 1
    Mapping of the binding site for LOX-PP on Hsp70. (A) A schematic representation of the Hsp70 mutants used in this study is shown. Full-length (WT) or deletion mutant Hsp70 cDNAs (ΔBglII and ΔSmaI) were inserted into the <t>pFLAG-CMV-2</t> vector.
    Pflag Cmv 1, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 101 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Millipore pflag myc cmv
    Mapping of the binding site for LOX-PP on Hsp70. (A) A schematic representation of the Hsp70 mutants used in this study is shown. Full-length (WT) or deletion mutant Hsp70 cDNAs (ΔBglII and ΔSmaI) were inserted into the <t>pFLAG-CMV-2</t> vector.
    Pflag Myc Cmv, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Millipore pflag cmv 5a vector
    Mapping of the binding site for LOX-PP on Hsp70. (A) A schematic representation of the Hsp70 mutants used in this study is shown. Full-length (WT) or deletion mutant Hsp70 cDNAs (ΔBglII and ΔSmaI) were inserted into the <t>pFLAG-CMV-2</t> vector.
    Pflag Cmv 5a Vector, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 71 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    FcRL4 and FcRL5 bind human immunoglobulins HEK293T cells were transiently transfected with human cDNA for FcRL1, 3, 4, 5, 6 and CD200R expressed in pFLAG-CMV-3, or CD32 in pEF6. Binding of heat-aggregated human serum IgG and IgA (Y-axis) to cells expressing human FcRL4 and FcRL5 was easily observable by FACS. Cell surface expression of each protein was confirmed using FITC-conjugated anti-FLAG M2 monoclonal antibody (X-axis), or by staining with anti-CD32 antibody. Binding to CD200R and CD32 are included as negative and positive controls respectively. No binding of human IgM to any of the FcRL proteins was observed (not shown).

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    Article Title: Human FcRL4 and FcRL5 are receptors for IgA and IgG FcRL4 and FcRL5 are receptors for IgA and IgG

    doi: 10.4049/jimmunol.1102651

    Figure Lengend Snippet: FcRL4 and FcRL5 bind human immunoglobulins HEK293T cells were transiently transfected with human cDNA for FcRL1, 3, 4, 5, 6 and CD200R expressed in pFLAG-CMV-3, or CD32 in pEF6. Binding of heat-aggregated human serum IgG and IgA (Y-axis) to cells expressing human FcRL4 and FcRL5 was easily observable by FACS. Cell surface expression of each protein was confirmed using FITC-conjugated anti-FLAG M2 monoclonal antibody (X-axis), or by staining with anti-CD32 antibody. Binding to CD200R and CD32 are included as negative and positive controls respectively. No binding of human IgM to any of the FcRL proteins was observed (not shown).

    Article Snippet: In order to test for Ig binding by FcRL family members, cDNA encoding mouse FcRH3/FcRL5, human CD200R, FcRL1, FcRL3, FcRL4, FcRL5 (Open Biosystems), and FcRL6( ) were ligated into pFLAG-CMV-3 (Sigma, St. Louis, MO). cDNA encoding human CD32 was ligated into pEF6 (Invitrogen).

    Techniques: Transfection, Binding Assay, Expressing, FACS, Staining

    RGR-d protein expression in cultured human fetal RPE cells. Immunoblots of whole cell proteins were probed with the HRGR-DE7 antibody. The analysis included polarized human fetal RPE cells from different samples: ( A ) donor HFRPE-A and ( B ) donors HFRPE-B and HFRPE-C. A specific protein band that corresponds in size to RGR-d was found in each donor RPE cell. Cell extracts with FLAG-RGR and FLAG-RGR-d fusion proteins were used both for size comparison and as protein controls. pFLAG-hRGR, pFLAG-hRGR-d, and pFLAG-CMV-4 control expression vectors were transfected into ARPE-19 cells. Untreated ARPE-19 cells did not express detectable levels of endogenous RGR or RGR-d proteins.

    Journal: Molecular Vision

    Article Title: Targeting of exon VI-skipping human RGR-opsin to the plasma membrane of pigment epithelium and co-localization with terminal complement complex C5b-9

    doi:

    Figure Lengend Snippet: RGR-d protein expression in cultured human fetal RPE cells. Immunoblots of whole cell proteins were probed with the HRGR-DE7 antibody. The analysis included polarized human fetal RPE cells from different samples: ( A ) donor HFRPE-A and ( B ) donors HFRPE-B and HFRPE-C. A specific protein band that corresponds in size to RGR-d was found in each donor RPE cell. Cell extracts with FLAG-RGR and FLAG-RGR-d fusion proteins were used both for size comparison and as protein controls. pFLAG-hRGR, pFLAG-hRGR-d, and pFLAG-CMV-4 control expression vectors were transfected into ARPE-19 cells. Untreated ARPE-19 cells did not express detectable levels of endogenous RGR or RGR-d proteins.

    Article Snippet: The sense strand primer was designed to exclude the start codons of the RGR and RGR-d pcDNA3 templates since the start codon of the pFLAG-CMV-4 vector (Sigma-Aldrich, St. Louis, MO) was to be employed.

    Techniques: Expressing, Cell Culture, Western Blot, Transfection

    FAT10 and Vpr coimmunoprecipitate and colocalize to mitochondria. (A) HEK293T cells were cotransfected with pcDNA4-FAT10 and either pFLAG-Vpr or pFLAG-CMV-6c (FLAG control). Cell lysate was immunoprecipitated with anti-FLAG and immunoblotted using anti-FLAG and anti-FAT10. FAT10 was detected in immunoprecipitates from cells expressing FLAG-Vpr but not FLAG alone. Results were confirmed in a reciprocal experiment in which pFLAG-FAT10 or p-FLAG-CMV-6c (FLAG control) was cotransfected with pNL4-3:ΔG/P-EGFP. Immunoprecipitation of cell lysate using anti-FLAG and subsequent immunoblotting using anti-FLAG and anti-Vpr demonstrated that Vpr was present in immunoprecipitate from cells transfected FLAG-FAT10 but not the FLAG control. (B) HEK293T cells were cotransfected with pGFP-Vpr or pHR (GFP control) and pFLAG-FAT10. GFP-Vpr and FLAG-FAT10 colocalized with MitotrackerCMXRos, primarily in large aggregates. (C) Western blotting of mitochondrial and cytoplasmic protein from HEK293T cells after cotransfection with pHA-Vpr and pFLAG-FAT10 demonstrated that FAT10 and Vpr in were present mitochondrial and cytoplasmic fractions. Blots were stripped and reprobed with anti-COX4 and anti-β-actin to confirm the purity of mitochondrial fractions.

    Journal: Journal of Virology

    Article Title: FAT10: a Novel Mediator of Vpr-Induced Apoptosis in Human Immunodeficiency Virus-Associated Nephropathy ▿

    doi: 10.1128/JVI.00034-09

    Figure Lengend Snippet: FAT10 and Vpr coimmunoprecipitate and colocalize to mitochondria. (A) HEK293T cells were cotransfected with pcDNA4-FAT10 and either pFLAG-Vpr or pFLAG-CMV-6c (FLAG control). Cell lysate was immunoprecipitated with anti-FLAG and immunoblotted using anti-FLAG and anti-FAT10. FAT10 was detected in immunoprecipitates from cells expressing FLAG-Vpr but not FLAG alone. Results were confirmed in a reciprocal experiment in which pFLAG-FAT10 or p-FLAG-CMV-6c (FLAG control) was cotransfected with pNL4-3:ΔG/P-EGFP. Immunoprecipitation of cell lysate using anti-FLAG and subsequent immunoblotting using anti-FLAG and anti-Vpr demonstrated that Vpr was present in immunoprecipitate from cells transfected FLAG-FAT10 but not the FLAG control. (B) HEK293T cells were cotransfected with pGFP-Vpr or pHR (GFP control) and pFLAG-FAT10. GFP-Vpr and FLAG-FAT10 colocalized with MitotrackerCMXRos, primarily in large aggregates. (C) Western blotting of mitochondrial and cytoplasmic protein from HEK293T cells after cotransfection with pHA-Vpr and pFLAG-FAT10 demonstrated that FAT10 and Vpr in were present mitochondrial and cytoplasmic fractions. Blots were stripped and reprobed with anti-COX4 and anti-β-actin to confirm the purity of mitochondrial fractions.

    Article Snippet: To express FLAG-tagged Vpr, we created pFLAG-CMV-6c-Vpr and pFLAG-CMV-6c-FAT10 by subcloning Vpr and FAT10 into pFLAG-CMV-6c (Sigma). pFLAG-CMV-6c-Vpr or pFLAG-CMV-6c (FLAG control) was cotransfected with pcDNA4-FAT10 ( ) into HEK293T cells, and cellular lysate was immunoprecipitated using anti-FLAG.

    Techniques: Immunoprecipitation, Expressing, Transfection, Western Blot, Cotransfection

    Mapping of the binding site for LOX-PP on Hsp70. (A) A schematic representation of the Hsp70 mutants used in this study is shown. Full-length (WT) or deletion mutant Hsp70 cDNAs (ΔBglII and ΔSmaI) were inserted into the pFLAG-CMV-2 vector.

    Journal: Molecular and Cellular Biology

    Article Title: The Ras Signaling Inhibitor LOX-PP Interacts with Hsp70 and c-Raf To Reduce Erk Activation and Transformed Phenotype of Breast Cancer Cells ▿

    doi: 10.1128/MCB.01148-10

    Figure Lengend Snippet: Mapping of the binding site for LOX-PP on Hsp70. (A) A schematic representation of the Hsp70 mutants used in this study is shown. Full-length (WT) or deletion mutant Hsp70 cDNAs (ΔBglII and ΔSmaI) were inserted into the pFLAG-CMV-2 vector.

    Article Snippet: The cDNA encoding the Hsp70 wild type (WT) was inserted into pEBG, pGEX4T-3 (GE Healthcare, Uppsala, Sweden), and pFLAG-CMV-2 (Sigma-Aldrich, St. Louis, MO).

    Techniques: Binding Assay, Mutagenesis, Plasmid Preparation

    WEE1Hu phosphorylation at Ser 642 during late S to G 2 phase. 293T cells were transfected with the pFLAG-CMV-2 vector encoding ΔN214-WEE1Hu. After transfection for 24 h, the cells were treated with 1 μg/ml of aphidicolin for 24 h. Then, the cells were washed with PBS and changed to complete growth medium. Cells were harvested at the indicated time after the release. One-third of the cells were analyzed using a flow cytometer (right panel), and the others were analyzed by immunoprecipitation with an anti-FLAG agarose, following immunoblot analysis with an anti-phospho-Ser/Thr Akt substrate antibody or an anti-FLAG antibody (left, top and second panels, respectively). The expression level of endogenous WEE1Hu (endo-WEE1Hu), phosphorylated Cdc2 (Tyr 15 ), Cdc2, cyclin B1, cyclin A, and β-actin was confirmed by immunoblot analysis with the indicated antibodies (left, from third to bottom panels).

    Journal: Molecular and Cellular Biology

    Article Title: Akt/Protein Kinase B-Dependent Phosphorylation and Inactivation of WEE1Hu Promote Cell Cycle Progression at G2/M Transition

    doi: 10.1128/MCB.25.13.5725-5737.2005

    Figure Lengend Snippet: WEE1Hu phosphorylation at Ser 642 during late S to G 2 phase. 293T cells were transfected with the pFLAG-CMV-2 vector encoding ΔN214-WEE1Hu. After transfection for 24 h, the cells were treated with 1 μg/ml of aphidicolin for 24 h. Then, the cells were washed with PBS and changed to complete growth medium. Cells were harvested at the indicated time after the release. One-third of the cells were analyzed using a flow cytometer (right panel), and the others were analyzed by immunoprecipitation with an anti-FLAG agarose, following immunoblot analysis with an anti-phospho-Ser/Thr Akt substrate antibody or an anti-FLAG antibody (left, top and second panels, respectively). The expression level of endogenous WEE1Hu (endo-WEE1Hu), phosphorylated Cdc2 (Tyr 15 ), Cdc2, cyclin B1, cyclin A, and β-actin was confirmed by immunoblot analysis with the indicated antibodies (left, from third to bottom panels).

    Article Snippet: Human wild-type and dominant-negative (AAA, K179A/T308A/S473A) akt1 cDNAs in a pFLAG-CMV-2 vector (Sigma) or a pHM6 vector (Roche Molecular Biochemicals, Mannheim, Germany), wild-type and phosphatase-inactive (C124S) PTEN in a pFLAG-CMV-2 vector, 14-3-3θ in a pFLAG-CMV-2 vector or a pHM6 vector, and aggrus (also known as podoplanin or T1α ) in a pcDNA3 vector were established in our laboratory (15-17, 24).

    Techniques: Transfection, Plasmid Preparation, Flow Cytometry, Cytometry, Immunoprecipitation, Expressing

    Overexpression of Akt together with 14-3-3θ overcomes the WEE1Hu-induced G 2 /M arrest. (A) 293T cells were cotransfected with the pFLAG-CMV-2 vector alone (a) or encoding wild-type WEE1Hu (b to e) and the pHM6 vector alone (a to c) or encoding 14-3-3θ (d and e) together with the pUSEamp vector not encoding Myr-Akt (a, b, and d) or encoding Myr-Akt (c and e). To monitor the transfected cells, all cells were simultaneously transfected with the pcDNA3 vector encoding Aggrus as a cell surface marker. The cells were harvested 48 h posttransfection. The Aggrus proteins were detected by staining with a rat monoclonal anti-Aggrus antibody and a fluorescein isothiocyanate-conjugated anti-rat antibody. Nuclei were also detected by staining with propidium iodide, and the cells were analyzed using a flow cytometer. (B) 293T cells were transfected as for panel A. Cell lysates were subjected to immunoblot analysis with the indicated antibodies.

    Journal: Molecular and Cellular Biology

    Article Title: Akt/Protein Kinase B-Dependent Phosphorylation and Inactivation of WEE1Hu Promote Cell Cycle Progression at G2/M Transition

    doi: 10.1128/MCB.25.13.5725-5737.2005

    Figure Lengend Snippet: Overexpression of Akt together with 14-3-3θ overcomes the WEE1Hu-induced G 2 /M arrest. (A) 293T cells were cotransfected with the pFLAG-CMV-2 vector alone (a) or encoding wild-type WEE1Hu (b to e) and the pHM6 vector alone (a to c) or encoding 14-3-3θ (d and e) together with the pUSEamp vector not encoding Myr-Akt (a, b, and d) or encoding Myr-Akt (c and e). To monitor the transfected cells, all cells were simultaneously transfected with the pcDNA3 vector encoding Aggrus as a cell surface marker. The cells were harvested 48 h posttransfection. The Aggrus proteins were detected by staining with a rat monoclonal anti-Aggrus antibody and a fluorescein isothiocyanate-conjugated anti-rat antibody. Nuclei were also detected by staining with propidium iodide, and the cells were analyzed using a flow cytometer. (B) 293T cells were transfected as for panel A. Cell lysates were subjected to immunoblot analysis with the indicated antibodies.

    Article Snippet: Human wild-type and dominant-negative (AAA, K179A/T308A/S473A) akt1 cDNAs in a pFLAG-CMV-2 vector (Sigma) or a pHM6 vector (Roche Molecular Biochemicals, Mannheim, Germany), wild-type and phosphatase-inactive (C124S) PTEN in a pFLAG-CMV-2 vector, 14-3-3θ in a pFLAG-CMV-2 vector or a pHM6 vector, and aggrus (also known as podoplanin or T1α ) in a pcDNA3 vector were established in our laboratory (15-17, 24).

    Techniques: Over Expression, Plasmid Preparation, Transfection, Marker, Staining, Flow Cytometry, Cytometry