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  • 86
    Thermo Fisher gene exp ngb mm00452101 m1
    <t>Ngb-IR</t> in cortical areas. In the piriform cortex Ngb-IR was only seen in few cells of WT mice ( A-B ). Conversely, intense Ngb-IR was seen in cell bodies and processes throughout the piriform cortex of Tg mice ( C-D ). In WT mice Ngb-IR was confined to 2–3 cell layers of the S1 ( E-F ) whereas in Tg mice Ngb-IR was seen scatter throughout all cortical layers ( G-H ). Black arrow exemplifies an Ngb over expressing cell body and black arrowhead over expressing processes. Abbreviations: Caudate putamen (CPu); Cingulate cortex, area 2 (Cg2); Corpus callosum (cc); Dysgranular insular cortex (DI); Piriform cortex (Pir); Primary somatosensory cortex (S1); Ventral hippocampal commissure (vhc). Scale bar 100 µm.
    Gene Exp Ngb Mm00452101 M1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore ngb 2904
    Dopamine receptor D3 (D3R) antagonist <t>NGB</t> 2904 treatment alone induced depressive-like behavior. (A) NGB 2904 treatment alone resulted in a lower body weight than in the vehicle group 24 h after treatment. (B) NGB 2904 treatment alone did not affect the locomotor activity. (C) NGB 2904 treatment alone induced an increase in the immobility time in the forced swim test (FST). (D) NGB 2904 treatment alone failed to affect immobility time in the tail suspension test (TST). The data are presented as the mean ± SEM. * P
    Ngb 2904, supplied by Millipore, used in various techniques. Bioz Stars score: 92/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology anti ngb
    Effect of <t>NGB</t> on H 2 O 2 - and Pb(IV)-induced apoptosis. (A ) Typical cytograms of vehicle–, E2- (10 nM), H 2 O 2 - (400 μM), and Pb(IV)- (200 μM) treated MCF-7 cells for 24 h (left) and relative analyses (right) obtained from <t>Annexin</t> V-FITC with PI assays. Data of viable (PI and Annexin V-FITC double negative) and dead (Annexin V-FITC positive and PI AnnexinV-FITC double positive) cells are means ± SD of three different experiments. P
    Anti Ngb, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 86/100, based on 20 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore rabbit polyclonal anti ngb
    Effect of <t>NGB</t> on H 2 O 2 - and Pb(IV)-induced apoptosis. (A ) Typical cytograms of vehicle–, E2- (10 nM), H 2 O 2 - (400 μM), and Pb(IV)- (200 μM) treated MCF-7 cells for 24 h (left) and relative analyses (right) obtained from <t>Annexin</t> V-FITC with PI assays. Data of viable (PI and Annexin V-FITC double negative) and dead (Annexin V-FITC positive and PI AnnexinV-FITC double positive) cells are means ± SD of three different experiments. P
    Rabbit Polyclonal Anti Ngb, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 34 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Strategic BioSolutions Inc ngb antibody
    Effect of <t>NGB</t> on H 2 O 2 - and Pb(IV)-induced apoptosis. (A ) Typical cytograms of vehicle–, E2- (10 nM), H 2 O 2 - (400 μM), and Pb(IV)- (200 μM) treated MCF-7 cells for 24 h (left) and relative analyses (right) obtained from <t>Annexin</t> V-FITC with PI assays. Data of viable (PI and Annexin V-FITC double negative) and dead (Annexin V-FITC positive and PI AnnexinV-FITC double positive) cells are means ± SD of three different experiments. P
    Ngb Antibody, supplied by Strategic BioSolutions Inc, used in various techniques. Bioz Stars score: 88/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Kaneka Corp ngb antibody
    Effect of <t>NGB</t> on H 2 O 2 - and Pb(IV)-induced apoptosis. (A ) Typical cytograms of vehicle–, E2- (10 nM), H 2 O 2 - (400 μM), and Pb(IV)- (200 μM) treated MCF-7 cells for 24 h (left) and relative analyses (right) obtained from <t>Annexin</t> V-FITC with PI assays. Data of viable (PI and Annexin V-FITC double negative) and dead (Annexin V-FITC positive and PI AnnexinV-FITC double positive) cells are means ± SD of three different experiments. P
    Ngb Antibody, supplied by Kaneka Corp, used in various techniques. Bioz Stars score: 88/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cambridge Biomedical ngb
    Effect of <t>NGB</t> on H 2 O 2 - and Pb(IV)-induced apoptosis. (A ) Typical cytograms of vehicle–, E2- (10 nM), H 2 O 2 - (400 μM), and Pb(IV)- (200 μM) treated MCF-7 cells for 24 h (left) and relative analyses (right) obtained from <t>Annexin</t> V-FITC with PI assays. Data of viable (PI and Annexin V-FITC double negative) and dead (Annexin V-FITC positive and PI AnnexinV-FITC double positive) cells are means ± SD of three different experiments. P
    Ngb, supplied by Cambridge Biomedical, used in various techniques. Bioz Stars score: 90/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore ngb sirna plasmids
    Silencing of <t>Ngb</t> induces caspase-3 activation and inhibits 14-3-3γ expression. (A) A Western blot analysis demonstrates that cleaved caspase-3 increased significantly in <t>N2a/Ngb-siRNA</t> cells. (B) RT-PCR demonstrates that the expression level of 14-3-3γ mRNA decreased significantly in N2a/Ngb-siRNA cells. (C) Western blot analysis demonstrates that the expression of 14-3-3γ protein decreased significantly in N2a/Ngb-siRNA cells. (D) A WST-8 assay demonstrates that overexpression of 14-3-3γ significantly enhanced the viability of N2a/Ngb-siRNA cells following treatment with 150 μmol/L of H 2 O 2 for 12 h compared with vector controls (p-EGFP-N1). All statistical results are expressed as means±SEM from three independent experiments. b P
    Ngb Sirna Plasmids, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 22 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abnova antibodies against ngb
    Interaction between <t>Ngb</t> and <t>VDAC</t>
    Antibodies Against Ngb, supplied by Abnova, used in various techniques. Bioz Stars score: 88/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore antibodies against ngb
    Upregulation and accumulation of <t>Ngb</t> in ischemic neurons are associated with axon regeneration during ischemic reperfusion. a Representative photograph showing the ipsilateral ischemic penumbra (Ipsi) and its contralateral counterpart (Contra) (indicated by squares) in the ischemic mouse brain (TTC staining) after I/R (tMCAo-1 h/reperfusion-24 h). b Western blotting analysis of Ngb and <t>GAP43</t> in the mouse brain after I/R. Adult mice were subjected to 1 h of tMCAo and various times (0.5, 1, 2, 3, 7 days) of reperfusion. Total soluble proteins extracted from cerebral cortices of Ipsi were subjected to western blotting analysis with anti-Ngb/GAP43/β-tubulin antibodies. Relative Ngb or GAP43 expression level was expressed as Ngb/β-tubulin or GAP43/β-tubulin normalized to that of sham. Data are presented as means ± S.E.M. * P
    Antibodies Against Ngb, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore anti neuroglobin antibody
    <t>Neuroglobin</t> (Ngb) expression and infarct distribution. Ngb expression (brown) and infarct distribution on Haematoxylin counter stained sections 24 hours after permanent middle cerebral artery occlusion (pMCAo). A. Uninjured, B. wild-type (WT) pMCAo and C. Ngb-null pMCAo. Note that there is no Ngb staining in Ngb-null mice, and that staining in cortex is very limited. The infarct is marked with red lines on the sections within red squares, which are shown in higher magnification in Figure 2 .
    Anti Neuroglobin Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam anti ngb antibody
    <t>NGB</t> regulates the phosphorylation of <t>AKT</t> in glioma cells. (A) qRT-PCR shows no changes of NGB mRNA levels after IGF-1 (100 ng/ml) treatment for 6 h in U87 cells. Error bars indicate SEM. N=3. (B and C) Representative images of western blots and quantification show no changes of NGB protein levels after IGF-1 (100 ng/ml) treatment for 6 h in U87 cells. Error bars indicate SEM, **p
    Anti Ngb Antibody, supplied by Abcam, used in various techniques. Bioz Stars score: 91/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    MyBiosource chicken anti ngb
    <t>VEGF</t> expression in wild type and <t>Ngb-overexpressing</t> transgenic cultures
    Chicken Anti Ngb, supplied by MyBiosource, used in various techniques. Bioz Stars score: 85/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Thermo Fisher mouse polyclonal antibodies against ngb
    Neuroglobin <t>(Ngb)</t> interacts with Dvl1 in SKNSH cells. ( A ) HA-Ngb was co-transfected with Myc-Dvl1 in SK-N-SH cells. Cells were harvested after 24 h post-transfection. Cells extract (600 μg) were precipitated by anti-Myc rabbit <t>polyclonal</t> antibody or control Immunoglobulin G (IgG). Western blot was used to detect Myc-Dvl1 and HA-Ngb. ( B ) Cells extract (600 μg) were prepared and immunoprecipitation (IP) with rabbit anti-Dvl1 antibody or control rabbit IgG. Western blot was used to detect Dvl1 and Ngb. ( C ) Dvl1(1-250), Dvl1(1-378), Dvl1(337-670) or Myc-Dvl1 was co-transfected with HA-Ngb. After 24 h post-transfection, cells were harvested and IP with mouse anti-Myc monoclonal antibody. Western blot was used to detect Ngb. ( D ) SK-N-SH cells were seeded in 6-well plate. The mouse anti-Ngb monoclonal antibody and Texas Red-conjugated anti-mouse IgG (red) were used to detect Ngb protein. The rabbit anti-Dvl1 monoclonal antibody and Texas Green-conjugated anti-rabbit IgG were used to detect Dvl1 protein. Nuclei were stained by Hoechst 33258. The merged image showed the co-localization of Dvl1 and Ngb. ( E ) SK-N-SH cells were co-transfected with Myc-Dvl1 and HA-Ngb. The rabbit anti-HA polyclonal antibody and Texas Green-conjugated anti-rabbit IgG were used to detect HA-Ngb protein. The mouse anti-Myc monoclonal antibody and Texas Red-conjugated anti-rabbit IgG were used to detect Myc-Dvl1 protein. Nuclei were stained by Hoechst 33258. The merged image showed the co-localization of Myc-Dvl1 and HA-Ngb.
    Mouse Polyclonal Antibodies Against Ngb, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Genoway ngb null mouse model
    Neuroglobin <t>(Ngb)</t> interacts with Dvl1 in SKNSH cells. ( A ) HA-Ngb was co-transfected with Myc-Dvl1 in SK-N-SH cells. Cells were harvested after 24 h post-transfection. Cells extract (600 μg) were precipitated by anti-Myc rabbit <t>polyclonal</t> antibody or control Immunoglobulin G (IgG). Western blot was used to detect Myc-Dvl1 and HA-Ngb. ( B ) Cells extract (600 μg) were prepared and immunoprecipitation (IP) with rabbit anti-Dvl1 antibody or control rabbit IgG. Western blot was used to detect Dvl1 and Ngb. ( C ) Dvl1(1-250), Dvl1(1-378), Dvl1(337-670) or Myc-Dvl1 was co-transfected with HA-Ngb. After 24 h post-transfection, cells were harvested and IP with mouse anti-Myc monoclonal antibody. Western blot was used to detect Ngb. ( D ) SK-N-SH cells were seeded in 6-well plate. The mouse anti-Ngb monoclonal antibody and Texas Red-conjugated anti-mouse IgG (red) were used to detect Ngb protein. The rabbit anti-Dvl1 monoclonal antibody and Texas Green-conjugated anti-rabbit IgG were used to detect Dvl1 protein. Nuclei were stained by Hoechst 33258. The merged image showed the co-localization of Dvl1 and Ngb. ( E ) SK-N-SH cells were co-transfected with Myc-Dvl1 and HA-Ngb. The rabbit anti-HA polyclonal antibody and Texas Green-conjugated anti-rabbit IgG were used to detect HA-Ngb protein. The mouse anti-Myc monoclonal antibody and Texas Red-conjugated anti-rabbit IgG were used to detect Myc-Dvl1 protein. Nuclei were stained by Hoechst 33258. The merged image showed the co-localization of Myc-Dvl1 and HA-Ngb.
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    GenScript ngb protein
    Neuroprotective effect of <t>Ngb</t> in an in vivo model of ocular hypertension. A–D: Representative epifluorescence photomicrographs of retinal flat mounts immunolabeled by <t>Tuj-1</t> antibody (RGC marker). The retinas were taken from the eyes of WT ( A and
    Ngb Protein, supplied by GenScript, used in various techniques. Bioz Stars score: 85/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc ngb
    Neuroprotective effect of <t>Ngb</t> in an in vivo model of ocular hypertension. A–D: Representative epifluorescence photomicrographs of retinal flat mounts immunolabeled by <t>Tuj-1</t> antibody (RGC marker). The retinas were taken from the eyes of WT ( A and
    Ngb, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology ngb
    Silencing of <t>Ngb</t> induces caspase-3 activation and inhibits <t>14-3-3γ</t> expression. (A) A Western blot analysis demonstrates that cleaved caspase-3 increased significantly in N2a/Ngb-siRNA cells. (B) RT-PCR demonstrates that the expression level of 14-3-3γ mRNA decreased significantly in N2a/Ngb-siRNA cells. (C) Western blot analysis demonstrates that the expression of 14-3-3γ protein decreased significantly in N2a/Ngb-siRNA cells. (D) A WST-8 assay demonstrates that overexpression of 14-3-3γ significantly enhanced the viability of N2a/Ngb-siRNA cells following treatment with 150 μmol/L of H 2 O 2 for 12 h compared with vector controls (p-EGFP-N1). All statistical results are expressed as means±SEM from three independent experiments. b P
    Ngb, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 89/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology anti ngb polyclonal rabbit antibody
    Silencing of <t>Ngb</t> induces caspase-3 activation and inhibits <t>14-3-3γ</t> expression. (A) A Western blot analysis demonstrates that cleaved caspase-3 increased significantly in N2a/Ngb-siRNA cells. (B) RT-PCR demonstrates that the expression level of 14-3-3γ mRNA decreased significantly in N2a/Ngb-siRNA cells. (C) Western blot analysis demonstrates that the expression of 14-3-3γ protein decreased significantly in N2a/Ngb-siRNA cells. (D) A WST-8 assay demonstrates that overexpression of 14-3-3γ significantly enhanced the viability of N2a/Ngb-siRNA cells following treatment with 150 μmol/L of H 2 O 2 for 12 h compared with vector controls (p-EGFP-N1). All statistical results are expressed as means±SEM from three independent experiments. b P
    Anti Ngb Polyclonal Rabbit Antibody, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 84/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam monoclonal anti human ngb 13c8 antibody
    Silencing of <t>Ngb</t> induces caspase-3 activation and inhibits <t>14-3-3γ</t> expression. (A) A Western blot analysis demonstrates that cleaved caspase-3 increased significantly in N2a/Ngb-siRNA cells. (B) RT-PCR demonstrates that the expression level of 14-3-3γ mRNA decreased significantly in N2a/Ngb-siRNA cells. (C) Western blot analysis demonstrates that the expression of 14-3-3γ protein decreased significantly in N2a/Ngb-siRNA cells. (D) A WST-8 assay demonstrates that overexpression of 14-3-3γ significantly enhanced the viability of N2a/Ngb-siRNA cells following treatment with 150 μmol/L of H 2 O 2 for 12 h compared with vector controls (p-EGFP-N1). All statistical results are expressed as means±SEM from three independent experiments. b P
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    Santa Cruz Biotechnology polyclonal antibodies against mouse ngb
    Silencing of <t>Ngb</t> induces caspase-3 activation and inhibits <t>14-3-3γ</t> expression. (A) A Western blot analysis demonstrates that cleaved caspase-3 increased significantly in N2a/Ngb-siRNA cells. (B) RT-PCR demonstrates that the expression level of 14-3-3γ mRNA decreased significantly in N2a/Ngb-siRNA cells. (C) Western blot analysis demonstrates that the expression of 14-3-3γ protein decreased significantly in N2a/Ngb-siRNA cells. (D) A WST-8 assay demonstrates that overexpression of 14-3-3γ significantly enhanced the viability of N2a/Ngb-siRNA cells following treatment with 150 μmol/L of H 2 O 2 for 12 h compared with vector controls (p-EGFP-N1). All statistical results are expressed as means±SEM from three independent experiments. b P
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    Image Search Results


    Ngb-IR in cortical areas. In the piriform cortex Ngb-IR was only seen in few cells of WT mice ( A-B ). Conversely, intense Ngb-IR was seen in cell bodies and processes throughout the piriform cortex of Tg mice ( C-D ). In WT mice Ngb-IR was confined to 2–3 cell layers of the S1 ( E-F ) whereas in Tg mice Ngb-IR was seen scatter throughout all cortical layers ( G-H ). Black arrow exemplifies an Ngb over expressing cell body and black arrowhead over expressing processes. Abbreviations: Caudate putamen (CPu); Cingulate cortex, area 2 (Cg2); Corpus callosum (cc); Dysgranular insular cortex (DI); Piriform cortex (Pir); Primary somatosensory cortex (S1); Ventral hippocampal commissure (vhc). Scale bar 100 µm.

    Journal: PLoS ONE

    Article Title: Neuroglobin Over Expressing Mice: Expression Pattern and Effect on Brain Ischemic Infarct Size

    doi: 10.1371/journal.pone.0076565

    Figure Lengend Snippet: Ngb-IR in cortical areas. In the piriform cortex Ngb-IR was only seen in few cells of WT mice ( A-B ). Conversely, intense Ngb-IR was seen in cell bodies and processes throughout the piriform cortex of Tg mice ( C-D ). In WT mice Ngb-IR was confined to 2–3 cell layers of the S1 ( E-F ) whereas in Tg mice Ngb-IR was seen scatter throughout all cortical layers ( G-H ). Black arrow exemplifies an Ngb over expressing cell body and black arrowhead over expressing processes. Abbreviations: Caudate putamen (CPu); Cingulate cortex, area 2 (Cg2); Corpus callosum (cc); Dysgranular insular cortex (DI); Piriform cortex (Pir); Primary somatosensory cortex (S1); Ventral hippocampal commissure (vhc). Scale bar 100 µm.

    Article Snippet: For RT-QPCR cDNA was diluted 1∶1 in replicas of four and expression of the Ngb transcript was quantified, using a Taqman gene expression assay (Life Technologies, Mm00452101_m1).

    Techniques: Mouse Assay, Expressing

    Ngb immunoreactivity (IR) in the forebrain. WT Ngb-IR was mostly confined to a thin layer of the Il and Cg1 ( A-B ). In Tg mice Ngb-IR was seen throughout the forebrain in medium intense stained cells ( C-D ). Intense Ngb-IR could be seen in cell bodies and processes in LS of WT ( E-F ) and Tg mice ( G-H ). In Tg mice weak Ngb-IR could also be seen scattered throughout the CPu in small sized cells. Black arrow exemplifies an over expressing cell. Abbreviations: Anterior commissure, anterior part (aca); Cingulate cortex, area 1 (Cg1); Infralimbic area (Il); Lateral septum (LS); Primary somatosensory cortex, jaw region (SIJ). Scale bar 100 µm.

    Journal: PLoS ONE

    Article Title: Neuroglobin Over Expressing Mice: Expression Pattern and Effect on Brain Ischemic Infarct Size

    doi: 10.1371/journal.pone.0076565

    Figure Lengend Snippet: Ngb immunoreactivity (IR) in the forebrain. WT Ngb-IR was mostly confined to a thin layer of the Il and Cg1 ( A-B ). In Tg mice Ngb-IR was seen throughout the forebrain in medium intense stained cells ( C-D ). Intense Ngb-IR could be seen in cell bodies and processes in LS of WT ( E-F ) and Tg mice ( G-H ). In Tg mice weak Ngb-IR could also be seen scattered throughout the CPu in small sized cells. Black arrow exemplifies an over expressing cell. Abbreviations: Anterior commissure, anterior part (aca); Cingulate cortex, area 1 (Cg1); Infralimbic area (Il); Lateral septum (LS); Primary somatosensory cortex, jaw region (SIJ). Scale bar 100 µm.

    Article Snippet: For RT-QPCR cDNA was diluted 1∶1 in replicas of four and expression of the Ngb transcript was quantified, using a Taqman gene expression assay (Life Technologies, Mm00452101_m1).

    Techniques: Mouse Assay, Staining, Expressing

    Ngb-IR in the penumbra and infarct volumes 24 hours after permanent middle cerebral artery occlusion. Ngb-IR in the cortex of a representative pMCAo operated Ngb-Tg mouse is shown in A . No increase in Ngb-IR intensity was observed in pMCAo operated Ngb-Tg mice when compared to sham-operated Ngb-Tg mice ( B ). The red line marks the penumbra. Black arrows exemplify Ngb-IR neurons and a black arrowhead a necrotic cell. No increase in Ngb expression is seen within nor adjacent to the penumbra area in the pMCAo operated Ngb-Tg animals compared to Tg-sham ( A - B ), which suggests no selective sparing of neurons expressing Ngb. Infarct volumes 24 hours after pMCAo estimated with 2D nucleator and Cavalierís principle is shown in C . The estimated infarct volume in cortex was significantly larger in WT Tg background mice (n = 5, total mean volume 3.5 mm 3 SE±0.77 mm 3 ) compared to Ngb-Tg (n = 7, total mean volume 1.7 mm 3 SE±0.27 mm 3 ) littermates. p

    Journal: PLoS ONE

    Article Title: Neuroglobin Over Expressing Mice: Expression Pattern and Effect on Brain Ischemic Infarct Size

    doi: 10.1371/journal.pone.0076565

    Figure Lengend Snippet: Ngb-IR in the penumbra and infarct volumes 24 hours after permanent middle cerebral artery occlusion. Ngb-IR in the cortex of a representative pMCAo operated Ngb-Tg mouse is shown in A . No increase in Ngb-IR intensity was observed in pMCAo operated Ngb-Tg mice when compared to sham-operated Ngb-Tg mice ( B ). The red line marks the penumbra. Black arrows exemplify Ngb-IR neurons and a black arrowhead a necrotic cell. No increase in Ngb expression is seen within nor adjacent to the penumbra area in the pMCAo operated Ngb-Tg animals compared to Tg-sham ( A - B ), which suggests no selective sparing of neurons expressing Ngb. Infarct volumes 24 hours after pMCAo estimated with 2D nucleator and Cavalierís principle is shown in C . The estimated infarct volume in cortex was significantly larger in WT Tg background mice (n = 5, total mean volume 3.5 mm 3 SE±0.77 mm 3 ) compared to Ngb-Tg (n = 7, total mean volume 1.7 mm 3 SE±0.27 mm 3 ) littermates. p

    Article Snippet: For RT-QPCR cDNA was diluted 1∶1 in replicas of four and expression of the Ngb transcript was quantified, using a Taqman gene expression assay (Life Technologies, Mm00452101_m1).

    Techniques: Mouse Assay, Expressing

    Genotyping and Western blot analysis of Ngb over expression. ( A ) RT-QPCR showed an approximately four fold increase in Ngb mRNA expression in Tg (n = 4, 0.446 SD±0.092) mice compared to WT (n = 8, 0.121 SD±0.048). ( B ) Western blot analysis confirmed the increase in mRNA showing an approximate five-fold increase in Ngb expression in Tg (n = 3, 1.864 SD±0.410) compared to WT (n = 3, 0.317 SD±0.091) mice. WB of ß-actin and Ngb are shown under the graph.

    Journal: PLoS ONE

    Article Title: Neuroglobin Over Expressing Mice: Expression Pattern and Effect on Brain Ischemic Infarct Size

    doi: 10.1371/journal.pone.0076565

    Figure Lengend Snippet: Genotyping and Western blot analysis of Ngb over expression. ( A ) RT-QPCR showed an approximately four fold increase in Ngb mRNA expression in Tg (n = 4, 0.446 SD±0.092) mice compared to WT (n = 8, 0.121 SD±0.048). ( B ) Western blot analysis confirmed the increase in mRNA showing an approximate five-fold increase in Ngb expression in Tg (n = 3, 1.864 SD±0.410) compared to WT (n = 3, 0.317 SD±0.091) mice. WB of ß-actin and Ngb are shown under the graph.

    Article Snippet: For RT-QPCR cDNA was diluted 1∶1 in replicas of four and expression of the Ngb transcript was quantified, using a Taqman gene expression assay (Life Technologies, Mm00452101_m1).

    Techniques: Western Blot, Over Expression, Quantitative RT-PCR, Expressing, Mouse Assay

    Overview of Ngb mRNA and protein expression in WT and Ngb-Tg brain. ( A ) In Situ Hybridization (ISH) and ( B ) immunohistochemistry (IHC) showed distinct and pronounced Ngb expression in hypothalamus and tegmental areas of the brain of both WT and Tg mice. Differential Ngb expression was observed primarily in the cortical areas, hippocampus caudate putamen and cerebellum of Tg mice. Abbreviations: Caudate putamen (CPu); Cerebellum (Cb); Hypothalamus (Hyp); Hippocampus (Hippo); Infralimbic area (Il); Laterodorsal tegmental nucleus (LDTg); Lateral septum (LS); Piriform cortex (Pir); Primary somatosensory cortex (S1).

    Journal: PLoS ONE

    Article Title: Neuroglobin Over Expressing Mice: Expression Pattern and Effect on Brain Ischemic Infarct Size

    doi: 10.1371/journal.pone.0076565

    Figure Lengend Snippet: Overview of Ngb mRNA and protein expression in WT and Ngb-Tg brain. ( A ) In Situ Hybridization (ISH) and ( B ) immunohistochemistry (IHC) showed distinct and pronounced Ngb expression in hypothalamus and tegmental areas of the brain of both WT and Tg mice. Differential Ngb expression was observed primarily in the cortical areas, hippocampus caudate putamen and cerebellum of Tg mice. Abbreviations: Caudate putamen (CPu); Cerebellum (Cb); Hypothalamus (Hyp); Hippocampus (Hippo); Infralimbic area (Il); Laterodorsal tegmental nucleus (LDTg); Lateral septum (LS); Piriform cortex (Pir); Primary somatosensory cortex (S1).

    Article Snippet: For RT-QPCR cDNA was diluted 1∶1 in replicas of four and expression of the Ngb transcript was quantified, using a Taqman gene expression assay (Life Technologies, Mm00452101_m1).

    Techniques: Expressing, In Situ Hybridization, Immunohistochemistry, Mouse Assay

    Ngb-IR in the hypothalamus and hippocampus. No difference could be seen in localization or intensity of Ngb-IR in the hypothalamus of WT ( A-B ) and Tg mice ( C-D ). The hippocampus was voided of Ngb-IR in WT mice (E-F). In Tg mice Ngb-IR was seen in cell bodies (black arrow) and processes of most structures of the hippocampus ( G-H ). Abbreviations: 3 rd ventricle (3V); field CA3 of hippocampus (CA3); Dentate gyrus (DG); Medial habenular nucleus (MHb); Optic chiasm (Ox). Scale bar 100 µm.

    Journal: PLoS ONE

    Article Title: Neuroglobin Over Expressing Mice: Expression Pattern and Effect on Brain Ischemic Infarct Size

    doi: 10.1371/journal.pone.0076565

    Figure Lengend Snippet: Ngb-IR in the hypothalamus and hippocampus. No difference could be seen in localization or intensity of Ngb-IR in the hypothalamus of WT ( A-B ) and Tg mice ( C-D ). The hippocampus was voided of Ngb-IR in WT mice (E-F). In Tg mice Ngb-IR was seen in cell bodies (black arrow) and processes of most structures of the hippocampus ( G-H ). Abbreviations: 3 rd ventricle (3V); field CA3 of hippocampus (CA3); Dentate gyrus (DG); Medial habenular nucleus (MHb); Optic chiasm (Ox). Scale bar 100 µm.

    Article Snippet: For RT-QPCR cDNA was diluted 1∶1 in replicas of four and expression of the Ngb transcript was quantified, using a Taqman gene expression assay (Life Technologies, Mm00452101_m1).

    Techniques: Mouse Assay

    Ngb-IR, GFAP-IR and NeuN-IR in the cerebellum. In A - C Ngb (green) and the astroglia marker GFAP (red) in the cerebellum is shown. No co-localization of Ngb-IR and GFAP-IR was observed ( C ). In D - E Ngb (green) and the neuronal marker NeuN (red) is shown. The intensely Ngb stained Purkinje neurons are not co-localized with NeuN. Scale bar 20 µm in A - C and 50 µm in D - E

    Journal: PLoS ONE

    Article Title: Neuroglobin Over Expressing Mice: Expression Pattern and Effect on Brain Ischemic Infarct Size

    doi: 10.1371/journal.pone.0076565

    Figure Lengend Snippet: Ngb-IR, GFAP-IR and NeuN-IR in the cerebellum. In A - C Ngb (green) and the astroglia marker GFAP (red) in the cerebellum is shown. No co-localization of Ngb-IR and GFAP-IR was observed ( C ). In D - E Ngb (green) and the neuronal marker NeuN (red) is shown. The intensely Ngb stained Purkinje neurons are not co-localized with NeuN. Scale bar 20 µm in A - C and 50 µm in D - E

    Article Snippet: For RT-QPCR cDNA was diluted 1∶1 in replicas of four and expression of the Ngb transcript was quantified, using a Taqman gene expression assay (Life Technologies, Mm00452101_m1).

    Techniques: Marker, Staining

    Ngb-IR in the hindbrain and cerebellum. In the hindbrain pontine nuclei no difference was seen between WT ( A-B ) and Tg ( C-D ) mice in localization, intensity or morphology of the Ngb-IR neurons. No Ngb-IR could be seen in the cerebellum of WT mice ( E-F ). In contrast, Ngb-IR was seen in Purkinje cells (black arrow) and granule cells (black arrowhead) of the cerebellar lobule in Tg mice ( G-H ). Abbreviations: Aqueduct (Aq); Cerebellum (Cb); Laterodorsal tegmental nucleus (LDTg). Scale bar 100 µm.

    Journal: PLoS ONE

    Article Title: Neuroglobin Over Expressing Mice: Expression Pattern and Effect on Brain Ischemic Infarct Size

    doi: 10.1371/journal.pone.0076565

    Figure Lengend Snippet: Ngb-IR in the hindbrain and cerebellum. In the hindbrain pontine nuclei no difference was seen between WT ( A-B ) and Tg ( C-D ) mice in localization, intensity or morphology of the Ngb-IR neurons. No Ngb-IR could be seen in the cerebellum of WT mice ( E-F ). In contrast, Ngb-IR was seen in Purkinje cells (black arrow) and granule cells (black arrowhead) of the cerebellar lobule in Tg mice ( G-H ). Abbreviations: Aqueduct (Aq); Cerebellum (Cb); Laterodorsal tegmental nucleus (LDTg). Scale bar 100 µm.

    Article Snippet: For RT-QPCR cDNA was diluted 1∶1 in replicas of four and expression of the Ngb transcript was quantified, using a Taqman gene expression assay (Life Technologies, Mm00452101_m1).

    Techniques: Mouse Assay

    Ngb-IR, GFAP-IR and NeuN-IR in the piriform cortex. No co-localization could be observed between Ngb-IR ( A , C ) and GFAP-IR ( B , C ) in the piriform cortex. Ngb-IR ( D , F ) and NeuN-IR ( E , F ) was found to be highly co-localized in the piriform cortex. Single white arrows indicate Ngb-IR cells, double-headed white arrows exemplifies GFAP-IR cell. Yellow arrows exemplify Ngb-IR and NeuN-IR co-localized neurons. Scale bar 50 µm

    Journal: PLoS ONE

    Article Title: Neuroglobin Over Expressing Mice: Expression Pattern and Effect on Brain Ischemic Infarct Size

    doi: 10.1371/journal.pone.0076565

    Figure Lengend Snippet: Ngb-IR, GFAP-IR and NeuN-IR in the piriform cortex. No co-localization could be observed between Ngb-IR ( A , C ) and GFAP-IR ( B , C ) in the piriform cortex. Ngb-IR ( D , F ) and NeuN-IR ( E , F ) was found to be highly co-localized in the piriform cortex. Single white arrows indicate Ngb-IR cells, double-headed white arrows exemplifies GFAP-IR cell. Yellow arrows exemplify Ngb-IR and NeuN-IR co-localized neurons. Scale bar 50 µm

    Article Snippet: For RT-QPCR cDNA was diluted 1∶1 in replicas of four and expression of the Ngb transcript was quantified, using a Taqman gene expression assay (Life Technologies, Mm00452101_m1).

    Techniques:

    Dopamine receptor D3 (D3R) antagonist NGB 2904 treatment alone induced depressive-like behavior. (A) NGB 2904 treatment alone resulted in a lower body weight than in the vehicle group 24 h after treatment. (B) NGB 2904 treatment alone did not affect the locomotor activity. (C) NGB 2904 treatment alone induced an increase in the immobility time in the forced swim test (FST). (D) NGB 2904 treatment alone failed to affect immobility time in the tail suspension test (TST). The data are presented as the mean ± SEM. * P

    Journal: International Journal of Neuropsychopharmacology

    Article Title: The Dopamine Receptor D3 Regulates Lipopolysaccharide-Induced Depressive-Like Behavior in Mice

    doi: 10.1093/ijnp/pyy005

    Figure Lengend Snippet: Dopamine receptor D3 (D3R) antagonist NGB 2904 treatment alone induced depressive-like behavior. (A) NGB 2904 treatment alone resulted in a lower body weight than in the vehicle group 24 h after treatment. (B) NGB 2904 treatment alone did not affect the locomotor activity. (C) NGB 2904 treatment alone induced an increase in the immobility time in the forced swim test (FST). (D) NGB 2904 treatment alone failed to affect immobility time in the tail suspension test (TST). The data are presented as the mean ± SEM. * P

    Article Snippet: Drugs LPS from Escherichia coli (L-3129, serotype 0127:B8), PPX, and NGB 2904 were purchased from Sigma-Aldrich.

    Techniques: Activity Assay

    Effect of NGB on H 2 O 2 - and Pb(IV)-induced apoptosis. (A ) Typical cytograms of vehicle–, E2- (10 nM), H 2 O 2 - (400 μM), and Pb(IV)- (200 μM) treated MCF-7 cells for 24 h (left) and relative analyses (right) obtained from Annexin V-FITC with PI assays. Data of viable (PI and Annexin V-FITC double negative) and dead (Annexin V-FITC positive and PI AnnexinV-FITC double positive) cells are means ± SD of three different experiments. P

    Journal: PLoS ONE

    Article Title: Neuroglobin in Breast Cancer Cells: Effect of Hypoxia and Oxidative Stress on Protein Level, Localization, and Anti-Apoptotic Function

    doi: 10.1371/journal.pone.0154959

    Figure Lengend Snippet: Effect of NGB on H 2 O 2 - and Pb(IV)-induced apoptosis. (A ) Typical cytograms of vehicle–, E2- (10 nM), H 2 O 2 - (400 μM), and Pb(IV)- (200 μM) treated MCF-7 cells for 24 h (left) and relative analyses (right) obtained from Annexin V-FITC with PI assays. Data of viable (PI and Annexin V-FITC double negative) and dead (Annexin V-FITC positive and PI AnnexinV-FITC double positive) cells are means ± SD of three different experiments. P

    Article Snippet: Short hairpin RNA (shRNA) of NGB Lentiviral Particles, Control shRNA Lentiviral Particles, anti-poly(ADP ribose) polymerase (PARP-1), anti-NGB, anti-Bcl2 antibodies and Annexin V-FITC Apoptosis Detection Kit were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

    Techniques:

    Silencing of Ngb induces caspase-3 activation and inhibits 14-3-3γ expression. (A) A Western blot analysis demonstrates that cleaved caspase-3 increased significantly in N2a/Ngb-siRNA cells. (B) RT-PCR demonstrates that the expression level of 14-3-3γ mRNA decreased significantly in N2a/Ngb-siRNA cells. (C) Western blot analysis demonstrates that the expression of 14-3-3γ protein decreased significantly in N2a/Ngb-siRNA cells. (D) A WST-8 assay demonstrates that overexpression of 14-3-3γ significantly enhanced the viability of N2a/Ngb-siRNA cells following treatment with 150 μmol/L of H 2 O 2 for 12 h compared with vector controls (p-EGFP-N1). All statistical results are expressed as means±SEM from three independent experiments. b P

    Journal: Acta Pharmacologica Sinica

    Article Title: Silencing neuroglobin enhances neuronal vulnerability to oxidative injury by down-regulating 14-3-3γ

    doi: 10.1038/aps.2009.70

    Figure Lengend Snippet: Silencing of Ngb induces caspase-3 activation and inhibits 14-3-3γ expression. (A) A Western blot analysis demonstrates that cleaved caspase-3 increased significantly in N2a/Ngb-siRNA cells. (B) RT-PCR demonstrates that the expression level of 14-3-3γ mRNA decreased significantly in N2a/Ngb-siRNA cells. (C) Western blot analysis demonstrates that the expression of 14-3-3γ protein decreased significantly in N2a/Ngb-siRNA cells. (D) A WST-8 assay demonstrates that overexpression of 14-3-3γ significantly enhanced the viability of N2a/Ngb-siRNA cells following treatment with 150 μmol/L of H 2 O 2 for 12 h compared with vector controls (p-EGFP-N1). All statistical results are expressed as means±SEM from three independent experiments. b P

    Article Snippet: N2a cells were transfected with Ngb-siRNA plasmids or control vectors, and stable cell lines were screened using a neomycin antibiotic (Sigma, USA) starting at a concentration of 800 μg/mL.

    Techniques: Activation Assay, Expressing, Western Blot, Reverse Transcription Polymerase Chain Reaction, Over Expression, Plasmid Preparation

    Knock-down of Ngb in N2a neuroblastoma cells. (A) Ngb-siRNA plasmids were co-transfected with p-Ngb-EGFP-N1 plasmids into N2a cells. The expression of Ngb-GFP fusion protein (green) was suppressed after Ngb-siRNA transfection (right panel). Bar, 20 μm. (B) Ngb-siRNA and control plasmids (vec) were transfected into N2a cells and stable cell lines were established by neomycin selection. A representative RT-PCR result shows that the expression level of Ngb mRNA decreased in Ngb-siRNA transfected (N2a/Ngb-siRNA) cells compared with vector transfected (N2a/vec) cells. β-actin served as an internal control and p-Ngb-EGFP-N1 plasmids served as positive controls. (C) A representative Western blot analysis shows decreased expression of Ngb protein in N2a/Ngb-siRNA cells. β-actin served as an internal control. All statistical results are expressed as means±SEM from three independent experiments. c P

    Journal: Acta Pharmacologica Sinica

    Article Title: Silencing neuroglobin enhances neuronal vulnerability to oxidative injury by down-regulating 14-3-3γ

    doi: 10.1038/aps.2009.70

    Figure Lengend Snippet: Knock-down of Ngb in N2a neuroblastoma cells. (A) Ngb-siRNA plasmids were co-transfected with p-Ngb-EGFP-N1 plasmids into N2a cells. The expression of Ngb-GFP fusion protein (green) was suppressed after Ngb-siRNA transfection (right panel). Bar, 20 μm. (B) Ngb-siRNA and control plasmids (vec) were transfected into N2a cells and stable cell lines were established by neomycin selection. A representative RT-PCR result shows that the expression level of Ngb mRNA decreased in Ngb-siRNA transfected (N2a/Ngb-siRNA) cells compared with vector transfected (N2a/vec) cells. β-actin served as an internal control and p-Ngb-EGFP-N1 plasmids served as positive controls. (C) A representative Western blot analysis shows decreased expression of Ngb protein in N2a/Ngb-siRNA cells. β-actin served as an internal control. All statistical results are expressed as means±SEM from three independent experiments. c P

    Article Snippet: N2a cells were transfected with Ngb-siRNA plasmids or control vectors, and stable cell lines were screened using a neomycin antibiotic (Sigma, USA) starting at a concentration of 800 μg/mL.

    Techniques: Transfection, Expressing, Stable Transfection, Selection, Reverse Transcription Polymerase Chain Reaction, Plasmid Preparation, Western Blot

    Silencing of Ngb enhances cell death following H 2 O 2 treatment. (A) N2a/Ngb-siRNA and N2a/vec cells were exposed to various concentration of H 2 O 2 for 12 h. A WST-8 assay demonstrates that the viability of N2a/Ngb-siRNA cells significantly decreased compared with N2a/vec cells following treatment with 150 and 300 μmol/L of H 2 O 2 . (B) N2a/Ngb-siRNA and N2a/vec cells were exposed to 150 μmol/L of H 2 O 2 for 12h and the nuclei were stained with Hoechst 33258. Apoptotic nuclei (condensed heavy staining nuclei indicated by arrow heads) increased in N2a/Ngb-siRNA cells after 12 h of H 2 O 2 incubation. Bar, 10 μm. (C) Wild-type N2a cells were transiently transfected with Ngb-siRNA plasmids or control vectors and exposed to various concentrations of H 2 O 2 for 12 h. A WST-8 assay demonstrates that the viability of N2a cells after Ngb-siRNA transfection significantly decreased following treatments with 150 and 250 μmol/L of H 2 O 2 compared with N2a cells transfected with vectors. (D) N2a/Ngb-siRNA and N2a/vec cells were exposed to 150 μmol/L of H 2 O 2 for 0.5, 1, 2, 3, 4, and 5 h. An LDH assay demonstrates that cell death significantly increased in N2a/Ngb-siRNA cells. All statistical results are expressed as means±SEM from at least three independent experiments. b P

    Journal: Acta Pharmacologica Sinica

    Article Title: Silencing neuroglobin enhances neuronal vulnerability to oxidative injury by down-regulating 14-3-3γ

    doi: 10.1038/aps.2009.70

    Figure Lengend Snippet: Silencing of Ngb enhances cell death following H 2 O 2 treatment. (A) N2a/Ngb-siRNA and N2a/vec cells were exposed to various concentration of H 2 O 2 for 12 h. A WST-8 assay demonstrates that the viability of N2a/Ngb-siRNA cells significantly decreased compared with N2a/vec cells following treatment with 150 and 300 μmol/L of H 2 O 2 . (B) N2a/Ngb-siRNA and N2a/vec cells were exposed to 150 μmol/L of H 2 O 2 for 12h and the nuclei were stained with Hoechst 33258. Apoptotic nuclei (condensed heavy staining nuclei indicated by arrow heads) increased in N2a/Ngb-siRNA cells after 12 h of H 2 O 2 incubation. Bar, 10 μm. (C) Wild-type N2a cells were transiently transfected with Ngb-siRNA plasmids or control vectors and exposed to various concentrations of H 2 O 2 for 12 h. A WST-8 assay demonstrates that the viability of N2a cells after Ngb-siRNA transfection significantly decreased following treatments with 150 and 250 μmol/L of H 2 O 2 compared with N2a cells transfected with vectors. (D) N2a/Ngb-siRNA and N2a/vec cells were exposed to 150 μmol/L of H 2 O 2 for 0.5, 1, 2, 3, 4, and 5 h. An LDH assay demonstrates that cell death significantly increased in N2a/Ngb-siRNA cells. All statistical results are expressed as means±SEM from at least three independent experiments. b P

    Article Snippet: N2a cells were transfected with Ngb-siRNA plasmids or control vectors, and stable cell lines were screened using a neomycin antibiotic (Sigma, USA) starting at a concentration of 800 μg/mL.

    Techniques: Concentration Assay, Staining, Incubation, Transfection, Lactate Dehydrogenase Assay

    Interaction between Ngb and VDAC

    Journal: Neurobiology of disease

    Article Title: Neuroglobin Overexpression Inhibits Oxygen-Glucose Deprivation-induced Mitochondrial Permeability Transition Pore Opening in Primary Cultured Mouse Cortical Neurons

    doi: 10.1016/j.nbd.2013.04.015

    Figure Lengend Snippet: Interaction between Ngb and VDAC

    Article Snippet: Next the cells were incubated with primary antibodies against Ngb (mouse poly Ab, 1:100; Abnova) or VDAC (rabbit poly Ab, 1:200; Abcam) at 4°C overnight.

    Techniques:

    Upregulation and accumulation of Ngb in ischemic neurons are associated with axon regeneration during ischemic reperfusion. a Representative photograph showing the ipsilateral ischemic penumbra (Ipsi) and its contralateral counterpart (Contra) (indicated by squares) in the ischemic mouse brain (TTC staining) after I/R (tMCAo-1 h/reperfusion-24 h). b Western blotting analysis of Ngb and GAP43 in the mouse brain after I/R. Adult mice were subjected to 1 h of tMCAo and various times (0.5, 1, 2, 3, 7 days) of reperfusion. Total soluble proteins extracted from cerebral cortices of Ipsi were subjected to western blotting analysis with anti-Ngb/GAP43/β-tubulin antibodies. Relative Ngb or GAP43 expression level was expressed as Ngb/β-tubulin or GAP43/β-tubulin normalized to that of sham. Data are presented as means ± S.E.M. * P

    Journal: Cell Death & Disease

    Article Title: Neuroglobin boosts axon regeneration during ischemic reperfusion via p38 binding and activation depending on oxygen signal

    doi: 10.1038/s41419-017-0260-8

    Figure Lengend Snippet: Upregulation and accumulation of Ngb in ischemic neurons are associated with axon regeneration during ischemic reperfusion. a Representative photograph showing the ipsilateral ischemic penumbra (Ipsi) and its contralateral counterpart (Contra) (indicated by squares) in the ischemic mouse brain (TTC staining) after I/R (tMCAo-1 h/reperfusion-24 h). b Western blotting analysis of Ngb and GAP43 in the mouse brain after I/R. Adult mice were subjected to 1 h of tMCAo and various times (0.5, 1, 2, 3, 7 days) of reperfusion. Total soluble proteins extracted from cerebral cortices of Ipsi were subjected to western blotting analysis with anti-Ngb/GAP43/β-tubulin antibodies. Relative Ngb or GAP43 expression level was expressed as Ngb/β-tubulin or GAP43/β-tubulin normalized to that of sham. Data are presented as means ± S.E.M. * P

    Article Snippet: Antibodies against Ngb, Flag (Sigma, Saint Louis, MO, USA), GAP43, GFAP, NeuN, NF200, p-p38, p38 (Cell Signaling Technology, Boston, USA), β-tubulin, green fluorescent protein (GFP), Cygb, GST, His (Santa Cruz Biotechnology, Santa Cruz, USA), and Tau-1 (Merk Millipore Ltd., Darmstadt, Germany) were purchased.

    Techniques: Staining, Western Blot, Mouse Assay, Expressing

    Ngb is an axon-regeneration inducer in primary cultured neurons after OGD/Re. a Western blotting analysis of Ngb and GAP43 in primary cultures of mouse cerebral cortical neurons. Cultured neurons at DIV 7 were incubated with OGD media at 1% O 2 for 1 h and then re-incubated with normal neurobasal media at 21% O 2 for various times (OGD/Re). Total soluble proteins were extracted and subjected to western blotting analysis with anti-Ngb/Cygb/GAP43/β-tubulin antibodies. Statistical analysis demonstrated that Ngb and GAP43 were significantly upregulated at 6, 12, and 24 h of OGD/Re. Data are presented as means ± S.E.M. * P

    Journal: Cell Death & Disease

    Article Title: Neuroglobin boosts axon regeneration during ischemic reperfusion via p38 binding and activation depending on oxygen signal

    doi: 10.1038/s41419-017-0260-8

    Figure Lengend Snippet: Ngb is an axon-regeneration inducer in primary cultured neurons after OGD/Re. a Western blotting analysis of Ngb and GAP43 in primary cultures of mouse cerebral cortical neurons. Cultured neurons at DIV 7 were incubated with OGD media at 1% O 2 for 1 h and then re-incubated with normal neurobasal media at 21% O 2 for various times (OGD/Re). Total soluble proteins were extracted and subjected to western blotting analysis with anti-Ngb/Cygb/GAP43/β-tubulin antibodies. Statistical analysis demonstrated that Ngb and GAP43 were significantly upregulated at 6, 12, and 24 h of OGD/Re. Data are presented as means ± S.E.M. * P

    Article Snippet: Antibodies against Ngb, Flag (Sigma, Saint Louis, MO, USA), GAP43, GFAP, NeuN, NF200, p-p38, p38 (Cell Signaling Technology, Boston, USA), β-tubulin, green fluorescent protein (GFP), Cygb, GST, His (Santa Cruz Biotechnology, Santa Cruz, USA), and Tau-1 (Merk Millipore Ltd., Darmstadt, Germany) were purchased.

    Techniques: Cell Culture, Western Blot, Incubation

    Neuroglobin (Ngb) expression and infarct distribution. Ngb expression (brown) and infarct distribution on Haematoxylin counter stained sections 24 hours after permanent middle cerebral artery occlusion (pMCAo). A. Uninjured, B. wild-type (WT) pMCAo and C. Ngb-null pMCAo. Note that there is no Ngb staining in Ngb-null mice, and that staining in cortex is very limited. The infarct is marked with red lines on the sections within red squares, which are shown in higher magnification in Figure 2 .

    Journal: Experimental & Translational Stroke Medicine

    Article Title: Reduced infarct size in neuroglobin-null mice after experimental stroke in vivo

    doi: 10.1186/2040-7378-4-15

    Figure Lengend Snippet: Neuroglobin (Ngb) expression and infarct distribution. Ngb expression (brown) and infarct distribution on Haematoxylin counter stained sections 24 hours after permanent middle cerebral artery occlusion (pMCAo). A. Uninjured, B. wild-type (WT) pMCAo and C. Ngb-null pMCAo. Note that there is no Ngb staining in Ngb-null mice, and that staining in cortex is very limited. The infarct is marked with red lines on the sections within red squares, which are shown in higher magnification in Figure 2 .

    Article Snippet: Antibody validation IHC was performed as described above using the following primary antibodies: Ngb was detected with either an in-house made polyclonal rabbit antibody (code# 4836/5, in 1:100.000 dilution), polyclonal rabbit antibody against amino acid 55–70 of human Ngb (code# N7162, Sigma-Aldrich, in 1:500 dilution) or polyclonal goat antibody against the an internal region of human Ngb (code# sc-22001, Santa Cruz Biotechnology, in 1:500 dilution).

    Techniques: Expressing, Staining, Mouse Assay

    NGB regulates the phosphorylation of AKT in glioma cells. (A) qRT-PCR shows no changes of NGB mRNA levels after IGF-1 (100 ng/ml) treatment for 6 h in U87 cells. Error bars indicate SEM. N=3. (B and C) Representative images of western blots and quantification show no changes of NGB protein levels after IGF-1 (100 ng/ml) treatment for 6 h in U87 cells. Error bars indicate SEM, **p

    Journal: Redox Biology

    Article Title: Tumor grade related expression of neuroglobin is negatively regulated by PPARγ and confers antioxidant activity in glioma progression

    doi: 10.1016/j.redox.2017.03.023

    Figure Lengend Snippet: NGB regulates the phosphorylation of AKT in glioma cells. (A) qRT-PCR shows no changes of NGB mRNA levels after IGF-1 (100 ng/ml) treatment for 6 h in U87 cells. Error bars indicate SEM. N=3. (B and C) Representative images of western blots and quantification show no changes of NGB protein levels after IGF-1 (100 ng/ml) treatment for 6 h in U87 cells. Error bars indicate SEM, **p

    Article Snippet: 2.5 Western blotting Standard Western blotting procedures were carried out as previously reported with the following antibodies: anti-4-Hydroxy-2-Nonenal (4-HNE) antibody (ab48506, Abcam, MA, USA; 1:1000) to detect lipid peroxidation, anti-PTEN (ab32199, Abcam, MA, USA; 1:500) and anti-AKT (ab8805, Abcam, MA, USA; 1:1000), anti-NGB antibody (ab37258, Abcam, MA, USA; 1:500), anti-PPARγ (ab45036, Abcam, MA, USA; 1:1000), anti-Phospho-Akt (Ser473) antibody (mAb #4060, Cell signaling, Danvers, MA; 1:1000), anti- Phospho-Akt (T308) (mAb #13038, Cell signaling, Danvers, MA; 1:1000), anti-β-actin (BA0410, Boster, Wuhan, China; 1:1000), anti-GAPDH (A00227, Boster, Wuhan, China; 1:1000).

    Techniques: Quantitative RT-PCR, Western Blot

    PPARγ negatively regulates the expression of NGB. (A and B) Representative images of western blots and quantification show decreased PTEN and increased phosphorylation of AKT in high-grade gliomas (N=21) than those in low-grade gliomas (N=18). Error bars indicate SEM. *p

    Journal: Redox Biology

    Article Title: Tumor grade related expression of neuroglobin is negatively regulated by PPARγ and confers antioxidant activity in glioma progression

    doi: 10.1016/j.redox.2017.03.023

    Figure Lengend Snippet: PPARγ negatively regulates the expression of NGB. (A and B) Representative images of western blots and quantification show decreased PTEN and increased phosphorylation of AKT in high-grade gliomas (N=21) than those in low-grade gliomas (N=18). Error bars indicate SEM. *p

    Article Snippet: 2.5 Western blotting Standard Western blotting procedures were carried out as previously reported with the following antibodies: anti-4-Hydroxy-2-Nonenal (4-HNE) antibody (ab48506, Abcam, MA, USA; 1:1000) to detect lipid peroxidation, anti-PTEN (ab32199, Abcam, MA, USA; 1:500) and anti-AKT (ab8805, Abcam, MA, USA; 1:1000), anti-NGB antibody (ab37258, Abcam, MA, USA; 1:500), anti-PPARγ (ab45036, Abcam, MA, USA; 1:1000), anti-Phospho-Akt (Ser473) antibody (mAb #4060, Cell signaling, Danvers, MA; 1:1000), anti- Phospho-Akt (T308) (mAb #13038, Cell signaling, Danvers, MA; 1:1000), anti-β-actin (BA0410, Boster, Wuhan, China; 1:1000), anti-GAPDH (A00227, Boster, Wuhan, China; 1:1000).

    Techniques: Expressing, Western Blot

    Tumor grade related expression of NGB is negatively correlated with that of PPARγ. (A) qRT-PCR shows a robust expression of NGB mRNA in high-grade gliomas (N=21) compared with low-grade gliomas (N=18). Error bars indicate SEM. *p

    Journal: Redox Biology

    Article Title: Tumor grade related expression of neuroglobin is negatively regulated by PPARγ and confers antioxidant activity in glioma progression

    doi: 10.1016/j.redox.2017.03.023

    Figure Lengend Snippet: Tumor grade related expression of NGB is negatively correlated with that of PPARγ. (A) qRT-PCR shows a robust expression of NGB mRNA in high-grade gliomas (N=21) compared with low-grade gliomas (N=18). Error bars indicate SEM. *p

    Article Snippet: 2.5 Western blotting Standard Western blotting procedures were carried out as previously reported with the following antibodies: anti-4-Hydroxy-2-Nonenal (4-HNE) antibody (ab48506, Abcam, MA, USA; 1:1000) to detect lipid peroxidation, anti-PTEN (ab32199, Abcam, MA, USA; 1:500) and anti-AKT (ab8805, Abcam, MA, USA; 1:1000), anti-NGB antibody (ab37258, Abcam, MA, USA; 1:500), anti-PPARγ (ab45036, Abcam, MA, USA; 1:1000), anti-Phospho-Akt (Ser473) antibody (mAb #4060, Cell signaling, Danvers, MA; 1:1000), anti- Phospho-Akt (T308) (mAb #13038, Cell signaling, Danvers, MA; 1:1000), anti-β-actin (BA0410, Boster, Wuhan, China; 1:1000), anti-GAPDH (A00227, Boster, Wuhan, China; 1:1000).

    Techniques: Expressing, Quantitative RT-PCR

    Overview of the potential mechanisms involved in the DpC-mediated effects on neuroblastoma. DpC increases TNFα expression in neuroblastoma cells, which may (1) activate cytotoxic T cells to destroy tumor cells and/or (2) acts on the TNFα receptor ( TNFR ) to activate down-stream signaling pathways. These include the MAPK/p38/JNK and NF-ĸB signaling cascades, which lead to nuclear transcription of numerous genes, including those that induce apoptosis, as well as cytokines such as TNFα. Activation of TNFR also promotes cleavage of caspase 8, leading to caspase 3 cleavage and subsequent apoptosis. Moreover, DpC is also highly redox active, resulting in the production of reactive oxygen species (ROS) [ 13 ]. The generation of ROS triggers the release of cytochrome c from mitochondria [ 8 ], leading to cleavage of caspase 9, which then also cleaves caspase 3, leading to apoptosis. Further, the increased ROS also leads to upregulation of neuroglobin ( Ngb ) and cytoglobin ( Cygb ) expression as both of these proteins respond to oxidative stress. Together, these molecular effects, which promote apoptosis, could contribute to the anti-cancer activity of DpC in neuroblastoma

    Journal: Journal of Hematology & Oncology

    Article Title: The novel thiosemicarbazone, di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC), inhibits neuroblastoma growth in vitro and in vivo via multiple mechanisms

    doi: 10.1186/s13045-016-0330-x

    Figure Lengend Snippet: Overview of the potential mechanisms involved in the DpC-mediated effects on neuroblastoma. DpC increases TNFα expression in neuroblastoma cells, which may (1) activate cytotoxic T cells to destroy tumor cells and/or (2) acts on the TNFα receptor ( TNFR ) to activate down-stream signaling pathways. These include the MAPK/p38/JNK and NF-ĸB signaling cascades, which lead to nuclear transcription of numerous genes, including those that induce apoptosis, as well as cytokines such as TNFα. Activation of TNFR also promotes cleavage of caspase 8, leading to caspase 3 cleavage and subsequent apoptosis. Moreover, DpC is also highly redox active, resulting in the production of reactive oxygen species (ROS) [ 13 ]. The generation of ROS triggers the release of cytochrome c from mitochondria [ 8 ], leading to cleavage of caspase 9, which then also cleaves caspase 3, leading to apoptosis. Further, the increased ROS also leads to upregulation of neuroglobin ( Ngb ) and cytoglobin ( Cygb ) expression as both of these proteins respond to oxidative stress. Together, these molecular effects, which promote apoptosis, could contribute to the anti-cancer activity of DpC in neuroblastoma

    Article Snippet: Special thanks are given to Dr. Tan-Un for provision of the antibodies against Ngb and Cygb described in the section.

    Techniques: Expressing, Activation Assay, Activity Assay

    a Incubation of DpC (25 μM) with SK-N-LP neuroblastoma cells and HK2 non-tumorigenic, immortalized kidney cells induces Cygb and Ngb expression after a 24-h incubation. In these studies, non-tumorigenic, immortalized cell lines (i.e., MSC, H9c2, or HK2), or neoplastic, neuroblastoma (SK-N-LP) cells, were incubated for either 0, 12 or 24 h/37 °C with either control medium ( Con ; no agent added), Dp44mT (25 μM), or DpC (25 μM) and then flow cytometric analysis performed using Flow Jo 8.8.2. Black , red , and blue lines represent the control, or the cells treated with the iron chelators for 12 and 24 h, respectively. Results shown are typical experiments of three performed. b Western blot analysis of Cygb and Ngb expression in SK-N-LP cells following incubation with control media ( Con ), Dp44mT (25 μM), or DpC (25 μM) for 24 h/37 °C. The bands presented in the blots are representative of three repeats and the lanes have been cropped from raw data images containing all three repeats (raw data shown in Additional File 1 ) for clarity (lanes separated by dotted lines ). Densitometry data in ( b ) are presented as the mean ± SEM ( n = 3). * p

    Journal: Journal of Hematology & Oncology

    Article Title: The novel thiosemicarbazone, di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC), inhibits neuroblastoma growth in vitro and in vivo via multiple mechanisms

    doi: 10.1186/s13045-016-0330-x

    Figure Lengend Snippet: a Incubation of DpC (25 μM) with SK-N-LP neuroblastoma cells and HK2 non-tumorigenic, immortalized kidney cells induces Cygb and Ngb expression after a 24-h incubation. In these studies, non-tumorigenic, immortalized cell lines (i.e., MSC, H9c2, or HK2), or neoplastic, neuroblastoma (SK-N-LP) cells, were incubated for either 0, 12 or 24 h/37 °C with either control medium ( Con ; no agent added), Dp44mT (25 μM), or DpC (25 μM) and then flow cytometric analysis performed using Flow Jo 8.8.2. Black , red , and blue lines represent the control, or the cells treated with the iron chelators for 12 and 24 h, respectively. Results shown are typical experiments of three performed. b Western blot analysis of Cygb and Ngb expression in SK-N-LP cells following incubation with control media ( Con ), Dp44mT (25 μM), or DpC (25 μM) for 24 h/37 °C. The bands presented in the blots are representative of three repeats and the lanes have been cropped from raw data images containing all three repeats (raw data shown in Additional File 1 ) for clarity (lanes separated by dotted lines ). Densitometry data in ( b ) are presented as the mean ± SEM ( n = 3). * p

    Article Snippet: Special thanks are given to Dr. Tan-Un for provision of the antibodies against Ngb and Cygb described in the section.

    Techniques: Incubation, Expressing, Flow Cytometry, Western Blot

    VEGF expression in wild type and Ngb-overexpressing transgenic cultures

    Journal: Neuroscience Letters

    Article Title: Interactions between Vascular Endothelial Growth Factor and Neuroglobin

    doi: 10.1016/j.neulet.2012.05.018

    Figure Lengend Snippet: VEGF expression in wild type and Ngb-overexpressing transgenic cultures

    Article Snippet: Supernatants were collected and protein concentration determined by BCA protein assay (Biorad, Hercules, CA); 50 μg of protein was placed in SDS running buffer with dithiothreitol, incubated for 10 min at 70°C, run on 12% SDS-PAGE for 35 min at 200 V, and transferred to a 0.2-μm PVDF membrane (Thermo Fisher Scientific) at 70 V for 2 h. Membranes were blocked in 5% dry milk and incubated with rabbit anti-VEGF-A (1:500; Santa Cruz Biotechnology, Santa Cruz, CA) or chicken anti-Ngb (1:1000; My Biosource, San Diego, CA) overnight.

    Techniques: Expressing, Transgenic Assay

    Effect of VEGFR2/Flk1 receptor kinase inhibitors on VEGF-induced Ngb protein expression

    Journal: Neuroscience Letters

    Article Title: Interactions between Vascular Endothelial Growth Factor and Neuroglobin

    doi: 10.1016/j.neulet.2012.05.018

    Figure Lengend Snippet: Effect of VEGFR2/Flk1 receptor kinase inhibitors on VEGF-induced Ngb protein expression

    Article Snippet: Supernatants were collected and protein concentration determined by BCA protein assay (Biorad, Hercules, CA); 50 μg of protein was placed in SDS running buffer with dithiothreitol, incubated for 10 min at 70°C, run on 12% SDS-PAGE for 35 min at 200 V, and transferred to a 0.2-μm PVDF membrane (Thermo Fisher Scientific) at 70 V for 2 h. Membranes were blocked in 5% dry milk and incubated with rabbit anti-VEGF-A (1:500; Santa Cruz Biotechnology, Santa Cruz, CA) or chicken anti-Ngb (1:1000; My Biosource, San Diego, CA) overnight.

    Techniques: Expressing

    Effect of MAPK inhibition on VEGF-induced Ngb expression

    Journal: Neuroscience Letters

    Article Title: Interactions between Vascular Endothelial Growth Factor and Neuroglobin

    doi: 10.1016/j.neulet.2012.05.018

    Figure Lengend Snippet: Effect of MAPK inhibition on VEGF-induced Ngb expression

    Article Snippet: Supernatants were collected and protein concentration determined by BCA protein assay (Biorad, Hercules, CA); 50 μg of protein was placed in SDS running buffer with dithiothreitol, incubated for 10 min at 70°C, run on 12% SDS-PAGE for 35 min at 200 V, and transferred to a 0.2-μm PVDF membrane (Thermo Fisher Scientific) at 70 V for 2 h. Membranes were blocked in 5% dry milk and incubated with rabbit anti-VEGF-A (1:500; Santa Cruz Biotechnology, Santa Cruz, CA) or chicken anti-Ngb (1:1000; My Biosource, San Diego, CA) overnight.

    Techniques: Inhibition, Expressing

    VEGF-induced Ngb protein expression

    Journal: Neuroscience Letters

    Article Title: Interactions between Vascular Endothelial Growth Factor and Neuroglobin

    doi: 10.1016/j.neulet.2012.05.018

    Figure Lengend Snippet: VEGF-induced Ngb protein expression

    Article Snippet: Supernatants were collected and protein concentration determined by BCA protein assay (Biorad, Hercules, CA); 50 μg of protein was placed in SDS running buffer with dithiothreitol, incubated for 10 min at 70°C, run on 12% SDS-PAGE for 35 min at 200 V, and transferred to a 0.2-μm PVDF membrane (Thermo Fisher Scientific) at 70 V for 2 h. Membranes were blocked in 5% dry milk and incubated with rabbit anti-VEGF-A (1:500; Santa Cruz Biotechnology, Santa Cruz, CA) or chicken anti-Ngb (1:1000; My Biosource, San Diego, CA) overnight.

    Techniques: Expressing

    Neuroglobin (Ngb) interacts with Dvl1 in SKNSH cells. ( A ) HA-Ngb was co-transfected with Myc-Dvl1 in SK-N-SH cells. Cells were harvested after 24 h post-transfection. Cells extract (600 μg) were precipitated by anti-Myc rabbit polyclonal antibody or control Immunoglobulin G (IgG). Western blot was used to detect Myc-Dvl1 and HA-Ngb. ( B ) Cells extract (600 μg) were prepared and immunoprecipitation (IP) with rabbit anti-Dvl1 antibody or control rabbit IgG. Western blot was used to detect Dvl1 and Ngb. ( C ) Dvl1(1-250), Dvl1(1-378), Dvl1(337-670) or Myc-Dvl1 was co-transfected with HA-Ngb. After 24 h post-transfection, cells were harvested and IP with mouse anti-Myc monoclonal antibody. Western blot was used to detect Ngb. ( D ) SK-N-SH cells were seeded in 6-well plate. The mouse anti-Ngb monoclonal antibody and Texas Red-conjugated anti-mouse IgG (red) were used to detect Ngb protein. The rabbit anti-Dvl1 monoclonal antibody and Texas Green-conjugated anti-rabbit IgG were used to detect Dvl1 protein. Nuclei were stained by Hoechst 33258. The merged image showed the co-localization of Dvl1 and Ngb. ( E ) SK-N-SH cells were co-transfected with Myc-Dvl1 and HA-Ngb. The rabbit anti-HA polyclonal antibody and Texas Green-conjugated anti-rabbit IgG were used to detect HA-Ngb protein. The mouse anti-Myc monoclonal antibody and Texas Red-conjugated anti-rabbit IgG were used to detect Myc-Dvl1 protein. Nuclei were stained by Hoechst 33258. The merged image showed the co-localization of Myc-Dvl1 and HA-Ngb.

    Journal: International Journal of Molecular Sciences

    Article Title: Neuroglobin Regulates Wnt/β-Catenin and NFκB Signaling Pathway through Dvl1

    doi: 10.3390/ijms19072133

    Figure Lengend Snippet: Neuroglobin (Ngb) interacts with Dvl1 in SKNSH cells. ( A ) HA-Ngb was co-transfected with Myc-Dvl1 in SK-N-SH cells. Cells were harvested after 24 h post-transfection. Cells extract (600 μg) were precipitated by anti-Myc rabbit polyclonal antibody or control Immunoglobulin G (IgG). Western blot was used to detect Myc-Dvl1 and HA-Ngb. ( B ) Cells extract (600 μg) were prepared and immunoprecipitation (IP) with rabbit anti-Dvl1 antibody or control rabbit IgG. Western blot was used to detect Dvl1 and Ngb. ( C ) Dvl1(1-250), Dvl1(1-378), Dvl1(337-670) or Myc-Dvl1 was co-transfected with HA-Ngb. After 24 h post-transfection, cells were harvested and IP with mouse anti-Myc monoclonal antibody. Western blot was used to detect Ngb. ( D ) SK-N-SH cells were seeded in 6-well plate. The mouse anti-Ngb monoclonal antibody and Texas Red-conjugated anti-mouse IgG (red) were used to detect Ngb protein. The rabbit anti-Dvl1 monoclonal antibody and Texas Green-conjugated anti-rabbit IgG were used to detect Dvl1 protein. Nuclei were stained by Hoechst 33258. The merged image showed the co-localization of Dvl1 and Ngb. ( E ) SK-N-SH cells were co-transfected with Myc-Dvl1 and HA-Ngb. The rabbit anti-HA polyclonal antibody and Texas Green-conjugated anti-rabbit IgG were used to detect HA-Ngb protein. The mouse anti-Myc monoclonal antibody and Texas Red-conjugated anti-rabbit IgG were used to detect Myc-Dvl1 protein. Nuclei were stained by Hoechst 33258. The merged image showed the co-localization of Myc-Dvl1 and HA-Ngb.

    Article Snippet: Immunocytochemistry SK-N-SH cells were seeded on glass coverslips and cultured for 24 h. Then cells were fixed by 4% paraformaldehyde and co-incubated with rabbit polyclonal antibodies against Dvl1 and mouse polyclonal antibodies against Ngb for 4 h. Green-conjugated anti-rabbit IgG and Red-conjugated anti-mouse IgG (Thermo Fisher Scientific, Waltham, MA, USA) were added into cells and incubated for 2 h. Nucleus was stained with Hoechst 33258 (Sigma, St Louis, MO, USA).

    Techniques: Transfection, Western Blot, Immunoprecipitation, Staining

    Neuroprotective effect of Ngb in an in vivo model of ocular hypertension. A–D: Representative epifluorescence photomicrographs of retinal flat mounts immunolabeled by Tuj-1 antibody (RGC marker). The retinas were taken from the eyes of WT ( A and

    Journal: The American Journal of Pathology

    Article Title: Neuroglobin Is an Endogenous Neuroprotectant for Retinal Ganglion Cells against Glaucomatous Damage

    doi: 10.1016/j.ajpath.2011.08.015

    Figure Lengend Snippet: Neuroprotective effect of Ngb in an in vivo model of ocular hypertension. A–D: Representative epifluorescence photomicrographs of retinal flat mounts immunolabeled by Tuj-1 antibody (RGC marker). The retinas were taken from the eyes of WT ( A and

    Article Snippet: Retinal flat mounts and sections were incubated with a primary antibody against a RGC-specific marker, β-III-tubulin (Tuj-1; Sigma-Aldrich, St. Louis, MO) and/or NGB protein (GenScript, Piscataway, NJ), followed by reaction with a biotin-conjugated secondary antibody.

    Techniques: In Vivo, Immunolabeling, Marker

    Transient increase of Ngb expression in RGCs induced by elevation in IOP. Representative epifluorescence photomicrographs of a retinal section taken from WT mice at 3 days after induction of IOP elevation. The retinal section was triple-labeled by Tuj-1

    Journal: The American Journal of Pathology

    Article Title: Neuroglobin Is an Endogenous Neuroprotectant for Retinal Ganglion Cells against Glaucomatous Damage

    doi: 10.1016/j.ajpath.2011.08.015

    Figure Lengend Snippet: Transient increase of Ngb expression in RGCs induced by elevation in IOP. Representative epifluorescence photomicrographs of a retinal section taken from WT mice at 3 days after induction of IOP elevation. The retinal section was triple-labeled by Tuj-1

    Article Snippet: Retinal flat mounts and sections were incubated with a primary antibody against a RGC-specific marker, β-III-tubulin (Tuj-1; Sigma-Aldrich, St. Louis, MO) and/or NGB protein (GenScript, Piscataway, NJ), followed by reaction with a biotin-conjugated secondary antibody.

    Techniques: Expressing, Mouse Assay, Labeling

    Silencing of Ngb induces caspase-3 activation and inhibits 14-3-3γ expression. (A) A Western blot analysis demonstrates that cleaved caspase-3 increased significantly in N2a/Ngb-siRNA cells. (B) RT-PCR demonstrates that the expression level of 14-3-3γ mRNA decreased significantly in N2a/Ngb-siRNA cells. (C) Western blot analysis demonstrates that the expression of 14-3-3γ protein decreased significantly in N2a/Ngb-siRNA cells. (D) A WST-8 assay demonstrates that overexpression of 14-3-3γ significantly enhanced the viability of N2a/Ngb-siRNA cells following treatment with 150 μmol/L of H 2 O 2 for 12 h compared with vector controls (p-EGFP-N1). All statistical results are expressed as means±SEM from three independent experiments. b P

    Journal: Acta Pharmacologica Sinica

    Article Title: Silencing neuroglobin enhances neuronal vulnerability to oxidative injury by down-regulating 14-3-3γ

    doi: 10.1038/aps.2009.70

    Figure Lengend Snippet: Silencing of Ngb induces caspase-3 activation and inhibits 14-3-3γ expression. (A) A Western blot analysis demonstrates that cleaved caspase-3 increased significantly in N2a/Ngb-siRNA cells. (B) RT-PCR demonstrates that the expression level of 14-3-3γ mRNA decreased significantly in N2a/Ngb-siRNA cells. (C) Western blot analysis demonstrates that the expression of 14-3-3γ protein decreased significantly in N2a/Ngb-siRNA cells. (D) A WST-8 assay demonstrates that overexpression of 14-3-3γ significantly enhanced the viability of N2a/Ngb-siRNA cells following treatment with 150 μmol/L of H 2 O 2 for 12 h compared with vector controls (p-EGFP-N1). All statistical results are expressed as means±SEM from three independent experiments. b P

    Article Snippet: The membranes were blocked with 5% (w /v ) nonfat dried milk in TBST buffer [0.1 mol/L Tris-HCl, pH 8.0, 0.9% (w/v ) NaCl, and 0.1% (v/v ) Tween-20] and probed with rabbit polyclonal antibodies to Ngb (Santa Cruz Biotechnology, USA) and 14-3-3γ (Immuno-Biological Laboratories, Japan) and mouse monoclonal antibody to β-actin (Santa Cruz Biotechnology, USA) in TBST overnight at 4 °C.

    Techniques: Activation Assay, Expressing, Western Blot, Reverse Transcription Polymerase Chain Reaction, Over Expression, Plasmid Preparation