pp2 Millipore Search Results


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  • 95
    Millipore pp2
    Caveolin-1 and EGFR co-localization as a feature of non-canonical EGFR activation. Epithelial cells (RLE-6TN) were exposed (5 min) to carbon nanoparticles (CNP), non-nano carbon particles (CP), each 10 µg/cm 2 , (50 µM) H 2 O 2 , (5 µM) C6 ceramide, or EGF (100 ng/mL), respectively. ( A ) Subcellular localization of EGFR (red Alexa flour 594) and caveolin-1 (green Alexa flour 488). Co-localization is visualized by the yellow color in merged images; ( B ) Subcellular localization of EGFR and caveolin-1 in cells pre-treated with inhibitors of carbon nanoparticle-specific signaling prior to particle or EGF exposure: SFK inhibitor <t>PP2</t> (10 µM), 1 mM ectoine ( E ), 75 µM α-tocopherol (Toco), and 1 µg/mL filipin III (Fil). Co-localization is visualized by the yellow color in merged images; ( C ) Subcellular localization of caveolin-1 (red Alexa fluor 594) after exposure to carbon nanoparticles (CNP), carbon particles (CP), or hydrogen peroxide (H 2 O 2 ); ( D ) Quantification and representative Western blots of caveolin-1 in lipid raft fraction of RLE-6TN cells exposed to CNP (10 µg/cm 2 ). Raft and non-raft fraction were isolated from density gradients after ultracentrifugation. Pre-treatment of cells with α-tocopherol (Toco) was applied as an antioxidant strategy. GAPDH was used as a control protein not associated with lipid rafts. The bars in the graph represent the additive immune signals of raft and non-raft fractions, which was indicated in the representative original Western blots; ( E ) Quantification and representative Western-blot of EGFR phosphorylation at Tyr 845 . Nuclei were stained with DAPI (blue). Scale bars represent 20 µm; *, which was significantly different to PBS control ( p
    Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 1801 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Millipore src pp2
    Caveolin-1 and EGFR co-localization as a feature of non-canonical EGFR activation. Epithelial cells (RLE-6TN) were exposed (5 min) to carbon nanoparticles (CNP), non-nano carbon particles (CP), each 10 µg/cm 2 , (50 µM) H 2 O 2 , (5 µM) C6 ceramide, or EGF (100 ng/mL), respectively. ( A ) Subcellular localization of EGFR (red Alexa flour 594) and caveolin-1 (green Alexa flour 488). Co-localization is visualized by the yellow color in merged images; ( B ) Subcellular localization of EGFR and caveolin-1 in cells pre-treated with inhibitors of carbon nanoparticle-specific signaling prior to particle or EGF exposure: SFK inhibitor <t>PP2</t> (10 µM), 1 mM ectoine ( E ), 75 µM α-tocopherol (Toco), and 1 µg/mL filipin III (Fil). Co-localization is visualized by the yellow color in merged images; ( C ) Subcellular localization of caveolin-1 (red Alexa fluor 594) after exposure to carbon nanoparticles (CNP), carbon particles (CP), or hydrogen peroxide (H 2 O 2 ); ( D ) Quantification and representative Western blots of caveolin-1 in lipid raft fraction of RLE-6TN cells exposed to CNP (10 µg/cm 2 ). Raft and non-raft fraction were isolated from density gradients after ultracentrifugation. Pre-treatment of cells with α-tocopherol (Toco) was applied as an antioxidant strategy. GAPDH was used as a control protein not associated with lipid rafts. The bars in the graph represent the additive immune signals of raft and non-raft fractions, which was indicated in the representative original Western blots; ( E ) Quantification and representative Western-blot of EGFR phosphorylation at Tyr 845 . Nuclei were stained with DAPI (blue). Scale bars represent 20 µm; *, which was significantly different to PBS control ( p
    Src Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 90/100, based on 30 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    77
    Millipore well validated inhibitor pp2
    Quantitative analysis of radial maturation. A , Schema of the five boxes encompassing the outer (1) and inner (2) tiers of the cortical ribbon (excluding the meninges), the outer (3) and inner (4) tiers of the intermediate zone of migration, and the ventricular–subventricular zones (5) in which BrdU- or THY-positive neurons were counted. B , Between 300–400 BrdU- and 300–400 THY-labeled cells were counted in four different sections from normal, reeler, <t>PP2-,</t> and BIM1-treated slices, and the proportion in boxes 1–5 was graphed in gray (BrdU) and black (THY). Boxes 1 and 2 defined in the cortical plate are the most relevant, because the proportion of labeled cells in the deep boxes varied more from experiment to experiment.
    Well Validated Inhibitor Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 77/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore sfk inhibitor pp2
    Src-family kinase signaling, but not Syk or PI3K signaling, is indispensable for reorganization of macrophage surfaces. (A) Immunoblots of phosphorylated AKT in nonactivated (PLL) or hIgG-activated human macrophages pretreated with vehicle (DMSO), as a control, 10 µM <t>PP2</t> (left), 100 µM piceatannol (PCT; middle), or 1 µM wortmannin (Wort; right). Blots represent two independent experiments. (B) TIRF image of FcγRI (white; bars, 20 µm) and dSTORM images (bars, 5 µm) of FcγRI (green) and SIRPα (red) at the surface of human macrophages incubated with vehicle (DMSO), PP2, PCT, or Wort, pretreated as in A. Cells were then seeded onto slides coated with PLL (nonactivated) or hIgG for 10 min and stained with anti–FcγRI-AF488 and anti–SIRPα-AF647 mAbs. In each condition, regions outlined by the white squares (middle column) are shown enlarged (right column). Bars, 1 µm. (C) CBC histograms for FcγRI and SIRPα in cells pretreated as in A and seeded onto slides coated with PLL (gray) or hIgG (DMSO, dark gray; PP2, red; PCT, green; and Wort, blue) for 10 min, as indicated. Data are from a minimum of 30 cells from three independent donors. Bars show mean ± SD. (D) NND analysis from data shown in C. Each symbol represents the median NND of all paired single-molecule localizations from one cell. Horizontal lines and error bars represent mean ± SD. ns, not significant; ****, P
    Sfk Inhibitor Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 79 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    76
    Millipore src family kinase inhibitor pp2 4 amino 5 4 chlorophenyl 7 t butyl pyrazololo 3 4 d pyrimidine
    Src-family kinase signaling, but not Syk or PI3K signaling, is indispensable for reorganization of macrophage surfaces. (A) Immunoblots of phosphorylated AKT in nonactivated (PLL) or hIgG-activated human macrophages pretreated with vehicle (DMSO), as a control, 10 µM <t>PP2</t> (left), 100 µM piceatannol (PCT; middle), or 1 µM wortmannin (Wort; right). Blots represent two independent experiments. (B) TIRF image of FcγRI (white; bars, 20 µm) and dSTORM images (bars, 5 µm) of FcγRI (green) and SIRPα (red) at the surface of human macrophages incubated with vehicle (DMSO), PP2, PCT, or Wort, pretreated as in A. Cells were then seeded onto slides coated with PLL (nonactivated) or hIgG for 10 min and stained with anti–FcγRI-AF488 and anti–SIRPα-AF647 mAbs. In each condition, regions outlined by the white squares (middle column) are shown enlarged (right column). Bars, 1 µm. (C) CBC histograms for FcγRI and SIRPα in cells pretreated as in A and seeded onto slides coated with PLL (gray) or hIgG (DMSO, dark gray; PP2, red; PCT, green; and Wort, blue) for 10 min, as indicated. Data are from a minimum of 30 cells from three independent donors. Bars show mean ± SD. (D) NND analysis from data shown in C. Each symbol represents the median NND of all paired single-molecule localizations from one cell. Horizontal lines and error bars represent mean ± SD. ns, not significant; ****, P
    Src Family Kinase Inhibitor Pp2 4 Amino 5 4 Chlorophenyl 7 T Butyl Pyrazololo 3 4 D Pyrimidine, supplied by Millipore, used in various techniques. Bioz Stars score: 76/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    79
    Millipore pp2 kinase inhibitor
    Antitumor effects of Src inhibitor <t>PP2</t> on U251 glioma cells ( A ) Cytotoxic effects of PP2 by MTT assays 24 h after treatment. ( B ) Decreased phosphorylation of Src, Akt, and STAT3 in PP2-treated U251 glioma cells by immunoblots. ( C ) Decrease in NF-κB activity in U251 glioma cells after PP2 by luciferase reporter gene assays. U251 glioma cells were transfected with NF-κB-Luc and β-galactosidase for 24 h before ARP treatment. ( D ) Suppressed migration of U251 glioma cells following PP2 treatment determined by wound-healing assays. * P
    Pp2 Kinase Inhibitor, supplied by Millipore, used in various techniques. Bioz Stars score: 79/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    77
    Millipore csrc pp2
    Lipid raft preparations demonstrating increased the presence of p-PKC α , phosphorylated and total caveolin-1 (p-Cav-1 and t-Cav-1, respectively) and phosphorylated <t>cSrc</t> (p-cSrc) in the same raft fractions 1 h after viral infection (V) as compared to mock infection (M), and the effect of chemical inhibition of cSrc by <t>PP2</t> (A) and (B), and of PKC α by calphostin C (Cal.C) in (C). (A) Western blot for p-PKC α shows increased signal upon viral infection, with slight reduction upon PP2 treatment (10 µ M, or DMSO control). (B) Western blot for p-Cav-1, with or without pretreatment by PP2 (or DMSO control), stripped and reprobed for p-cSrc and then stripped and reprobed for t-Cav-1, shows increased p-Cav-1 and p-cSrc in the same lipid raft fractions with a reduction in phosphorylation for both in cells pretreated with PP2. t-Cav-1 in the lipid raft fractions was also slightly reduced by PP2 pretreatment. (C) Western blots on cells pretreated with either DMSO control or calphostin C (1 µ M) for 3 h prior to mock or virus infection, and stripped and reprobed as in (B), show a dramatic reduction in p-Cav-1 but not t-Cav-1, and complete abrogation of p-cSrc expression by calphostin C in the same lipid raft fractions.
    Csrc Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 77/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    77
    Millipore pki pp2
    Lipid raft preparations demonstrating increased the presence of p-PKC α , phosphorylated and total caveolin-1 (p-Cav-1 and t-Cav-1, respectively) and phosphorylated <t>cSrc</t> (p-cSrc) in the same raft fractions 1 h after viral infection (V) as compared to mock infection (M), and the effect of chemical inhibition of cSrc by <t>PP2</t> (A) and (B), and of PKC α by calphostin C (Cal.C) in (C). (A) Western blot for p-PKC α shows increased signal upon viral infection, with slight reduction upon PP2 treatment (10 µ M, or DMSO control). (B) Western blot for p-Cav-1, with or without pretreatment by PP2 (or DMSO control), stripped and reprobed for p-cSrc and then stripped and reprobed for t-Cav-1, shows increased p-Cav-1 and p-cSrc in the same lipid raft fractions with a reduction in phosphorylation for both in cells pretreated with PP2. t-Cav-1 in the lipid raft fractions was also slightly reduced by PP2 pretreatment. (C) Western blots on cells pretreated with either DMSO control or calphostin C (1 µ M) for 3 h prior to mock or virus infection, and stripped and reprobed as in (B), show a dramatic reduction in p-Cav-1 but not t-Cav-1, and complete abrogation of p-cSrc expression by calphostin C in the same lipid raft fractions.
    Pki Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 77/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore src kinase inhibitor pp2
    Effect of IL-1RA, <t>PP2</t> or 3OMS on IL-1β-induced <t>Src</t> kinase in primary cortical neurones. Cultures were treated with vehicle (C) or IL-1β (0.3 U ml −1 ) for 5, 15, 30 or 60 min, in the absence or presence of
    Src Kinase Inhibitor Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 173 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    83
    Millipore sfks inhibitor pp2
    <t>SFKs</t> are necessary for TLR signalling events in human pDCs. ( a – f ) WT or LYN − ( d – f ) CAL-1 cells were pre-treated for 1 h with <t>PP2</t> (10 μM; a – c ; blue) or DMSO control ( a – c ; black) or left untreated ( d – f ; black: LYN + , orange: LYN − ), before stimulation with R848 for the indicated time periods (minutes). Level of p-IKKα/β and total IKKα/β ( a , d ) as well as IκBα ( b , e ) and GAPDH were assessed by immunoblot. p-NF-κB was determined by flow cytometry. The histogram depicts the 15 min time point ( c , f ). ( g , h ) Human PBMCs were pre-treated as in c and stimulated for 15 min with R848. p-NF-κB ( g ) and p-IRF-7 ( h ) were determined by flow cytometry in gated pDCs. Lines in graphs connect the same donor and histograms depict one representative donor; dotted lines, unstimulated; solid lines, R848 stimulation. ( i ) CAL-1 cells were pre-treated as in c before stimulation with recombinant IFN-β (1,000 U ml −1 ) for the indicated time periods (minutes). p-STAT1 were determined by flow cytometry. Histogram depicts the 30 min time point. Data are representative of two ( b , e , f , i ) or three ( a , c , d ) independent experiments and four ( g ) or five ( h ) donors processed separately. Graphs depict mean±s.d. of replicates within one representative experiment ( a – f , i ) or individual donors ( g , h ). Two-way ANOVA ( c , f , i ) and two-ways Student's t -test ( g , h ) were used for statistical analyses. * P
    Sfks Inhibitor Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 83/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore src inhibitor
    Effect of tumor primary culture (TPC) conditioned medium (CM) and leukemia inhibitory factor (LIF)-neutralizing antibody. (a) Stat3 (signal transduction and activators of transcription 3) and phospho-Stat3 (p-Stat3) in HC11 and LM3 cells treated with increasing concentrations of CM (30%, 50% and 80%). (b) Stat3 and p-Stat3 levels in HC11 and TPC cells treated with CM that had been preincubated with or without LIF-neutralizing antibody. (c) Phosphorylation levels of Stat3 and extracellular signal-regulated kinase (ERK)1/2 in HC11 cells treated with LIF (3 and 5 ng/ml) preincubated with or without LIF-neutralizing antibody. (d) Expression of CCAAT-enhancer-binding protein (C/EBP)δ in TPC cells treated with CM with or without neutralizing antibody. (e) Phosphorylation of Stat3 and ERK1/2 in HC11 cells treated with LIF with or without the MAP kinase/ERK kinase inhibitor PD98059. (f) p-Stat3 and Stat3 in HC11 and TPC cells treated with LIF with or without the <t>Src-specific</t> inhibitor <t>PP2.</t> Experiments were repeated at least three times with similar results. No effect was observed on phosphorylation levels of either Stat3 or ERK1/2 when HC11 cells were treated with the PD98059 vehicle, dimethyl sulfoxide (data not shown).
    Src Inhibitor, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 127 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    78
    Millipore sfk inhibitor proteinphosphatase 2
    Effect of tumor primary culture (TPC) conditioned medium (CM) and leukemia inhibitory factor (LIF)-neutralizing antibody. (a) Stat3 (signal transduction and activators of transcription 3) and phospho-Stat3 (p-Stat3) in HC11 and LM3 cells treated with increasing concentrations of CM (30%, 50% and 80%). (b) Stat3 and p-Stat3 levels in HC11 and TPC cells treated with CM that had been preincubated with or without LIF-neutralizing antibody. (c) Phosphorylation levels of Stat3 and extracellular signal-regulated kinase (ERK)1/2 in HC11 cells treated with LIF (3 and 5 ng/ml) preincubated with or without LIF-neutralizing antibody. (d) Expression of CCAAT-enhancer-binding protein (C/EBP)δ in TPC cells treated with CM with or without neutralizing antibody. (e) Phosphorylation of Stat3 and ERK1/2 in HC11 cells treated with LIF with or without the MAP kinase/ERK kinase inhibitor PD98059. (f) p-Stat3 and Stat3 in HC11 and TPC cells treated with LIF with or without the <t>Src-specific</t> inhibitor <t>PP2.</t> Experiments were repeated at least three times with similar results. No effect was observed on phosphorylation levels of either Stat3 or ERK1/2 when HC11 cells were treated with the PD98059 vehicle, dimethyl sulfoxide (data not shown).
    Sfk Inhibitor Proteinphosphatase 2, supplied by Millipore, used in various techniques. Bioz Stars score: 78/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Millipore csrc kinase inhibitor pp2
    Effect of tumor primary culture (TPC) conditioned medium (CM) and leukemia inhibitory factor (LIF)-neutralizing antibody. (a) Stat3 (signal transduction and activators of transcription 3) and phospho-Stat3 (p-Stat3) in HC11 and LM3 cells treated with increasing concentrations of CM (30%, 50% and 80%). (b) Stat3 and p-Stat3 levels in HC11 and TPC cells treated with CM that had been preincubated with or without LIF-neutralizing antibody. (c) Phosphorylation levels of Stat3 and extracellular signal-regulated kinase (ERK)1/2 in HC11 cells treated with LIF (3 and 5 ng/ml) preincubated with or without LIF-neutralizing antibody. (d) Expression of CCAAT-enhancer-binding protein (C/EBP)δ in TPC cells treated with CM with or without neutralizing antibody. (e) Phosphorylation of Stat3 and ERK1/2 in HC11 cells treated with LIF with or without the MAP kinase/ERK kinase inhibitor PD98059. (f) p-Stat3 and Stat3 in HC11 and TPC cells treated with LIF with or without the <t>Src-specific</t> inhibitor <t>PP2.</t> Experiments were repeated at least three times with similar results. No effect was observed on phosphorylation levels of either Stat3 or ERK1/2 when HC11 cells were treated with the PD98059 vehicle, dimethyl sulfoxide (data not shown).
    Csrc Kinase Inhibitor Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    Millipore pyrazolopyrimidine 2 pp2
    Effect of tumor primary culture (TPC) conditioned medium (CM) and leukemia inhibitory factor (LIF)-neutralizing antibody. (a) Stat3 (signal transduction and activators of transcription 3) and phospho-Stat3 (p-Stat3) in HC11 and LM3 cells treated with increasing concentrations of CM (30%, 50% and 80%). (b) Stat3 and p-Stat3 levels in HC11 and TPC cells treated with CM that had been preincubated with or without LIF-neutralizing antibody. (c) Phosphorylation levels of Stat3 and extracellular signal-regulated kinase (ERK)1/2 in HC11 cells treated with LIF (3 and 5 ng/ml) preincubated with or without LIF-neutralizing antibody. (d) Expression of CCAAT-enhancer-binding protein (C/EBP)δ in TPC cells treated with CM with or without neutralizing antibody. (e) Phosphorylation of Stat3 and ERK1/2 in HC11 cells treated with LIF with or without the MAP kinase/ERK kinase inhibitor PD98059. (f) p-Stat3 and Stat3 in HC11 and TPC cells treated with LIF with or without the <t>Src-specific</t> inhibitor <t>PP2.</t> Experiments were repeated at least three times with similar results. No effect was observed on phosphorylation levels of either Stat3 or ERK1/2 when HC11 cells were treated with the PD98059 vehicle, dimethyl sulfoxide (data not shown).
    Pyrazolopyrimidine 2 Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 97/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore chemical inhibitors
    Effect of tumor primary culture (TPC) conditioned medium (CM) and leukemia inhibitory factor (LIF)-neutralizing antibody. (a) Stat3 (signal transduction and activators of transcription 3) and phospho-Stat3 (p-Stat3) in HC11 and LM3 cells treated with increasing concentrations of CM (30%, 50% and 80%). (b) Stat3 and p-Stat3 levels in HC11 and TPC cells treated with CM that had been preincubated with or without LIF-neutralizing antibody. (c) Phosphorylation levels of Stat3 and extracellular signal-regulated kinase (ERK)1/2 in HC11 cells treated with LIF (3 and 5 ng/ml) preincubated with or without LIF-neutralizing antibody. (d) Expression of CCAAT-enhancer-binding protein (C/EBP)δ in TPC cells treated with CM with or without neutralizing antibody. (e) Phosphorylation of Stat3 and ERK1/2 in HC11 cells treated with LIF with or without the MAP kinase/ERK kinase inhibitor PD98059. (f) p-Stat3 and Stat3 in HC11 and TPC cells treated with LIF with or without the <t>Src-specific</t> inhibitor <t>PP2.</t> Experiments were repeated at least three times with similar results. No effect was observed on phosphorylation levels of either Stat3 or ERK1/2 when HC11 cells were treated with the PD98059 vehicle, dimethyl sulfoxide (data not shown).
    Chemical Inhibitors, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 424 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    76
    Millipore pan rtk inhibitor pp2
    Effect of tumor primary culture (TPC) conditioned medium (CM) and leukemia inhibitory factor (LIF)-neutralizing antibody. (a) Stat3 (signal transduction and activators of transcription 3) and phospho-Stat3 (p-Stat3) in HC11 and LM3 cells treated with increasing concentrations of CM (30%, 50% and 80%). (b) Stat3 and p-Stat3 levels in HC11 and TPC cells treated with CM that had been preincubated with or without LIF-neutralizing antibody. (c) Phosphorylation levels of Stat3 and extracellular signal-regulated kinase (ERK)1/2 in HC11 cells treated with LIF (3 and 5 ng/ml) preincubated with or without LIF-neutralizing antibody. (d) Expression of CCAAT-enhancer-binding protein (C/EBP)δ in TPC cells treated with CM with or without neutralizing antibody. (e) Phosphorylation of Stat3 and ERK1/2 in HC11 cells treated with LIF with or without the MAP kinase/ERK kinase inhibitor PD98059. (f) p-Stat3 and Stat3 in HC11 and TPC cells treated with LIF with or without the <t>Src-specific</t> inhibitor <t>PP2.</t> Experiments were repeated at least three times with similar results. No effect was observed on phosphorylation levels of either Stat3 or ERK1/2 when HC11 cells were treated with the PD98059 vehicle, dimethyl sulfoxide (data not shown).
    Pan Rtk Inhibitor Pp2, supplied by Millipore, used in various techniques. Bioz Stars score: 76/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Millipore pp2 4 amino 5 4 chlorophenyl 7 t butyl pyrazolo 3 4 d pyrimidine
    Effect of tumor primary culture (TPC) conditioned medium (CM) and leukemia inhibitory factor (LIF)-neutralizing antibody. (a) Stat3 (signal transduction and activators of transcription 3) and phospho-Stat3 (p-Stat3) in HC11 and LM3 cells treated with increasing concentrations of CM (30%, 50% and 80%). (b) Stat3 and p-Stat3 levels in HC11 and TPC cells treated with CM that had been preincubated with or without LIF-neutralizing antibody. (c) Phosphorylation levels of Stat3 and extracellular signal-regulated kinase (ERK)1/2 in HC11 cells treated with LIF (3 and 5 ng/ml) preincubated with or without LIF-neutralizing antibody. (d) Expression of CCAAT-enhancer-binding protein (C/EBP)δ in TPC cells treated with CM with or without neutralizing antibody. (e) Phosphorylation of Stat3 and ERK1/2 in HC11 cells treated with LIF with or without the MAP kinase/ERK kinase inhibitor PD98059. (f) p-Stat3 and Stat3 in HC11 and TPC cells treated with LIF with or without the <t>Src-specific</t> inhibitor <t>PP2.</t> Experiments were repeated at least three times with similar results. No effect was observed on phosphorylation levels of either Stat3 or ERK1/2 when HC11 cells were treated with the PD98059 vehicle, dimethyl sulfoxide (data not shown).
    Pp2 4 Amino 5 4 Chlorophenyl 7 T Butyl Pyrazolo 3 4 D Pyrimidine, supplied by Millipore, used in various techniques. Bioz Stars score: 89/100, based on 37 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Mechanically induced phosphorylation of the Y667-β-cat conserved E-cadherin interaction site in the mesoderm of Danio . ( a ) Phospho-β-cat labelling in zebrafish marginal cells before epiboly (sphere stage). ( b ) Phospho-β-cat labelling at the start of epiboly (dome stage). ( c ) Phospho-β-cat labelling after blebbistatin treatment that suppressed movements. ( d ) Phospho-β-cat labelling after blebbistatin treatment with epiboly movements rescued by global compression. ( e ) Phospho-β-cat labelling after blebbistatin treatment with epiboly movements rescued by drug washing. ( f ) Phospho-β-cat labelling after blebbistatin treatment rescued by magnetic manipulation of UML-injected embryos leading to epiboly movement resumption. ( g ) Phospho-β-cat labelling in the presence of <t>PP2</t> Src-family inhibitor treatment at dome (associated β-cat nuclear translocation tests in Supplementary Fig. S15b ). ( h ) β-cat labelling in blebbistatin globally compressed embryos treated with PP2. ( i ) β-cat labelling in blebbistatin magnetically deformed UML-injected embryos treated with PP2. Associated quantitative results in Supplementary Fig. S15b . ( j ) Levels of pY667 β-cat in the margin relative to the blastoderm centre and in the margin relative to the background in sphere stage ( n =6), dome stage ( n =9), PP2 treated ( n =7), blebbistatin-treated ( n =22), blebbistatin-treated and washed ( n =7), and blebbistatin-treated UML-injected magnetically rescued embryos ( n =9). Differences between control dome, blebbistatin-washed or blebbistatin-compressed embryos and all other conditions are statistically significant ( P
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    OxLDL-induced Vav1 phosphorylation is mediated by <t>src</t> kinase Fyn. (A) Human platelets were incubated with the broad-spectrum src inhibitor <t>AG1879</t> before incubation with 50 μg/mL oxLDL. The platelet lysates were then analyzed as in for
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    OxLDL-induced Vav1 phosphorylation is mediated by <t>src</t> kinase Fyn. (A) Human platelets were incubated with the broad-spectrum src inhibitor <t>AG1879</t> before incubation with 50 μg/mL oxLDL. The platelet lysates were then analyzed as in for
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    OxLDL-induced Vav1 phosphorylation is mediated by <t>src</t> kinase Fyn. (A) Human platelets were incubated with the broad-spectrum src inhibitor <t>AG1879</t> before incubation with 50 μg/mL oxLDL. The platelet lysates were then analyzed as in for
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    OxLDL-induced Vav1 phosphorylation is mediated by <t>src</t> kinase Fyn. (A) Human platelets were incubated with the broad-spectrum src inhibitor <t>AG1879</t> before incubation with 50 μg/mL oxLDL. The platelet lysates were then analyzed as in for
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    Phosphatidylinositol 3 (PI3) kinase activity mediates oxidative stress-induced phosphorylation of c-Src. A : Caco-2 cell monolayers were pretreated with wortmannin (0.1 μM), LY294002 (25 μM), or <t>4-amino-5[chlorophyll]-7-[t-butyl]pyrazolo[3–4-d]pyrimidine</t>
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    Phosphatidylinositol 3 (PI3) kinase activity mediates oxidative stress-induced phosphorylation of c-Src. A : Caco-2 cell monolayers were pretreated with wortmannin (0.1 μM), LY294002 (25 μM), or <t>4-amino-5[chlorophyll]-7-[t-butyl]pyrazolo[3–4-d]pyrimidine</t>
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    Image Search Results


    Caveolin-1 and EGFR co-localization as a feature of non-canonical EGFR activation. Epithelial cells (RLE-6TN) were exposed (5 min) to carbon nanoparticles (CNP), non-nano carbon particles (CP), each 10 µg/cm 2 , (50 µM) H 2 O 2 , (5 µM) C6 ceramide, or EGF (100 ng/mL), respectively. ( A ) Subcellular localization of EGFR (red Alexa flour 594) and caveolin-1 (green Alexa flour 488). Co-localization is visualized by the yellow color in merged images; ( B ) Subcellular localization of EGFR and caveolin-1 in cells pre-treated with inhibitors of carbon nanoparticle-specific signaling prior to particle or EGF exposure: SFK inhibitor PP2 (10 µM), 1 mM ectoine ( E ), 75 µM α-tocopherol (Toco), and 1 µg/mL filipin III (Fil). Co-localization is visualized by the yellow color in merged images; ( C ) Subcellular localization of caveolin-1 (red Alexa fluor 594) after exposure to carbon nanoparticles (CNP), carbon particles (CP), or hydrogen peroxide (H 2 O 2 ); ( D ) Quantification and representative Western blots of caveolin-1 in lipid raft fraction of RLE-6TN cells exposed to CNP (10 µg/cm 2 ). Raft and non-raft fraction were isolated from density gradients after ultracentrifugation. Pre-treatment of cells with α-tocopherol (Toco) was applied as an antioxidant strategy. GAPDH was used as a control protein not associated with lipid rafts. The bars in the graph represent the additive immune signals of raft and non-raft fractions, which was indicated in the representative original Western blots; ( E ) Quantification and representative Western-blot of EGFR phosphorylation at Tyr 845 . Nuclei were stained with DAPI (blue). Scale bars represent 20 µm; *, which was significantly different to PBS control ( p

    Journal: Nanomaterials

    Article Title: Non-Canonical Activation of the Epidermal Growth Factor Receptor by Carbon Nanoparticles

    doi: 10.3390/nano8040267

    Figure Lengend Snippet: Caveolin-1 and EGFR co-localization as a feature of non-canonical EGFR activation. Epithelial cells (RLE-6TN) were exposed (5 min) to carbon nanoparticles (CNP), non-nano carbon particles (CP), each 10 µg/cm 2 , (50 µM) H 2 O 2 , (5 µM) C6 ceramide, or EGF (100 ng/mL), respectively. ( A ) Subcellular localization of EGFR (red Alexa flour 594) and caveolin-1 (green Alexa flour 488). Co-localization is visualized by the yellow color in merged images; ( B ) Subcellular localization of EGFR and caveolin-1 in cells pre-treated with inhibitors of carbon nanoparticle-specific signaling prior to particle or EGF exposure: SFK inhibitor PP2 (10 µM), 1 mM ectoine ( E ), 75 µM α-tocopherol (Toco), and 1 µg/mL filipin III (Fil). Co-localization is visualized by the yellow color in merged images; ( C ) Subcellular localization of caveolin-1 (red Alexa fluor 594) after exposure to carbon nanoparticles (CNP), carbon particles (CP), or hydrogen peroxide (H 2 O 2 ); ( D ) Quantification and representative Western blots of caveolin-1 in lipid raft fraction of RLE-6TN cells exposed to CNP (10 µg/cm 2 ). Raft and non-raft fraction were isolated from density gradients after ultracentrifugation. Pre-treatment of cells with α-tocopherol (Toco) was applied as an antioxidant strategy. GAPDH was used as a control protein not associated with lipid rafts. The bars in the graph represent the additive immune signals of raft and non-raft fractions, which was indicated in the representative original Western blots; ( E ) Quantification and representative Western-blot of EGFR phosphorylation at Tyr 845 . Nuclei were stained with DAPI (blue). Scale bars represent 20 µm; *, which was significantly different to PBS control ( p

    Article Snippet: Filipin III (from Streptomyces filipinensis , Sigma-Aldrich Chemie, Schnelldorf, Germany), PP2 (Calbiochem, Schwalbach, Germany), and DSP (dithiobis-succimidylpropionate, Thermo Scientific, Waltham, MA, USA) were solubilized in DMSO (dimethyl sulfoxide) and diluted in PBS to the indicated concentrations. α-tocopherol ( d -alpha-tocopherol succinate, semi-synthetic, Sigma-Aldrich Chemie, Schnelldorf, Germany) was solubilized in ethanol and further diluted in PBS before use.

    Techniques: Activation Assay, Western Blot, Isolation, Staining

    Src activation, via FRS2, is essential to NGF-dependent macropinocytosis and cell death. (A) Schematic showing that Src is recruited into TrkA signaling via FRS2, and this facilitates activation of H-Ras. (B) Daoy-TrkA cells were treated with PP2 or DMSO (1 h) prior to addition of NGF and incubated for an additional 12 h. Cells were scored for cell death using a trypan blue exclusion assay ( n = 3). The asterisks represent a statistically significant decrease ( P

    Journal: Molecular and Cellular Biology

    Article Title: Unravelling the Mechanism of TrkA-Induced Cell Death by Macropinocytosis in Medulloblastoma Daoy Cells

    doi: 10.1128/MCB.00255-16

    Figure Lengend Snippet: Src activation, via FRS2, is essential to NGF-dependent macropinocytosis and cell death. (A) Schematic showing that Src is recruited into TrkA signaling via FRS2, and this facilitates activation of H-Ras. (B) Daoy-TrkA cells were treated with PP2 or DMSO (1 h) prior to addition of NGF and incubated for an additional 12 h. Cells were scored for cell death using a trypan blue exclusion assay ( n = 3). The asterisks represent a statistically significant decrease ( P

    Article Snippet: The Src inhibitor PP2 (Sigma-Aldrich) was used at the concentrations indicated.

    Techniques: Activation Assay, Incubation, Trypan Blue Exclusion Assay

    The UNC-5 cytosolic domain interacts with SRC-1 in vitro. (A) SRC-1 binds the UNC-5 cytosolic domain. HEK293 cells were transfected with the indicated expression vectors. SRC-1-myc was immunoprecipitated (IP) with anti-myc antibody (9E10) and subjected to Western blot analysis (WB) with the indicated antibodies. (B) The SH2 domain of SRC-1 is important for its interaction with UNC-5. Lysates were subjected to immunoprecipitation with an anti-myc antibody (9E10) and probed for the indicated Western blot analysis. SH2M and SH3M denote SRC-1 mutations in the SH2 domain (R165A) and SH3 domain (W108R W109R), respectively. WT, wild type. (C) PP2 inhibits the interaction between SRC-1 and UNC-5. Transfected HEK293 cells were treated with PP2, an inhibitor of Src family kinases, for 30 min at the indicated concentrations (μM). Lysates were immunoprecipitated with anti-myc antibody (9E10) and probed for the indicated Western blot analysis.

    Journal: Molecular and Cellular Biology

    Article Title: SRC-1 Mediates UNC-5 Signaling in Caenorhabditis elegans

    doi: 10.1128/MCB.25.15.6485-6495.2005

    Figure Lengend Snippet: The UNC-5 cytosolic domain interacts with SRC-1 in vitro. (A) SRC-1 binds the UNC-5 cytosolic domain. HEK293 cells were transfected with the indicated expression vectors. SRC-1-myc was immunoprecipitated (IP) with anti-myc antibody (9E10) and subjected to Western blot analysis (WB) with the indicated antibodies. (B) The SH2 domain of SRC-1 is important for its interaction with UNC-5. Lysates were subjected to immunoprecipitation with an anti-myc antibody (9E10) and probed for the indicated Western blot analysis. SH2M and SH3M denote SRC-1 mutations in the SH2 domain (R165A) and SH3 domain (W108R W109R), respectively. WT, wild type. (C) PP2 inhibits the interaction between SRC-1 and UNC-5. Transfected HEK293 cells were treated with PP2, an inhibitor of Src family kinases, for 30 min at the indicated concentrations (μM). Lysates were immunoprecipitated with anti-myc antibody (9E10) and probed for the indicated Western blot analysis.

    Article Snippet: In order to inhibit Src family kinase activity, cells were starved for 3 h in the serum-free medium and then the inhibitor PP2 (Calbiochem) was treated at the concentrations indicated in Fig. for 30 min ( ).

    Techniques: In Vitro, Transfection, Expressing, Immunoprecipitation, Western Blot

    Src and p38 kinases inhibitors effect on the expression of p38/Src protein phosphorylation and heparanase protein. A: The expression of heparanase protein did not change in human gastric cancer SGC-7901 cells treated with 5 μmol/L pp2 or 1 μmol/L

    Journal: International Journal of Clinical and Experimental Pathology

    Article Title: Heparanase promotes human gastric cancer cells migration and invasion by increasing Src and p38 phosphorylation expression

    doi:

    Figure Lengend Snippet: Src and p38 kinases inhibitors effect on the expression of p38/Src protein phosphorylation and heparanase protein. A: The expression of heparanase protein did not change in human gastric cancer SGC-7901 cells treated with 5 μmol/L pp2 or 1 μmol/L

    Article Snippet: The selective p38 (SB 203580) and Src (PP2) inhibitors were purchased from Calbiochem (San Diego, CA) and were dissolved in DMSO as stock solutions.

    Techniques: Expressing

    Dual infection of H. pylori strains expressing different single phosphorylatable or phosphomimetic EPIYA motifs induces AGS cell elongation. AGS cells were infected for 4 hours with CagA EPIYA Y > F ( A ) or EPIYA Y > D ( B ) mutant strains as indicated. The available single phosphorylatable or phosphomimetic EPIYA motifs for each double infection are indicated. The number of elongated cells in each experiment was quantitated in triplicate in 10 different 0.25-mm 2 fields, and CagA phosphorylation was examined using α–PY-99 and α-CagA antibodies (arrows). ( C ) AGS infection for 4 hours with the indicated CagA-expressing strains in the presence or absence of c-Src inhibitor PP2 (10 μM) or c-Abl inhibitor SKI-DV2-43 (1 μM) revealed significant changes in AGS cell elongation as quantitated in triplicate in 10 different 0.25-mm 2 fields. * P ≤ 0.01; ** P ≤ 0.001.

    Journal: The Journal of Clinical Investigation

    Article Title: c-Src and c-Abl kinases control hierarchic phosphorylation and function of the CagA effector protein in Western and East Asian Helicobacter pylori strains

    doi: 10.1172/JCI61143

    Figure Lengend Snippet: Dual infection of H. pylori strains expressing different single phosphorylatable or phosphomimetic EPIYA motifs induces AGS cell elongation. AGS cells were infected for 4 hours with CagA EPIYA Y > F ( A ) or EPIYA Y > D ( B ) mutant strains as indicated. The available single phosphorylatable or phosphomimetic EPIYA motifs for each double infection are indicated. The number of elongated cells in each experiment was quantitated in triplicate in 10 different 0.25-mm 2 fields, and CagA phosphorylation was examined using α–PY-99 and α-CagA antibodies (arrows). ( C ) AGS infection for 4 hours with the indicated CagA-expressing strains in the presence or absence of c-Src inhibitor PP2 (10 μM) or c-Abl inhibitor SKI-DV2-43 (1 μM) revealed significant changes in AGS cell elongation as quantitated in triplicate in 10 different 0.25-mm 2 fields. * P ≤ 0.01; ** P ≤ 0.001.

    Article Snippet: The c-Abl tyrosine kinase inhibitor SKI-DV2-43 ( ) and c-Src inhibitor PP2 (Calbiochem) were dissolved in Me2 SO and added to the cells 30 minutes before infection.

    Techniques: Infection, Expressing, Mutagenesis

    Analysis of CagA PY protein species during infection with H. pylori by 1-DE and 2-DE. ( A ). ( B ) AGS cells were infected for the indicated times with strain 26695. The resulting protein lysates were separated by 1-DE, and phosphorylation of injected CagA was examined using α–PY-99 and α-CagA antibodies (arrows). ( C ) Separation of CagA protein species from B by 2-DE. Depending on the time of infection, full-length CagA PY appeared as 1 spot (spot 1, red arrows, pI = 7.0) or 2 spots (spots 1 and 2; spot 2, green arrows, pI = 6.5) as indicated. The α-CagA antibody probe revealed a third spot (spot 3, blue arrows, pI = 7.5). Overlay of both exposures yielded 2 or 3 spots as shown. Strain TN2-GF4 exhibited the same pattern as 26695 (bottom). ( D ) Inhibition of Src with PP2 (10 μM) or Abl with SKI-DV2-43 (1 μM) revealed significant changes in spot intensity depending on the time of infection.

    Journal: The Journal of Clinical Investigation

    Article Title: c-Src and c-Abl kinases control hierarchic phosphorylation and function of the CagA effector protein in Western and East Asian Helicobacter pylori strains

    doi: 10.1172/JCI61143

    Figure Lengend Snippet: Analysis of CagA PY protein species during infection with H. pylori by 1-DE and 2-DE. ( A ). ( B ) AGS cells were infected for the indicated times with strain 26695. The resulting protein lysates were separated by 1-DE, and phosphorylation of injected CagA was examined using α–PY-99 and α-CagA antibodies (arrows). ( C ) Separation of CagA protein species from B by 2-DE. Depending on the time of infection, full-length CagA PY appeared as 1 spot (spot 1, red arrows, pI = 7.0) or 2 spots (spots 1 and 2; spot 2, green arrows, pI = 6.5) as indicated. The α-CagA antibody probe revealed a third spot (spot 3, blue arrows, pI = 7.5). Overlay of both exposures yielded 2 or 3 spots as shown. Strain TN2-GF4 exhibited the same pattern as 26695 (bottom). ( D ) Inhibition of Src with PP2 (10 μM) or Abl with SKI-DV2-43 (1 μM) revealed significant changes in spot intensity depending on the time of infection.

    Article Snippet: The c-Abl tyrosine kinase inhibitor SKI-DV2-43 ( ) and c-Src inhibitor PP2 (Calbiochem) were dissolved in Me2 SO and added to the cells 30 minutes before infection.

    Techniques: Infection, Injection, Inhibition

    Particulate-triggered IL-2 release depends on actin polymerization, Syk and Src kinases, and NFAT activation. DCs were primed with LPS (100 ng/ml) for 30 min prior to addition of inhibitors: cytochalasin D ( A ), Syk inhibitor ( B ), Src inhibitor PP2 ( C

    Journal: The Journal of Immunology Author Choice

    Article Title: The Syk–NFAT–IL-2 Pathway in Dendritic Cells Is Required for Optimal Sterile Immunity Elicited by Alum Adjuvants

    doi: 10.4049/jimmunol.1600420

    Figure Lengend Snippet: Particulate-triggered IL-2 release depends on actin polymerization, Syk and Src kinases, and NFAT activation. DCs were primed with LPS (100 ng/ml) for 30 min prior to addition of inhibitors: cytochalasin D ( A ), Syk inhibitor ( B ), Src inhibitor PP2 ( C

    Article Snippet: The following small molecule inhibitors were used: cytochalasin D, PP2 Src inhibitor (Sigma-Aldrich), Syk inhibitor II CAS 227449-73-2 (Calbiochem), cyclosporin A, and FK506 (both from Cell Signaling Technology).

    Techniques: Activation Assay

    Distribution of β-catenin in monolayers treated with either DMSO or the Src Inhibitor PP2 8 hr after the application of No Strain or High Strain (15%) using the ISA. Scale bars: 25 μm. DOI: http://dx.doi.org/10.7554/eLife.19799.015

    Journal: eLife

    Article Title: Increasing β-catenin/Wnt3A activity levels drive mechanical strain-induced cell cycle progression through mitosis

    doi: 10.7554/eLife.19799

    Figure Lengend Snippet: Distribution of β-catenin in monolayers treated with either DMSO or the Src Inhibitor PP2 8 hr after the application of No Strain or High Strain (15%) using the ISA. Scale bars: 25 μm. DOI: http://dx.doi.org/10.7554/eLife.19799.015

    Article Snippet: Src Inhibitors PP2 (10 μM; EMD Millipore, Germany) and SU6656 (10 μM, Sigma-Aldrich), CKI inhibitor D4476 (10 μM; Abcam), EGFR inhibitor PD153035 (2.5 μM, Abcam, United Kingdom), and DMSO controls were added to strain device wells 15 min prior to strain application.

    Techniques:

    The Src Inhibitor PP2 blocks strain-induced increases in Y654 phosphorylated β-catenin and β-catenin transcriptional activity. Distribution of TOPdGFP at 8 hr ( A ), β-catenin at 8 hr (A, insets), EdU at 24 hr ( C ), and pY654 β-catenin at 8 hr ( E ) in MDCK Monolayers after No Strain or High Strain (15%) applied by the ISA, treated with either DMSO or the Src inhibitor PP2. Quantification of percent cells TOPdGFP- ( B ) or EdU- ( D ) positive and quantification of average pY654 β-catenin intensity per pixel ( F ). Quantifications were from at least 3 independent experiments and for the TOPdGFP and EdU quantifications included analysis of 707–1478 cells per experiment. Quantifications were mean +/- SEM; unpaired t-test ( B, D ) or Kolmogorov-Smirnoff ( F ) test p values

    Journal: eLife

    Article Title: Increasing β-catenin/Wnt3A activity levels drive mechanical strain-induced cell cycle progression through mitosis

    doi: 10.7554/eLife.19799

    Figure Lengend Snippet: The Src Inhibitor PP2 blocks strain-induced increases in Y654 phosphorylated β-catenin and β-catenin transcriptional activity. Distribution of TOPdGFP at 8 hr ( A ), β-catenin at 8 hr (A, insets), EdU at 24 hr ( C ), and pY654 β-catenin at 8 hr ( E ) in MDCK Monolayers after No Strain or High Strain (15%) applied by the ISA, treated with either DMSO or the Src inhibitor PP2. Quantification of percent cells TOPdGFP- ( B ) or EdU- ( D ) positive and quantification of average pY654 β-catenin intensity per pixel ( F ). Quantifications were from at least 3 independent experiments and for the TOPdGFP and EdU quantifications included analysis of 707–1478 cells per experiment. Quantifications were mean +/- SEM; unpaired t-test ( B, D ) or Kolmogorov-Smirnoff ( F ) test p values

    Article Snippet: Src Inhibitors PP2 (10 μM; EMD Millipore, Germany) and SU6656 (10 μM, Sigma-Aldrich), CKI inhibitor D4476 (10 μM; Abcam), EGFR inhibitor PD153035 (2.5 μM, Abcam, United Kingdom), and DMSO controls were added to strain device wells 15 min prior to strain application.

    Techniques: Activity Assay

    Src regulates motility and PG down-regulates Src activity in prostate cancer. A–B. PC3-M, C4-2B (A), and LNCaP cells (B) were plated in 6 well plates and allowed to attach and spread. LNCaP cells were transfected with control siRNA or PG siRNA (a pool of 4 sequences). PC3-M, C4-2B, and LNCaP cells were treated with medium containing PP2 (10 µM) or DMSO (solvent control) for 24 hours prior to performing the dispase assay. Inhibition of Src strengthens cell-cell adhesion in PCa cells in general and is able to rescue cell-cell adhesion in PG-deficient cells. C. PC3-M and ARCaP M cells were plated in 24-well plates and allowed to attach and spread. The cells were treated with medium containing the selective Src- family kinase inhibitor PP2 (10 µM) or DMSO (solvent control) for 24 hours prior to performing the scratch wound assay. D. ARCaP E cells were plated in 24-well plates and allowed to attach and spread. The cells were transduced with GFP-containing adenovirus or caSrc-containing adenovirus, and after 24 hours a scratch wound was made. Activity of Src is directly correlated with motility of PCa cells. E. ARCaP M , PC3-M, and LNCaP cells were plated in 6-well plates and allowed to attach and spread. LNCaP cells were transfected with control siRNA or PG siRNA pool and after 96 hours, the cells were lysed. ARCAP M and PC3-M cells were transduced with GFP-containing adenovirus or PG-containing adenovirus, and after 24 hours the cells were lysed. The lysates were subjected to SDS-PAGE followed by immunoblotting with antibodies against PG, pSrc, Src, and GAPDH. Levels of pSrc were determined by normalizing to Src levels for each cell line using Image J. Phosphorylation (activation) of Src is inversely correlated with PG levels. Representatives of at least three independent immunoblots are shown, with numbers representing GAPDH normalized pSrc/Src ratio for the blot shown. The average ratio and standard deviation of pSrc/Src normalized for loading by GAPDH is as follows: ARCaP M 0.6+/−0.2, LNCaP 1.7+/−0.2, PC3M 0.7+/−0.1. Graphs represent averages +/− SEM. *P

    Journal: PLoS ONE

    Article Title: The Desmosomal Armadillo Protein Plakoglobin Regulates Prostate Cancer Cell Adhesion and Motility through Vitronectin-Dependent Src Signaling

    doi: 10.1371/journal.pone.0042132

    Figure Lengend Snippet: Src regulates motility and PG down-regulates Src activity in prostate cancer. A–B. PC3-M, C4-2B (A), and LNCaP cells (B) were plated in 6 well plates and allowed to attach and spread. LNCaP cells were transfected with control siRNA or PG siRNA (a pool of 4 sequences). PC3-M, C4-2B, and LNCaP cells were treated with medium containing PP2 (10 µM) or DMSO (solvent control) for 24 hours prior to performing the dispase assay. Inhibition of Src strengthens cell-cell adhesion in PCa cells in general and is able to rescue cell-cell adhesion in PG-deficient cells. C. PC3-M and ARCaP M cells were plated in 24-well plates and allowed to attach and spread. The cells were treated with medium containing the selective Src- family kinase inhibitor PP2 (10 µM) or DMSO (solvent control) for 24 hours prior to performing the scratch wound assay. D. ARCaP E cells were plated in 24-well plates and allowed to attach and spread. The cells were transduced with GFP-containing adenovirus or caSrc-containing adenovirus, and after 24 hours a scratch wound was made. Activity of Src is directly correlated with motility of PCa cells. E. ARCaP M , PC3-M, and LNCaP cells were plated in 6-well plates and allowed to attach and spread. LNCaP cells were transfected with control siRNA or PG siRNA pool and after 96 hours, the cells were lysed. ARCAP M and PC3-M cells were transduced with GFP-containing adenovirus or PG-containing adenovirus, and after 24 hours the cells were lysed. The lysates were subjected to SDS-PAGE followed by immunoblotting with antibodies against PG, pSrc, Src, and GAPDH. Levels of pSrc were determined by normalizing to Src levels for each cell line using Image J. Phosphorylation (activation) of Src is inversely correlated with PG levels. Representatives of at least three independent immunoblots are shown, with numbers representing GAPDH normalized pSrc/Src ratio for the blot shown. The average ratio and standard deviation of pSrc/Src normalized for loading by GAPDH is as follows: ARCaP M 0.6+/−0.2, LNCaP 1.7+/−0.2, PC3M 0.7+/−0.1. Graphs represent averages +/− SEM. *P

    Article Snippet: PP2 selective Src inhibitor was from Calbiochem (San Diego, CA).

    Techniques: Activity Assay, Transfection, Inhibition, Scratch Wound Assay Assay, Transduction, SDS Page, Activation Assay, Western Blot, Standard Deviation

    Quantitative analysis of radial maturation. A , Schema of the five boxes encompassing the outer (1) and inner (2) tiers of the cortical ribbon (excluding the meninges), the outer (3) and inner (4) tiers of the intermediate zone of migration, and the ventricular–subventricular zones (5) in which BrdU- or THY-positive neurons were counted. B , Between 300–400 BrdU- and 300–400 THY-labeled cells were counted in four different sections from normal, reeler, PP2-, and BIM1-treated slices, and the proportion in boxes 1–5 was graphed in gray (BrdU) and black (THY). Boxes 1 and 2 defined in the cortical plate are the most relevant, because the proportion of labeled cells in the deep boxes varied more from experiment to experiment.

    Journal: The Journal of Neuroscience

    Article Title: Inhibition of Src Family Kinases and Non-Classical Protein Kinases C Induce a Reeler-Like Malformation of Cortical Plate Development

    doi: 10.1523/JNEUROSCI.23-30-09953.2003

    Figure Lengend Snippet: Quantitative analysis of radial maturation. A , Schema of the five boxes encompassing the outer (1) and inner (2) tiers of the cortical ribbon (excluding the meninges), the outer (3) and inner (4) tiers of the intermediate zone of migration, and the ventricular–subventricular zones (5) in which BrdU- or THY-positive neurons were counted. B , Between 300–400 BrdU- and 300–400 THY-labeled cells were counted in four different sections from normal, reeler, PP2-, and BIM1-treated slices, and the proportion in boxes 1–5 was graphed in gray (BrdU) and black (THY). Boxes 1 and 2 defined in the cortical plate are the most relevant, because the proportion of labeled cells in the deep boxes varied more from experiment to experiment.

    Article Snippet: To study that question further, we tested whether inhibition of Src family kinases with the well validated inhibitor PP2 ( ) (Calbiochem) would perturb neuronal migration and architectonic development in embryonic slices.

    Techniques: Migration, Labeling

    Alterations of the radial glial scaffold, revealed with an anti-nestin antibody. Although normal slices ( A ) are characterized by a finely organized radial glial scaffold, particularly in its intracortical portion and in the MZ, radial glial fibers in slices from reeler embryos ( B ) are distorted, with enlarged and branched segments, particularly in the cortical ribbon and in their subpial portion. A rather similar morphology is found when normal slices are treated with PP2 ( C ) or BIM1 ( D ): radial glial extensions are wavy and show enlarged portions and increased branching. Scale bar, 80 μm.

    Journal: The Journal of Neuroscience

    Article Title: Inhibition of Src Family Kinases and Non-Classical Protein Kinases C Induce a Reeler-Like Malformation of Cortical Plate Development

    doi: 10.1523/JNEUROSCI.23-30-09953.2003

    Figure Lengend Snippet: Alterations of the radial glial scaffold, revealed with an anti-nestin antibody. Although normal slices ( A ) are characterized by a finely organized radial glial scaffold, particularly in its intracortical portion and in the MZ, radial glial fibers in slices from reeler embryos ( B ) are distorted, with enlarged and branched segments, particularly in the cortical ribbon and in their subpial portion. A rather similar morphology is found when normal slices are treated with PP2 ( C ) or BIM1 ( D ): radial glial extensions are wavy and show enlarged portions and increased branching. Scale bar, 80 μm.

    Article Snippet: To study that question further, we tested whether inhibition of Src family kinases with the well validated inhibitor PP2 ( ) (Calbiochem) would perturb neuronal migration and architectonic development in embryonic slices.

    Techniques:

    Src-family kinase signaling, but not Syk or PI3K signaling, is indispensable for reorganization of macrophage surfaces. (A) Immunoblots of phosphorylated AKT in nonactivated (PLL) or hIgG-activated human macrophages pretreated with vehicle (DMSO), as a control, 10 µM PP2 (left), 100 µM piceatannol (PCT; middle), or 1 µM wortmannin (Wort; right). Blots represent two independent experiments. (B) TIRF image of FcγRI (white; bars, 20 µm) and dSTORM images (bars, 5 µm) of FcγRI (green) and SIRPα (red) at the surface of human macrophages incubated with vehicle (DMSO), PP2, PCT, or Wort, pretreated as in A. Cells were then seeded onto slides coated with PLL (nonactivated) or hIgG for 10 min and stained with anti–FcγRI-AF488 and anti–SIRPα-AF647 mAbs. In each condition, regions outlined by the white squares (middle column) are shown enlarged (right column). Bars, 1 µm. (C) CBC histograms for FcγRI and SIRPα in cells pretreated as in A and seeded onto slides coated with PLL (gray) or hIgG (DMSO, dark gray; PP2, red; PCT, green; and Wort, blue) for 10 min, as indicated. Data are from a minimum of 30 cells from three independent donors. Bars show mean ± SD. (D) NND analysis from data shown in C. Each symbol represents the median NND of all paired single-molecule localizations from one cell. Horizontal lines and error bars represent mean ± SD. ns, not significant; ****, P

    Journal: The Journal of Cell Biology

    Article Title: Membrane nanoclusters of FcγRI segregate from inhibitory SIRPα upon activation of human macrophages

    doi: 10.1083/jcb.201608094

    Figure Lengend Snippet: Src-family kinase signaling, but not Syk or PI3K signaling, is indispensable for reorganization of macrophage surfaces. (A) Immunoblots of phosphorylated AKT in nonactivated (PLL) or hIgG-activated human macrophages pretreated with vehicle (DMSO), as a control, 10 µM PP2 (left), 100 µM piceatannol (PCT; middle), or 1 µM wortmannin (Wort; right). Blots represent two independent experiments. (B) TIRF image of FcγRI (white; bars, 20 µm) and dSTORM images (bars, 5 µm) of FcγRI (green) and SIRPα (red) at the surface of human macrophages incubated with vehicle (DMSO), PP2, PCT, or Wort, pretreated as in A. Cells were then seeded onto slides coated with PLL (nonactivated) or hIgG for 10 min and stained with anti–FcγRI-AF488 and anti–SIRPα-AF647 mAbs. In each condition, regions outlined by the white squares (middle column) are shown enlarged (right column). Bars, 1 µm. (C) CBC histograms for FcγRI and SIRPα in cells pretreated as in A and seeded onto slides coated with PLL (gray) or hIgG (DMSO, dark gray; PP2, red; PCT, green; and Wort, blue) for 10 min, as indicated. Data are from a minimum of 30 cells from three independent donors. Bars show mean ± SD. (D) NND analysis from data shown in C. Each symbol represents the median NND of all paired single-molecule localizations from one cell. Horizontal lines and error bars represent mean ± SD. ns, not significant; ****, P

    Article Snippet: Drug treatments Primary human macrophages were pretreated with 10 µM of either the SFK inhibitor PP2 (Sigma-Aldrich) or the myosin II inhibitor blebbistatin (Sigma-Aldrich), with 100 µM of the Syk kinase inhibitor piceatannol (Sigma-Aldrich), or with 0.5 µM jasplakinolide (Sigma-Aldrich) for 30 min, or with 1 µM of either latrunculin A (EMD Millipore) or the PI3K inhibitor wortmannin (Sigma-Aldrich), or with 10 µM of the formin inhibitor SMIFH2 (Sigma-Aldrich) for 10 min, in PBS at 37°C.

    Techniques: Western Blot, Incubation, Staining

    β1- and β3-mediated CLAN formation are dependent upon a Src family kinase while only β1-mediated CLAN formation is PI3-K dependent

    Journal:

    Article Title: Distinct ?1 and ?3 integrin pathways converge to regulate cross-linked actin network (CLAN) formation in human trabecular meshwork (HTM) cells

    doi: 10.1167/iovs.08-3215

    Figure Lengend Snippet: β1- and β3-mediated CLAN formation are dependent upon a Src family kinase while only β1-mediated CLAN formation is PI3-K dependent

    Article Snippet: Prior to re-plating onto coverslips pre-coated with 20 nM fibronectin, suspended cells were pre-incubated for 30–60 minutes in the absence or presence of 0.25, 1, 5 or 20 μM of the Src family kinase (SFK) inhibitor PP2 (EMD Biosciences, Inc., San Diego, CA), 60 or 120 μM of the selective SFK inhibitor cis-5,8,11,14,17-eicosapentaenoic acid (EPA, Sigma), 20 μM of the phosphatidylinositol 3-kinase (PI3-K) inhibitor LY294002 (EMD Bioscience, Inc.), 20 μM of the Rac1 inhibitor NSC23766 (kindly provided by Yi Zheng, Children's Hospital Research Foundation, Cincinnati, OH), or 100 nM of the dominant negative (DN) form of the Rac1 guanine nucleotide exchange factor (GEF) Tiam1.

    Techniques:

    PP2 promotes expression of late chondrocyte marker genes. Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide (DMSO) or the Src inhibitor PP2 (10 μmol/l), and transcript levels of late chondrocyte marker genes were determined by real-time PCR. Expression levels of (a) Col10a1 , (b) Ihh , (c) Cdkn1c and (d) Atf3 were significantly increased upon Src inhibition ( n = 3; * P

    Journal: Arthritis Research & Therapy

    Article Title: Src kinase inhibition promotes the chondrocyte phenotype

    doi: 10.1186/ar2308

    Figure Lengend Snippet: PP2 promotes expression of late chondrocyte marker genes. Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide (DMSO) or the Src inhibitor PP2 (10 μmol/l), and transcript levels of late chondrocyte marker genes were determined by real-time PCR. Expression levels of (a) Col10a1 , (b) Ihh , (c) Cdkn1c and (d) Atf3 were significantly increased upon Src inhibition ( n = 3; * P

    Article Snippet: Primary chondrocyte cultures were treated with 1 μmol/l or 10 μmol/l of the Src family kinase inhibitor PP2 (Calbiochem; VWR, Mississauga, Ontario, Canada) or with dimethyl sulphoxide (DMSO; Sigma), the vehicle in which PP2 was dissolved, as control.

    Techniques: Expressing, Marker, Incubation, Real-time Polymerase Chain Reaction, Inhibition

    PP2 promotes cell rounding and cortical actin formation. Primary mouse chondrocytes were incubated for 24 hours with dimethyl sulphoxide (DMSO) or the Src inhibitor PP2 (10 μmol/l), and cells were stained with antibodies against total focal adhesion kinase (FAK) or FAK phosphorylated on residue tyrosine 397 (green), rhodamine-phalloidin (red) and DAPI (blue). In the presence of DMSO, total and phosphorylated actin localized to focal adhesions at the end of stress fibres. In cells treated with PP2, total FAK acquired a diffuse cytosolic staining, whereas the signal for phosphorylated FAK was greatly reduced (scale bar: 2 μm).

    Journal: Arthritis Research & Therapy

    Article Title: Src kinase inhibition promotes the chondrocyte phenotype

    doi: 10.1186/ar2308

    Figure Lengend Snippet: PP2 promotes cell rounding and cortical actin formation. Primary mouse chondrocytes were incubated for 24 hours with dimethyl sulphoxide (DMSO) or the Src inhibitor PP2 (10 μmol/l), and cells were stained with antibodies against total focal adhesion kinase (FAK) or FAK phosphorylated on residue tyrosine 397 (green), rhodamine-phalloidin (red) and DAPI (blue). In the presence of DMSO, total and phosphorylated actin localized to focal adhesions at the end of stress fibres. In cells treated with PP2, total FAK acquired a diffuse cytosolic staining, whereas the signal for phosphorylated FAK was greatly reduced (scale bar: 2 μm).

    Article Snippet: Primary chondrocyte cultures were treated with 1 μmol/l or 10 μmol/l of the Src family kinase inhibitor PP2 (Calbiochem; VWR, Mississauga, Ontario, Canada) or with dimethyl sulphoxide (DMSO; Sigma), the vehicle in which PP2 was dissolved, as control.

    Techniques: Incubation, Staining

    PP2 promotes cell rounding and cortical actin formation. Primary mouse chondrocytes were incubated for 24 hours with (a) dimethyl sulphoxide or (b) the Src inhibitor PP2 (10 μmol/l), and cells were stained with rhodamine-phalloidin (red) for polymerized actin and with DAPI for nuclei (blue). PP2 induced cell rounding, loss of stress fibre, and cortical organization of actin (scale bar: 2 μm).

    Journal: Arthritis Research & Therapy

    Article Title: Src kinase inhibition promotes the chondrocyte phenotype

    doi: 10.1186/ar2308

    Figure Lengend Snippet: PP2 promotes cell rounding and cortical actin formation. Primary mouse chondrocytes were incubated for 24 hours with (a) dimethyl sulphoxide or (b) the Src inhibitor PP2 (10 μmol/l), and cells were stained with rhodamine-phalloidin (red) for polymerized actin and with DAPI for nuclei (blue). PP2 induced cell rounding, loss of stress fibre, and cortical organization of actin (scale bar: 2 μm).

    Article Snippet: Primary chondrocyte cultures were treated with 1 μmol/l or 10 μmol/l of the Src family kinase inhibitor PP2 (Calbiochem; VWR, Mississauga, Ontario, Canada) or with dimethyl sulphoxide (DMSO; Sigma), the vehicle in which PP2 was dissolved, as control.

    Techniques: Incubation, Staining

    Interplay between RhoA/ROCK and Src kinase signalling. (a) Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide (DMSO) or the Src inhibitor PP2 (10 μmol/l), and Ras homology A (RhoA) activity was measured using the G-LISA™ kit (Cytoskeleton). No significant differences in RhoA activity were observed upon Src inhibition ( n = 3). (b) Primary mouse chondrocytes were incubated for 1 or 2 days with DMSO or the Rho-associated, coiled-coil containing protein kinase (ROCK) inhibitor Y27632 (10 μmol/l) or the actin-modifying drugs cytochalasin D (1 μmol/l) and jasplakinolide (50 nmol/l). Expression levels of (b) Lyn , (c) Frk and (d) Hck genes were determined using real-time PCR, which demonstrated regulation of all three genes by the employed drugs ( n = 3, * P

    Journal: Arthritis Research & Therapy

    Article Title: Src kinase inhibition promotes the chondrocyte phenotype

    doi: 10.1186/ar2308

    Figure Lengend Snippet: Interplay between RhoA/ROCK and Src kinase signalling. (a) Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide (DMSO) or the Src inhibitor PP2 (10 μmol/l), and Ras homology A (RhoA) activity was measured using the G-LISA™ kit (Cytoskeleton). No significant differences in RhoA activity were observed upon Src inhibition ( n = 3). (b) Primary mouse chondrocytes were incubated for 1 or 2 days with DMSO or the Rho-associated, coiled-coil containing protein kinase (ROCK) inhibitor Y27632 (10 μmol/l) or the actin-modifying drugs cytochalasin D (1 μmol/l) and jasplakinolide (50 nmol/l). Expression levels of (b) Lyn , (c) Frk and (d) Hck genes were determined using real-time PCR, which demonstrated regulation of all three genes by the employed drugs ( n = 3, * P

    Article Snippet: Primary chondrocyte cultures were treated with 1 μmol/l or 10 μmol/l of the Src family kinase inhibitor PP2 (Calbiochem; VWR, Mississauga, Ontario, Canada) or with dimethyl sulphoxide (DMSO; Sigma), the vehicle in which PP2 was dissolved, as control.

    Techniques: Incubation, Activity Assay, Inhibition, Expressing, Real-time Polymerase Chain Reaction

    Src kinase activity regulates chondrocyte cell numbers. Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide (DMSO), (a) the general tyrosine kinase inhibitors Tyr A23, Tyr A25 and Tyr A47, or (b) the Src inhibitor PP2 (1 and 10 μmol/l each). Cell numbers were determined by MTT assay. All inhibitors reduced cell numbers at the 10 μmol/l concentration, but the effects of PP2 were more pronounced ( n = 4; * P

    Journal: Arthritis Research & Therapy

    Article Title: Src kinase inhibition promotes the chondrocyte phenotype

    doi: 10.1186/ar2308

    Figure Lengend Snippet: Src kinase activity regulates chondrocyte cell numbers. Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide (DMSO), (a) the general tyrosine kinase inhibitors Tyr A23, Tyr A25 and Tyr A47, or (b) the Src inhibitor PP2 (1 and 10 μmol/l each). Cell numbers were determined by MTT assay. All inhibitors reduced cell numbers at the 10 μmol/l concentration, but the effects of PP2 were more pronounced ( n = 4; * P

    Article Snippet: Primary chondrocyte cultures were treated with 1 μmol/l or 10 μmol/l of the Src family kinase inhibitor PP2 (Calbiochem; VWR, Mississauga, Ontario, Canada) or with dimethyl sulphoxide (DMSO; Sigma), the vehicle in which PP2 was dissolved, as control.

    Techniques: Activity Assay, Incubation, MTT Assay, Concentration Assay

    PP2 promotes expression of early chondrocyte marker genes. Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide (DMSO) or the Src inhibitor PP2 (10 μmol/l), and transcript levels of early chondrocyte marker genes were determined by real-time PCR. Expression levels of (a) Sox9 , (b) Sox5 , (c) Sox6 , (d) Col2a1 and (e) Acan (aggrecan) were significantly increased upon Src inhibition ( n = 3; ** P

    Journal: Arthritis Research & Therapy

    Article Title: Src kinase inhibition promotes the chondrocyte phenotype

    doi: 10.1186/ar2308

    Figure Lengend Snippet: PP2 promotes expression of early chondrocyte marker genes. Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide (DMSO) or the Src inhibitor PP2 (10 μmol/l), and transcript levels of early chondrocyte marker genes were determined by real-time PCR. Expression levels of (a) Sox9 , (b) Sox5 , (c) Sox6 , (d) Col2a1 and (e) Acan (aggrecan) were significantly increased upon Src inhibition ( n = 3; ** P

    Article Snippet: Primary chondrocyte cultures were treated with 1 μmol/l or 10 μmol/l of the Src family kinase inhibitor PP2 (Calbiochem; VWR, Mississauga, Ontario, Canada) or with dimethyl sulphoxide (DMSO; Sigma), the vehicle in which PP2 was dissolved, as control.

    Techniques: Expressing, Marker, Incubation, Real-time Polymerase Chain Reaction, Inhibition

    PP2 promotes expression of genes involved in glycosaminoglycan synthesis. Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide or the Src inhibitor PP2 (10 μmol/l), and transcript levels of genes involved in glycosaminoglycan synthesis were determined by real-time PCR. Expression levels of (a) Xylt1 , (b) Xylt2 , (c) Chst3 and (d) Chst11 were significantly increased upon Src inhibition ( n = 3; * P

    Journal: Arthritis Research & Therapy

    Article Title: Src kinase inhibition promotes the chondrocyte phenotype

    doi: 10.1186/ar2308

    Figure Lengend Snippet: PP2 promotes expression of genes involved in glycosaminoglycan synthesis. Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide or the Src inhibitor PP2 (10 μmol/l), and transcript levels of genes involved in glycosaminoglycan synthesis were determined by real-time PCR. Expression levels of (a) Xylt1 , (b) Xylt2 , (c) Chst3 and (d) Chst11 were significantly increased upon Src inhibition ( n = 3; * P

    Article Snippet: Primary chondrocyte cultures were treated with 1 μmol/l or 10 μmol/l of the Src family kinase inhibitor PP2 (Calbiochem; VWR, Mississauga, Ontario, Canada) or with dimethyl sulphoxide (DMSO; Sigma), the vehicle in which PP2 was dissolved, as control.

    Techniques: Expressing, Incubation, Real-time Polymerase Chain Reaction, Inhibition

    PP2 represses expression of Src kinase genes. Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide (DMSO) or the Src inhibitor PP2 (10 μmol/l), and transcript levels of Src kinase genes were determined by real-time PCR. Expression levels of (a) Lyn , (b) Frk and (c) Hck were significantly decreased upon Src inhibition ( n = 3; *** P

    Journal: Arthritis Research & Therapy

    Article Title: Src kinase inhibition promotes the chondrocyte phenotype

    doi: 10.1186/ar2308

    Figure Lengend Snippet: PP2 represses expression of Src kinase genes. Primary mouse chondrocytes were incubated for 1 to 3 days with dimethyl sulphoxide (DMSO) or the Src inhibitor PP2 (10 μmol/l), and transcript levels of Src kinase genes were determined by real-time PCR. Expression levels of (a) Lyn , (b) Frk and (c) Hck were significantly decreased upon Src inhibition ( n = 3; *** P

    Article Snippet: Primary chondrocyte cultures were treated with 1 μmol/l or 10 μmol/l of the Src family kinase inhibitor PP2 (Calbiochem; VWR, Mississauga, Ontario, Canada) or with dimethyl sulphoxide (DMSO; Sigma), the vehicle in which PP2 was dissolved, as control.

    Techniques: Expressing, Incubation, Real-time Polymerase Chain Reaction, Inhibition

    Antitumor effects of Src inhibitor PP2 on U251 glioma cells ( A ) Cytotoxic effects of PP2 by MTT assays 24 h after treatment. ( B ) Decreased phosphorylation of Src, Akt, and STAT3 in PP2-treated U251 glioma cells by immunoblots. ( C ) Decrease in NF-κB activity in U251 glioma cells after PP2 by luciferase reporter gene assays. U251 glioma cells were transfected with NF-κB-Luc and β-galactosidase for 24 h before ARP treatment. ( D ) Suppressed migration of U251 glioma cells following PP2 treatment determined by wound-healing assays. * P

    Journal: Oncotarget

    Article Title: Src is the primary target of aripiprazole, an atypical antipsychotic drug, in its anti-tumor action

    doi: 10.18632/oncotarget.23192

    Figure Lengend Snippet: Antitumor effects of Src inhibitor PP2 on U251 glioma cells ( A ) Cytotoxic effects of PP2 by MTT assays 24 h after treatment. ( B ) Decreased phosphorylation of Src, Akt, and STAT3 in PP2-treated U251 glioma cells by immunoblots. ( C ) Decrease in NF-κB activity in U251 glioma cells after PP2 by luciferase reporter gene assays. U251 glioma cells were transfected with NF-κB-Luc and β-galactosidase for 24 h before ARP treatment. ( D ) Suppressed migration of U251 glioma cells following PP2 treatment determined by wound-healing assays. * P

    Article Snippet: ARP (purity > 98%), N-acetyl-L-cysteine, lipopolysaccharide (LPS), cytochalasin B, 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), dimethyl sulfoxide, Z-VAD-FMK, and the kinase inhibitor PP2 were from Sigma (St. Louis, MO, USA).

    Techniques: MTT Assay, Western Blot, Activity Assay, Luciferase, Transfection, Migration

    Lipid raft preparations demonstrating increased the presence of p-PKC α , phosphorylated and total caveolin-1 (p-Cav-1 and t-Cav-1, respectively) and phosphorylated cSrc (p-cSrc) in the same raft fractions 1 h after viral infection (V) as compared to mock infection (M), and the effect of chemical inhibition of cSrc by PP2 (A) and (B), and of PKC α by calphostin C (Cal.C) in (C). (A) Western blot for p-PKC α shows increased signal upon viral infection, with slight reduction upon PP2 treatment (10 µ M, or DMSO control). (B) Western blot for p-Cav-1, with or without pretreatment by PP2 (or DMSO control), stripped and reprobed for p-cSrc and then stripped and reprobed for t-Cav-1, shows increased p-Cav-1 and p-cSrc in the same lipid raft fractions with a reduction in phosphorylation for both in cells pretreated with PP2. t-Cav-1 in the lipid raft fractions was also slightly reduced by PP2 pretreatment. (C) Western blots on cells pretreated with either DMSO control or calphostin C (1 µ M) for 3 h prior to mock or virus infection, and stripped and reprobed as in (B), show a dramatic reduction in p-Cav-1 but not t-Cav-1, and complete abrogation of p-cSrc expression by calphostin C in the same lipid raft fractions.

    Journal: Biochemistry

    Article Title: Protein Kinase C Signaling in Adenoviral Infection

    doi: 10.1021/acs.biochem.6b00858

    Figure Lengend Snippet: Lipid raft preparations demonstrating increased the presence of p-PKC α , phosphorylated and total caveolin-1 (p-Cav-1 and t-Cav-1, respectively) and phosphorylated cSrc (p-cSrc) in the same raft fractions 1 h after viral infection (V) as compared to mock infection (M), and the effect of chemical inhibition of cSrc by PP2 (A) and (B), and of PKC α by calphostin C (Cal.C) in (C). (A) Western blot for p-PKC α shows increased signal upon viral infection, with slight reduction upon PP2 treatment (10 µ M, or DMSO control). (B) Western blot for p-Cav-1, with or without pretreatment by PP2 (or DMSO control), stripped and reprobed for p-cSrc and then stripped and reprobed for t-Cav-1, shows increased p-Cav-1 and p-cSrc in the same lipid raft fractions with a reduction in phosphorylation for both in cells pretreated with PP2. t-Cav-1 in the lipid raft fractions was also slightly reduced by PP2 pretreatment. (C) Western blots on cells pretreated with either DMSO control or calphostin C (1 µ M) for 3 h prior to mock or virus infection, and stripped and reprobed as in (B), show a dramatic reduction in p-Cav-1 but not t-Cav-1, and complete abrogation of p-cSrc expression by calphostin C in the same lipid raft fractions.

    Article Snippet: Inhibitors to PKC (calphostin C) and cSrc (PP2) were purchased from Calbiochem (La Jolla, CA).

    Techniques: Infection, Inhibition, Western Blot, Expressing

    Effect of IL-1RA, PP2 or 3OMS on IL-1β-induced Src kinase in primary cortical neurones. Cultures were treated with vehicle (C) or IL-1β (0.3 U ml −1 ) for 5, 15, 30 or 60 min, in the absence or presence of

    Journal:

    Article Title: Interleukin-1-induced interleukin-6 synthesis is mediated by the neutral sphingomyelinase/Src kinase pathway in neurones

    doi: 10.1038/sj.bjp.0707610

    Figure Lengend Snippet: Effect of IL-1RA, PP2 or 3OMS on IL-1β-induced Src kinase in primary cortical neurones. Cultures were treated with vehicle (C) or IL-1β (0.3 U ml −1 ) for 5, 15, 30 or 60 min, in the absence or presence of

    Article Snippet: To investigate the effect of IL-1β on the activation of MAPKs (extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and p38), Src kinase, CamKII and CREB, on the phosphorylation of NR2B, and on the synthesis of IL-6, primary cultures were incubated with vehicle or IL-1β (0.3 U ml−1 ), in the absence or presence of the Src kinase inhibitor PP2 (10 μ M , Calbiochem, Nottingham, UK), the nSMase inhibitor 3- O -methyl-sphingomyelin (3OMS) (15, 25 μ M , Biomol, USA), the NMDA receptor inhibitors MK-801 (20 μ M ), AP5 (50 μ M ) or ifenprodil (20 μ M ) (Tocris, UK), the Ca2+ /CamKII/IV inhibitor KN-62 (10 μ M ) (Calbiochem, UK) or interleukin-1 receptor antagonist (IL-1RA) (50 ng ml−1 ) at various time points from 5 min to 7 h.

    Techniques:

    Liprin-α1 depletion does not affect invadosome formation, but affects their motility. ( A,C,E ) NIH-Src cells co-transfected with siRNAs and GFP-LifeAct (invadosomes), fixed before ( A , T 0 ) or after incubation with nocodazole and PP2 ( C , T 60 ), and after washout and incubation for 5 min to rescue invadosomes ( E , R 5 ). Immunostaining for GFP-LifeAct (green), tubulin (red), and liprin-α1 (blue). Bars, 20 µm. ( B,D,F ) Graphs: top, mean percentage of cells with/without invadosomes; bottom, invadosome organization: cells classified based on the highest organization of their invadosomes (n = 186–309). χ 2 test: no differences with liprin-α1 and control siRNA; *** p

    Journal: Scientific Reports

    Article Title: Identification of a membrane-less compartment regulating invadosome function and motility

    doi: 10.1038/s41598-018-19447-2

    Figure Lengend Snippet: Liprin-α1 depletion does not affect invadosome formation, but affects their motility. ( A,C,E ) NIH-Src cells co-transfected with siRNAs and GFP-LifeAct (invadosomes), fixed before ( A , T 0 ) or after incubation with nocodazole and PP2 ( C , T 60 ), and after washout and incubation for 5 min to rescue invadosomes ( E , R 5 ). Immunostaining for GFP-LifeAct (green), tubulin (red), and liprin-α1 (blue). Bars, 20 µm. ( B,D,F ) Graphs: top, mean percentage of cells with/without invadosomes; bottom, invadosome organization: cells classified based on the highest organization of their invadosomes (n = 186–309). χ 2 test: no differences with liprin-α1 and control siRNA; *** p

    Article Snippet: 327 was a gift from S. Courtneidge Alexa-488, Alexa-568 and Alexa-647 secondary Abs and Oregon 488-gelatin (Life Technologies, Paisley, Scotland, UK); FITC- and TRITC-conjugated phalloidin (Sigma-Aldrich, St. Louis, MO); goat-anti rabbit nanogold–conjugated secondary Ab (Nanoprobes, Stony Brook, New York), fibronectin (BD Biosciences, San Jose, CA); peroxidase-conjugated anti-rabbit and anti-mouse secondary Abs, and Enhanced Chemiluminescence (ECL) Detection System (Amersham Biosciences, Little Chalfont, UK); Peroxidase-conjugated anti-hamster secondary Ab (Life Technologies); nocodazole and poly-L-lysine hydrobromid (Sigma-Aldrich, St. Louis, MO); PP2 Src kinase inhibitor (Calbiochem) and G418 (Merck, Darmstadt, Germany).

    Techniques: Transfection, Incubation, Immunostaining

    SFKs are necessary for TLR signalling events in human pDCs. ( a – f ) WT or LYN − ( d – f ) CAL-1 cells were pre-treated for 1 h with PP2 (10 μM; a – c ; blue) or DMSO control ( a – c ; black) or left untreated ( d – f ; black: LYN + , orange: LYN − ), before stimulation with R848 for the indicated time periods (minutes). Level of p-IKKα/β and total IKKα/β ( a , d ) as well as IκBα ( b , e ) and GAPDH were assessed by immunoblot. p-NF-κB was determined by flow cytometry. The histogram depicts the 15 min time point ( c , f ). ( g , h ) Human PBMCs were pre-treated as in c and stimulated for 15 min with R848. p-NF-κB ( g ) and p-IRF-7 ( h ) were determined by flow cytometry in gated pDCs. Lines in graphs connect the same donor and histograms depict one representative donor; dotted lines, unstimulated; solid lines, R848 stimulation. ( i ) CAL-1 cells were pre-treated as in c before stimulation with recombinant IFN-β (1,000 U ml −1 ) for the indicated time periods (minutes). p-STAT1 were determined by flow cytometry. Histogram depicts the 30 min time point. Data are representative of two ( b , e , f , i ) or three ( a , c , d ) independent experiments and four ( g ) or five ( h ) donors processed separately. Graphs depict mean±s.d. of replicates within one representative experiment ( a – f , i ) or individual donors ( g , h ). Two-way ANOVA ( c , f , i ) and two-ways Student's t -test ( g , h ) were used for statistical analyses. * P

    Journal: Nature Communications

    Article Title: Src family kinases Fyn and Lyn are constitutively activated and mediate plasmacytoid dendritic cell responses

    doi: 10.1038/ncomms14830

    Figure Lengend Snippet: SFKs are necessary for TLR signalling events in human pDCs. ( a – f ) WT or LYN − ( d – f ) CAL-1 cells were pre-treated for 1 h with PP2 (10 μM; a – c ; blue) or DMSO control ( a – c ; black) or left untreated ( d – f ; black: LYN + , orange: LYN − ), before stimulation with R848 for the indicated time periods (minutes). Level of p-IKKα/β and total IKKα/β ( a , d ) as well as IκBα ( b , e ) and GAPDH were assessed by immunoblot. p-NF-κB was determined by flow cytometry. The histogram depicts the 15 min time point ( c , f ). ( g , h ) Human PBMCs were pre-treated as in c and stimulated for 15 min with R848. p-NF-κB ( g ) and p-IRF-7 ( h ) were determined by flow cytometry in gated pDCs. Lines in graphs connect the same donor and histograms depict one representative donor; dotted lines, unstimulated; solid lines, R848 stimulation. ( i ) CAL-1 cells were pre-treated as in c before stimulation with recombinant IFN-β (1,000 U ml −1 ) for the indicated time periods (minutes). p-STAT1 were determined by flow cytometry. Histogram depicts the 30 min time point. Data are representative of two ( b , e , f , i ) or three ( a , c , d ) independent experiments and four ( g ) or five ( h ) donors processed separately. Graphs depict mean±s.d. of replicates within one representative experiment ( a – f , i ) or individual donors ( g , h ). Two-way ANOVA ( c , f , i ) and two-ways Student's t -test ( g , h ) were used for statistical analyses. * P

    Article Snippet: Cytokine measurements For cytokine measurement in culture supernatants, 1 × 106 total BMDCs, 5 × 104 FACS-purified pDCs, 1 × 105 CAL-1 or 3 × 106 PBMCs were left untreated or pre-treated for 1 h with the SFKs inhibitor PP2 (EMD Millipore), the inhibitor bafetinib (Adooq, Irvine, CA, USA) or the equivalent concentration of DMSO control, followed by 15 h in media in presence or absence of 0.1 μM CpG-B ODN 1668 (Roche), 1 μM CpG-A ODN 2216, 100 μM loxoribine (mouse cells) or 1 μg ml−1 R848 (human cells) (all from InvivoGen, San Diego, CA, USA).

    Techniques: Flow Cytometry, Cytometry, Recombinant

    SFKs are critical for human pDC cytokine production. ( a , b ) CAL-1 cells ( a ) or human PBMCs ( b ) were pre-treated for 1 h with PP2 or DMSO control and then stimulated with R848 for 15 h. Viability was assessed by flow cytometry, while type I IFN and TNF protein levels in the supernatant were assessed by bioassay and ELISA, respectively. ( c – e ) Human PBMCs were pre-treated for 1 h with PP2 (5 and 15 μM) or DMSO control and then stimulated with R848 for 6 h and subsequently intracellular stained for TNF and IFN-α. FACS plots show the co-expression of TNF and IFN-α in gated pDCs ( c ). The percentage of pDCs single positive to IFN-α and TNF staining ( d ). MFI of IFN-α and TNF in gated pDCs ( e ). ( f ) LYN and GAPDH protein expression in unmodified CAL-1 cells (LYN + ) and CAL-1 cells ablated for LYN by CRISPR/Cas9 (LYN − ) was determined by immunoblot. ( g , h ) The expression of IFN-β and TNF mRNAs was determined by qPCR ( g ) and the level of protein in the supernatant was quantified by bioassay or ELISA ( h ) in LYN + and LYN − CAL-1 cells after 1 or 15 h stimulation with R848, respectively. In g , the mRNA expression were normalized against GAPDH expression and plotted as mRNA fold increase compared to unstimulated cells. Data are representative of two ( g ) or three ( a , f , h ) independent experiments or four donors processed separately ( b – e ). Graphs depict mean±s.d. of replicates within one representative experiment/donor. Dunnett's multiple comparisons test ( a , b , d , e ) and two-ways Student's t -test ( f , g ) were used for statistical analyses. * P

    Journal: Nature Communications

    Article Title: Src family kinases Fyn and Lyn are constitutively activated and mediate plasmacytoid dendritic cell responses

    doi: 10.1038/ncomms14830

    Figure Lengend Snippet: SFKs are critical for human pDC cytokine production. ( a , b ) CAL-1 cells ( a ) or human PBMCs ( b ) were pre-treated for 1 h with PP2 or DMSO control and then stimulated with R848 for 15 h. Viability was assessed by flow cytometry, while type I IFN and TNF protein levels in the supernatant were assessed by bioassay and ELISA, respectively. ( c – e ) Human PBMCs were pre-treated for 1 h with PP2 (5 and 15 μM) or DMSO control and then stimulated with R848 for 6 h and subsequently intracellular stained for TNF and IFN-α. FACS plots show the co-expression of TNF and IFN-α in gated pDCs ( c ). The percentage of pDCs single positive to IFN-α and TNF staining ( d ). MFI of IFN-α and TNF in gated pDCs ( e ). ( f ) LYN and GAPDH protein expression in unmodified CAL-1 cells (LYN + ) and CAL-1 cells ablated for LYN by CRISPR/Cas9 (LYN − ) was determined by immunoblot. ( g , h ) The expression of IFN-β and TNF mRNAs was determined by qPCR ( g ) and the level of protein in the supernatant was quantified by bioassay or ELISA ( h ) in LYN + and LYN − CAL-1 cells after 1 or 15 h stimulation with R848, respectively. In g , the mRNA expression were normalized against GAPDH expression and plotted as mRNA fold increase compared to unstimulated cells. Data are representative of two ( g ) or three ( a , f , h ) independent experiments or four donors processed separately ( b – e ). Graphs depict mean±s.d. of replicates within one representative experiment/donor. Dunnett's multiple comparisons test ( a , b , d , e ) and two-ways Student's t -test ( f , g ) were used for statistical analyses. * P

    Article Snippet: Cytokine measurements For cytokine measurement in culture supernatants, 1 × 106 total BMDCs, 5 × 104 FACS-purified pDCs, 1 × 105 CAL-1 or 3 × 106 PBMCs were left untreated or pre-treated for 1 h with the SFKs inhibitor PP2 (EMD Millipore), the inhibitor bafetinib (Adooq, Irvine, CA, USA) or the equivalent concentration of DMSO control, followed by 15 h in media in presence or absence of 0.1 μM CpG-B ODN 1668 (Roche), 1 μM CpG-A ODN 2216, 100 μM loxoribine (mouse cells) or 1 μg ml−1 R848 (human cells) (all from InvivoGen, San Diego, CA, USA).

    Techniques: Flow Cytometry, Cytometry, Enzyme-linked Immunosorbent Assay, Staining, FACS, Expressing, CRISPR, Real-time Polymerase Chain Reaction

    SFKs are necessary for mTOR and BCAP phosphorylation in human pDCs. ( a ) CAL-1 cells were pre-treated for 1 h with PP2 (10 μM; blue) or DMSO control (black) before stimulation with R848 for the indicated time periods (minutes). p-MTOR was determined by flow cytometry. Histograms depict the 15 min time point. ( b ) Human PBMCs were pre-treated as in a and stimulated for 15 min with R848. p-MTOR was determined by flow cytometry in gated pDCs. Lines in graph connect the same donor and the histogram depicts a representative donor; dotted lines, unstimulated; solid lines, R848 stimulation. ( c ) CAL-1 cells were pre-treated as in a and then stimulated with R848 for 15 min. Protein immunoprecipitation was performed using anti-BCAP antibody or isotype control antibody and levels of tyrosine phosphorylation (p-Tyr) and BCAP were assessed by immunoblot. ( d ) CAL-1 cells were pre-treated as in a and then stimulated with R848 for the indicated time periods (minutes). p-AKT Ser473 and AKT levels were determined by immunoblot. Data are representative of two ( d ) or three ( a , c ) independent experiments or five donors processed separately ( b ). Graphs depict mean±s.d. of replicates within one representative experiment ( a , c , d ) or individual donors ( b ). Two-way ANOVA ( a ) and two-ways Student's t -test ( b ) were used for statistical analyses. * P

    Journal: Nature Communications

    Article Title: Src family kinases Fyn and Lyn are constitutively activated and mediate plasmacytoid dendritic cell responses

    doi: 10.1038/ncomms14830

    Figure Lengend Snippet: SFKs are necessary for mTOR and BCAP phosphorylation in human pDCs. ( a ) CAL-1 cells were pre-treated for 1 h with PP2 (10 μM; blue) or DMSO control (black) before stimulation with R848 for the indicated time periods (minutes). p-MTOR was determined by flow cytometry. Histograms depict the 15 min time point. ( b ) Human PBMCs were pre-treated as in a and stimulated for 15 min with R848. p-MTOR was determined by flow cytometry in gated pDCs. Lines in graph connect the same donor and the histogram depicts a representative donor; dotted lines, unstimulated; solid lines, R848 stimulation. ( c ) CAL-1 cells were pre-treated as in a and then stimulated with R848 for 15 min. Protein immunoprecipitation was performed using anti-BCAP antibody or isotype control antibody and levels of tyrosine phosphorylation (p-Tyr) and BCAP were assessed by immunoblot. ( d ) CAL-1 cells were pre-treated as in a and then stimulated with R848 for the indicated time periods (minutes). p-AKT Ser473 and AKT levels were determined by immunoblot. Data are representative of two ( d ) or three ( a , c ) independent experiments or five donors processed separately ( b ). Graphs depict mean±s.d. of replicates within one representative experiment ( a , c , d ) or individual donors ( b ). Two-way ANOVA ( a ) and two-ways Student's t -test ( b ) were used for statistical analyses. * P

    Article Snippet: Cytokine measurements For cytokine measurement in culture supernatants, 1 × 106 total BMDCs, 5 × 104 FACS-purified pDCs, 1 × 105 CAL-1 or 3 × 106 PBMCs were left untreated or pre-treated for 1 h with the SFKs inhibitor PP2 (EMD Millipore), the inhibitor bafetinib (Adooq, Irvine, CA, USA) or the equivalent concentration of DMSO control, followed by 15 h in media in presence or absence of 0.1 μM CpG-B ODN 1668 (Roche), 1 μM CpG-A ODN 2216, 100 μM loxoribine (mouse cells) or 1 μg ml−1 R848 (human cells) (all from InvivoGen, San Diego, CA, USA).

    Techniques: Flow Cytometry, Cytometry, Immunoprecipitation

    Effect of tumor primary culture (TPC) conditioned medium (CM) and leukemia inhibitory factor (LIF)-neutralizing antibody. (a) Stat3 (signal transduction and activators of transcription 3) and phospho-Stat3 (p-Stat3) in HC11 and LM3 cells treated with increasing concentrations of CM (30%, 50% and 80%). (b) Stat3 and p-Stat3 levels in HC11 and TPC cells treated with CM that had been preincubated with or without LIF-neutralizing antibody. (c) Phosphorylation levels of Stat3 and extracellular signal-regulated kinase (ERK)1/2 in HC11 cells treated with LIF (3 and 5 ng/ml) preincubated with or without LIF-neutralizing antibody. (d) Expression of CCAAT-enhancer-binding protein (C/EBP)δ in TPC cells treated with CM with or without neutralizing antibody. (e) Phosphorylation of Stat3 and ERK1/2 in HC11 cells treated with LIF with or without the MAP kinase/ERK kinase inhibitor PD98059. (f) p-Stat3 and Stat3 in HC11 and TPC cells treated with LIF with or without the Src-specific inhibitor PP2. Experiments were repeated at least three times with similar results. No effect was observed on phosphorylation levels of either Stat3 or ERK1/2 when HC11 cells were treated with the PD98059 vehicle, dimethyl sulfoxide (data not shown).

    Journal: Breast cancer research : BCR

    Article Title: Mouse mammary tumors display Stat3 activation dependent on leukemia inhibitory factor signaling

    doi: 10.1186/bcr1777

    Figure Lengend Snippet: Effect of tumor primary culture (TPC) conditioned medium (CM) and leukemia inhibitory factor (LIF)-neutralizing antibody. (a) Stat3 (signal transduction and activators of transcription 3) and phospho-Stat3 (p-Stat3) in HC11 and LM3 cells treated with increasing concentrations of CM (30%, 50% and 80%). (b) Stat3 and p-Stat3 levels in HC11 and TPC cells treated with CM that had been preincubated with or without LIF-neutralizing antibody. (c) Phosphorylation levels of Stat3 and extracellular signal-regulated kinase (ERK)1/2 in HC11 cells treated with LIF (3 and 5 ng/ml) preincubated with or without LIF-neutralizing antibody. (d) Expression of CCAAT-enhancer-binding protein (C/EBP)δ in TPC cells treated with CM with or without neutralizing antibody. (e) Phosphorylation of Stat3 and ERK1/2 in HC11 cells treated with LIF with or without the MAP kinase/ERK kinase inhibitor PD98059. (f) p-Stat3 and Stat3 in HC11 and TPC cells treated with LIF with or without the Src-specific inhibitor PP2. Experiments were repeated at least three times with similar results. No effect was observed on phosphorylation levels of either Stat3 or ERK1/2 when HC11 cells were treated with the PD98059 vehicle, dimethyl sulfoxide (data not shown).

    Article Snippet: Treatment with Src inhibitor (PP2; Calbiochem) was performed as described previously [ ].

    Techniques: Transduction, Expressing, Binding Assay

    Mechanically induced phosphorylation of the Y667-β-cat conserved E-cadherin interaction site in the mesoderm of Danio . ( a ) Phospho-β-cat labelling in zebrafish marginal cells before epiboly (sphere stage). ( b ) Phospho-β-cat labelling at the start of epiboly (dome stage). ( c ) Phospho-β-cat labelling after blebbistatin treatment that suppressed movements. ( d ) Phospho-β-cat labelling after blebbistatin treatment with epiboly movements rescued by global compression. ( e ) Phospho-β-cat labelling after blebbistatin treatment with epiboly movements rescued by drug washing. ( f ) Phospho-β-cat labelling after blebbistatin treatment rescued by magnetic manipulation of UML-injected embryos leading to epiboly movement resumption. ( g ) Phospho-β-cat labelling in the presence of PP2 Src-family inhibitor treatment at dome (associated β-cat nuclear translocation tests in Supplementary Fig. S15b ). ( h ) β-cat labelling in blebbistatin globally compressed embryos treated with PP2. ( i ) β-cat labelling in blebbistatin magnetically deformed UML-injected embryos treated with PP2. Associated quantitative results in Supplementary Fig. S15b . ( j ) Levels of pY667 β-cat in the margin relative to the blastoderm centre and in the margin relative to the background in sphere stage ( n =6), dome stage ( n =9), PP2 treated ( n =7), blebbistatin-treated ( n =22), blebbistatin-treated and washed ( n =7), and blebbistatin-treated UML-injected magnetically rescued embryos ( n =9). Differences between control dome, blebbistatin-washed or blebbistatin-compressed embryos and all other conditions are statistically significant ( P

    Journal: Nature Communications

    Article Title: Evolutionary conservation of early mesoderm specification by mechanotransduction in Bilateria

    doi: 10.1038/ncomms3821

    Figure Lengend Snippet: Mechanically induced phosphorylation of the Y667-β-cat conserved E-cadherin interaction site in the mesoderm of Danio . ( a ) Phospho-β-cat labelling in zebrafish marginal cells before epiboly (sphere stage). ( b ) Phospho-β-cat labelling at the start of epiboly (dome stage). ( c ) Phospho-β-cat labelling after blebbistatin treatment that suppressed movements. ( d ) Phospho-β-cat labelling after blebbistatin treatment with epiboly movements rescued by global compression. ( e ) Phospho-β-cat labelling after blebbistatin treatment with epiboly movements rescued by drug washing. ( f ) Phospho-β-cat labelling after blebbistatin treatment rescued by magnetic manipulation of UML-injected embryos leading to epiboly movement resumption. ( g ) Phospho-β-cat labelling in the presence of PP2 Src-family inhibitor treatment at dome (associated β-cat nuclear translocation tests in Supplementary Fig. S15b ). ( h ) β-cat labelling in blebbistatin globally compressed embryos treated with PP2. ( i ) β-cat labelling in blebbistatin magnetically deformed UML-injected embryos treated with PP2. Associated quantitative results in Supplementary Fig. S15b . ( j ) Levels of pY667 β-cat in the margin relative to the blastoderm centre and in the margin relative to the background in sphere stage ( n =6), dome stage ( n =9), PP2 treated ( n =7), blebbistatin-treated ( n =22), blebbistatin-treated and washed ( n =7), and blebbistatin-treated UML-injected magnetically rescued embryos ( n =9). Differences between control dome, blebbistatin-washed or blebbistatin-compressed embryos and all other conditions are statistically significant ( P

    Article Snippet: Zebrafish Src-kinase inhibition Src family kinases were inhibited by the chemical inhibitor PP2 purchased from Sigma Aldrich.

    Techniques: Injection, Translocation Assay

    OxLDL-induced Vav1 phosphorylation is mediated by src kinase Fyn. (A) Human platelets were incubated with the broad-spectrum src inhibitor AG1879 before incubation with 50 μg/mL oxLDL. The platelet lysates were then analyzed as in for

    Journal: Blood

    Article Title: Vav guanine nucleotide exchange factors link hyperlipidemia and a prothrombotic state

    doi: 10.1182/blood-2009-01-201970

    Figure Lengend Snippet: OxLDL-induced Vav1 phosphorylation is mediated by src kinase Fyn. (A) Human platelets were incubated with the broad-spectrum src inhibitor AG1879 before incubation with 50 μg/mL oxLDL. The platelet lysates were then analyzed as in for

    Article Snippet: The broad-spectrum src kinase inhibitor AG1879 was purchased from Calbiochem.

    Techniques: Incubation

    Phosphatidylinositol 3 (PI3) kinase activity mediates oxidative stress-induced phosphorylation of c-Src. A : Caco-2 cell monolayers were pretreated with wortmannin (0.1 μM), LY294002 (25 μM), or 4-amino-5[chlorophyll]-7-[t-butyl]pyrazolo[3–4-d]pyrimidine

    Journal: American Journal of Physiology - Gastrointestinal and Liver Physiology

    Article Title: Hydrogen peroxide activates focal adhesion kinase and c-Src by a phosphatidylinositol 3 kinase-dependent mechanism and promotes cell migration in Caco-2 cell monolayers

    doi: 10.1152/ajpgi.00368.2009

    Figure Lengend Snippet: Phosphatidylinositol 3 (PI3) kinase activity mediates oxidative stress-induced phosphorylation of c-Src. A : Caco-2 cell monolayers were pretreated with wortmannin (0.1 μM), LY294002 (25 μM), or 4-amino-5[chlorophyll]-7-[t-butyl]pyrazolo[3–4-d]pyrimidine

    Article Snippet: LY294002, Akt inhibitor VIII, and 4-amino-5[chlorophyll]-7-[t-butyl]pyrazolo[3–4-d]pyrimidine (PP2) were obtained from Calbiochem (San Diego, CA).

    Techniques: Activity Assay