human nt  (Alomone Labs)


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    Alomone Labs human nt
    Human Nt, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    nt3  (Alomone Labs)


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    Alomone Labs nt3
    Nt3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    human nt  (Alomone Labs)


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    Alomone Labs human nt
    Human Nt, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    rabbit polyclonal anti nt3  (Alomone Labs)


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    Alomone Labs rabbit polyclonal anti nt3
    Rabbit Polyclonal Anti Nt3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    neurotrophin 3  (Alomone Labs)


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    Alomone Labs neurotrophin 3
    Neurotrophin 3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    nt 3  (Alomone Labs)


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    Alomone Labs nt 3
    Nt 3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    recombinant human neurotrophin 3 nt 3 protein  (Alomone Labs)


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    Alomone Labs recombinant human neurotrophin 3 nt 3 protein
    (A, D, G) Representative STED (A), TREx (D) and dual-color SMLM (G) images of the ER and ribosomes in axons from neurons expressing GFP-Sec61β and stained for RpS12. (B, E, H) Magnifications and intensity profile lines from merged images for each microscopy method. (C, F, I) Quantification of RpS12 intensity in ER mask, enlarged ER mask, and one-color flipped images, for each microscopy method. (J) Representative STED images and intensity profile line for an axon segment of a neuron transfected as in (A) and co-labeled for puromycilated peptides. (K) Quantification of RpS12 intensity in ER mask and enlarged ER mask with or without high puromycin treatment using dual-color SMLM. (L-N) Schematic representation of split APEX system used to detect ER-ribosome contacts. When the ER protein Sec61β fused to AP module and the ribosomal protein RpL10A fused to EX module interact with each other, APEX is reconstituted and contact sites can be visualized as a biotinylation radius around the interactions (L). Representative images of split APEX assay in distal axons from neurons expressing RpL10A-3xHA-EX and V5-AP-Sec61β (left), or V5-AP-RTN4A as a negative control (right). Expression of constructs are visualized with V5 and HA antibodies, and biotinylation is detected with conjugated Strep-555 (M). Quantification of Strep signal in distal axons from neurons as in (M), and without H2O2 as a negative control for the biotinylation reaction (N). (O) Quantification of axonal ER-bound ribosomes using split APEX assay with RpL10A-3xHA-EX and V5-AP-Sec61β, in neurons stimulated for 30-minutes with BSA (control), BDNF, <t>NT-3</t> or NGF. Individual data points each represent a neuron in (C, F, I, K, N and O). Boxplots show 25/75-percentiles, the median, and whiskers represent min to max in (C, F, I and K). Data are presented as mean values ± SEM in (N, O). ns = not significant, *p < 0.05, **p < 0.01, ***p<0.001 comparing conditions to control using unpaired t-tests or ordinary one-way ANOVA tests. Scale bars represent 1μm (A, D, G, J) and 5μm (M).
    Recombinant Human Neurotrophin 3 Nt 3 Protein, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    recombinant human neurotrophin 3 nt 3 protein - by Bioz Stars, 2023-01
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    Images

    1) Product Images from "Axonal ER tubules regulate local translation via P180/RRBP1-mediated ribosome interactions"

    Article Title: Axonal ER tubules regulate local translation via P180/RRBP1-mediated ribosome interactions

    Journal: bioRxiv

    doi: 10.1101/2022.11.30.518484

    (A, D, G) Representative STED (A), TREx (D) and dual-color SMLM (G) images of the ER and ribosomes in axons from neurons expressing GFP-Sec61β and stained for RpS12. (B, E, H) Magnifications and intensity profile lines from merged images for each microscopy method. (C, F, I) Quantification of RpS12 intensity in ER mask, enlarged ER mask, and one-color flipped images, for each microscopy method. (J) Representative STED images and intensity profile line for an axon segment of a neuron transfected as in (A) and co-labeled for puromycilated peptides. (K) Quantification of RpS12 intensity in ER mask and enlarged ER mask with or without high puromycin treatment using dual-color SMLM. (L-N) Schematic representation of split APEX system used to detect ER-ribosome contacts. When the ER protein Sec61β fused to AP module and the ribosomal protein RpL10A fused to EX module interact with each other, APEX is reconstituted and contact sites can be visualized as a biotinylation radius around the interactions (L). Representative images of split APEX assay in distal axons from neurons expressing RpL10A-3xHA-EX and V5-AP-Sec61β (left), or V5-AP-RTN4A as a negative control (right). Expression of constructs are visualized with V5 and HA antibodies, and biotinylation is detected with conjugated Strep-555 (M). Quantification of Strep signal in distal axons from neurons as in (M), and without H2O2 as a negative control for the biotinylation reaction (N). (O) Quantification of axonal ER-bound ribosomes using split APEX assay with RpL10A-3xHA-EX and V5-AP-Sec61β, in neurons stimulated for 30-minutes with BSA (control), BDNF, NT-3 or NGF. Individual data points each represent a neuron in (C, F, I, K, N and O). Boxplots show 25/75-percentiles, the median, and whiskers represent min to max in (C, F, I and K). Data are presented as mean values ± SEM in (N, O). ns = not significant, *p < 0.05, **p < 0.01, ***p<0.001 comparing conditions to control using unpaired t-tests or ordinary one-way ANOVA tests. Scale bars represent 1μm (A, D, G, J) and 5μm (M).
    Figure Legend Snippet: (A, D, G) Representative STED (A), TREx (D) and dual-color SMLM (G) images of the ER and ribosomes in axons from neurons expressing GFP-Sec61β and stained for RpS12. (B, E, H) Magnifications and intensity profile lines from merged images for each microscopy method. (C, F, I) Quantification of RpS12 intensity in ER mask, enlarged ER mask, and one-color flipped images, for each microscopy method. (J) Representative STED images and intensity profile line for an axon segment of a neuron transfected as in (A) and co-labeled for puromycilated peptides. (K) Quantification of RpS12 intensity in ER mask and enlarged ER mask with or without high puromycin treatment using dual-color SMLM. (L-N) Schematic representation of split APEX system used to detect ER-ribosome contacts. When the ER protein Sec61β fused to AP module and the ribosomal protein RpL10A fused to EX module interact with each other, APEX is reconstituted and contact sites can be visualized as a biotinylation radius around the interactions (L). Representative images of split APEX assay in distal axons from neurons expressing RpL10A-3xHA-EX and V5-AP-Sec61β (left), or V5-AP-RTN4A as a negative control (right). Expression of constructs are visualized with V5 and HA antibodies, and biotinylation is detected with conjugated Strep-555 (M). Quantification of Strep signal in distal axons from neurons as in (M), and without H2O2 as a negative control for the biotinylation reaction (N). (O) Quantification of axonal ER-bound ribosomes using split APEX assay with RpL10A-3xHA-EX and V5-AP-Sec61β, in neurons stimulated for 30-minutes with BSA (control), BDNF, NT-3 or NGF. Individual data points each represent a neuron in (C, F, I, K, N and O). Boxplots show 25/75-percentiles, the median, and whiskers represent min to max in (C, F, I and K). Data are presented as mean values ± SEM in (N, O). ns = not significant, *p < 0.05, **p < 0.01, ***p<0.001 comparing conditions to control using unpaired t-tests or ordinary one-way ANOVA tests. Scale bars represent 1μm (A, D, G, J) and 5μm (M).

    Techniques Used: Expressing, Staining, Microscopy, Transfection, Labeling, Negative Control, Construct

    recombinant human neurotrophin 3 nt 3 protein  (Alomone Labs)


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    Alomone Labs recombinant human neurotrophin 3 nt 3 protein
    Expression of Bdnf isoforms increases over embryonic cortical neuron differentiation concomitant with movement of the gene locus away from the nuclear periphery (A) Schematic of neuronal precursor cell (NPC) differentiation into post-mitotic neurons (PMN). E12.5, embryonic day 12.5. FGF2, fibroblast growth factor 2. <t>NT-3,</t> <t>neurotrophin-3.</t> FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . (B) Expression profile of an NPC-marker, Nestin, and neuronal markers, Map2 and NeuN, in NPCs and PMNs, assessed by qRT-PCR and normalized to NPC. Bars represent mean ± SEM; points show results from different biological replicates . ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001; unpaired t-test (two-tailed). Nestin p = 0.0005, t = 4.707, n = 7, df = 12; Map2 p = 0.0481, t = 2.201, n = 7, df = 12; NeuN p = 0.0001, t = 14.15, n = 3, df = 4. (C) Expression of Bdnf variants and downstream gene Lin7c during differentiation, assessed by qRT-PCR and normalized to NPC levels. Bars represent mean ± SEM; points show results from different biological replicates (n = 7, df = 12). ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001, ∗∗∗∗p< 0.0001, unpaired t-test (two-tailed). Exon I p = 0.0001, t = 5.627; Exon II p = 0.0007, t = 4.549; Exon III p = 0.0001, t = 5.614; Exon IV p = 0.0019, t = 3.972; Exon V p = 0.0029, t = 3.734; Exon VI p = 0.0021, t = 3.899; Exon VIII p = 0.0397, t = 2.307; Exon IXa p = 0.0023, t = 3.846; Lin7c p = 0.0248, t = 2.565. (D) Relocation of the Bdnf gene during neuronal development assessed by DNA-FISH combined with measurements of the distance of the signal from the closest edge of the nucleus. Left panel; representative confocal sections of DNA-FISH showing nuclear localization of Bdnf loci (green) in NPCs and PMNs. Nuclei were stained with DAPI (gray). For each image, the distance between the center of the FISH signal and the edge of the nucleus is indicated. Scale bars, 5 μm. Right panel; scatter dot plot of the distribution of the distance between Bdnf locus and the edge of the DAPI staining. Solid gray lines denote medians. ∗∗∗∗p = 0.0002, Mann-Whitney test (two-tailed). n = 133 (NPC), 123 (PMN) foci across 4 biological replicates. See also <xref ref-type=Figure S1 . " width="250" height="auto" />
    Recombinant Human Neurotrophin 3 Nt 3 Protein, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "A novel intergenic enhancer that regulates Bdnf expression in developing cortical neurons"

    Article Title: A novel intergenic enhancer that regulates Bdnf expression in developing cortical neurons

    Journal: iScience

    doi: 10.1016/j.isci.2022.105695

    Expression of Bdnf isoforms increases over embryonic cortical neuron differentiation concomitant with movement of the gene locus away from the nuclear periphery (A) Schematic of neuronal precursor cell (NPC) differentiation into post-mitotic neurons (PMN). E12.5, embryonic day 12.5. FGF2, fibroblast growth factor 2. NT-3, neurotrophin-3. FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . (B) Expression profile of an NPC-marker, Nestin, and neuronal markers, Map2 and NeuN, in NPCs and PMNs, assessed by qRT-PCR and normalized to NPC. Bars represent mean ± SEM; points show results from different biological replicates . ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001; unpaired t-test (two-tailed). Nestin p = 0.0005, t = 4.707, n = 7, df = 12; Map2 p = 0.0481, t = 2.201, n = 7, df = 12; NeuN p = 0.0001, t = 14.15, n = 3, df = 4. (C) Expression of Bdnf variants and downstream gene Lin7c during differentiation, assessed by qRT-PCR and normalized to NPC levels. Bars represent mean ± SEM; points show results from different biological replicates (n = 7, df = 12). ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001, ∗∗∗∗p< 0.0001, unpaired t-test (two-tailed). Exon I p = 0.0001, t = 5.627; Exon II p = 0.0007, t = 4.549; Exon III p = 0.0001, t = 5.614; Exon IV p = 0.0019, t = 3.972; Exon V p = 0.0029, t = 3.734; Exon VI p = 0.0021, t = 3.899; Exon VIII p = 0.0397, t = 2.307; Exon IXa p = 0.0023, t = 3.846; Lin7c p = 0.0248, t = 2.565. (D) Relocation of the Bdnf gene during neuronal development assessed by DNA-FISH combined with measurements of the distance of the signal from the closest edge of the nucleus. Left panel; representative confocal sections of DNA-FISH showing nuclear localization of Bdnf loci (green) in NPCs and PMNs. Nuclei were stained with DAPI (gray). For each image, the distance between the center of the FISH signal and the edge of the nucleus is indicated. Scale bars, 5 μm. Right panel; scatter dot plot of the distribution of the distance between Bdnf locus and the edge of the DAPI staining. Solid gray lines denote medians. ∗∗∗∗p = 0.0002, Mann-Whitney test (two-tailed). n = 133 (NPC), 123 (PMN) foci across 4 biological replicates. See also <xref ref-type=Figure S1 . " title="... day 12.5. FGF2, fibroblast growth factor 2. NT-3, neurotrophin-3. FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . (B) ..." property="contentUrl" width="100%" height="100%"/>
    Figure Legend Snippet: Expression of Bdnf isoforms increases over embryonic cortical neuron differentiation concomitant with movement of the gene locus away from the nuclear periphery (A) Schematic of neuronal precursor cell (NPC) differentiation into post-mitotic neurons (PMN). E12.5, embryonic day 12.5. FGF2, fibroblast growth factor 2. NT-3, neurotrophin-3. FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . (B) Expression profile of an NPC-marker, Nestin, and neuronal markers, Map2 and NeuN, in NPCs and PMNs, assessed by qRT-PCR and normalized to NPC. Bars represent mean ± SEM; points show results from different biological replicates . ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001; unpaired t-test (two-tailed). Nestin p = 0.0005, t = 4.707, n = 7, df = 12; Map2 p = 0.0481, t = 2.201, n = 7, df = 12; NeuN p = 0.0001, t = 14.15, n = 3, df = 4. (C) Expression of Bdnf variants and downstream gene Lin7c during differentiation, assessed by qRT-PCR and normalized to NPC levels. Bars represent mean ± SEM; points show results from different biological replicates (n = 7, df = 12). ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001, ∗∗∗∗p< 0.0001, unpaired t-test (two-tailed). Exon I p = 0.0001, t = 5.627; Exon II p = 0.0007, t = 4.549; Exon III p = 0.0001, t = 5.614; Exon IV p = 0.0019, t = 3.972; Exon V p = 0.0029, t = 3.734; Exon VI p = 0.0021, t = 3.899; Exon VIII p = 0.0397, t = 2.307; Exon IXa p = 0.0023, t = 3.846; Lin7c p = 0.0248, t = 2.565. (D) Relocation of the Bdnf gene during neuronal development assessed by DNA-FISH combined with measurements of the distance of the signal from the closest edge of the nucleus. Left panel; representative confocal sections of DNA-FISH showing nuclear localization of Bdnf loci (green) in NPCs and PMNs. Nuclei were stained with DAPI (gray). For each image, the distance between the center of the FISH signal and the edge of the nucleus is indicated. Scale bars, 5 μm. Right panel; scatter dot plot of the distribution of the distance between Bdnf locus and the edge of the DAPI staining. Solid gray lines denote medians. ∗∗∗∗p = 0.0002, Mann-Whitney test (two-tailed). n = 133 (NPC), 123 (PMN) foci across 4 biological replicates. See also Figure S1 .

    Techniques Used: Expressing, In Vitro, Marker, Quantitative RT-PCR, Two Tailed Test, Staining, MANN-WHITNEY


    Figure Legend Snippet:

    Techniques Used: Plasmid Preparation, Recombinant, Nick Translation, Sensitive Assay, Isolation, CRISPR, Software

    nt 3  (Alomone Labs)


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    Alomone Labs nt 3
    Expression of Bdnf isoforms increases over embryonic cortical neuron differentiation concomitant with movement of the gene locus away from the nuclear periphery (A) Schematic of neuronal precursor cell (NPC) differentiation into post-mitotic neurons (PMN). E12.5, embryonic day 12.5. FGF2, fibroblast growth factor 2. <t>NT-3,</t> <t>neurotrophin-3.</t> FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . (B) Expression profile of an NPC-marker, Nestin, and neuronal markers, Map2 and NeuN, in NPCs and PMNs, assessed by qRT-PCR and normalized to NPC. Bars represent mean ± SEM; points show results from different biological replicates . ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001; unpaired t-test (two-tailed). Nestin p = 0.0005, t = 4.707, n = 7, df = 12; Map2 p = 0.0481, t = 2.201, n = 7, df = 12; NeuN p = 0.0001, t = 14.15, n = 3, df = 4. (C) Expression of Bdnf variants and downstream gene Lin7c during differentiation, assessed by qRT-PCR and normalized to NPC levels. Bars represent mean ± SEM; points show results from different biological replicates (n = 7, df = 12). ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001, ∗∗∗∗p< 0.0001, unpaired t-test (two-tailed). Exon I p = 0.0001, t = 5.627; Exon II p = 0.0007, t = 4.549; Exon III p = 0.0001, t = 5.614; Exon IV p = 0.0019, t = 3.972; Exon V p = 0.0029, t = 3.734; Exon VI p = 0.0021, t = 3.899; Exon VIII p = 0.0397, t = 2.307; Exon IXa p = 0.0023, t = 3.846; Lin7c p = 0.0248, t = 2.565. (D) Relocation of the Bdnf gene during neuronal development assessed by DNA-FISH combined with measurements of the distance of the signal from the closest edge of the nucleus. Left panel; representative confocal sections of DNA-FISH showing nuclear localization of Bdnf loci (green) in NPCs and PMNs. Nuclei were stained with DAPI (gray). For each image, the distance between the center of the FISH signal and the edge of the nucleus is indicated. Scale bars, 5 μm. Right panel; scatter dot plot of the distribution of the distance between Bdnf locus and the edge of the DAPI staining. Solid gray lines denote medians. ∗∗∗∗p = 0.0002, Mann-Whitney test (two-tailed). n = 133 (NPC), 123 (PMN) foci across 4 biological replicates. See also <xref ref-type=Figure S1 . " width="250" height="auto" />
    Nt 3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/nt 3/product/Alomone Labs
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    nt 3 - by Bioz Stars, 2023-01
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    Images

    1) Product Images from "A novel intergenic enhancer that regulates Bdnf expression in developing cortical neurons"

    Article Title: A novel intergenic enhancer that regulates Bdnf expression in developing cortical neurons

    Journal: iScience

    doi: 10.1016/j.isci.2022.105695

    Expression of Bdnf isoforms increases over embryonic cortical neuron differentiation concomitant with movement of the gene locus away from the nuclear periphery (A) Schematic of neuronal precursor cell (NPC) differentiation into post-mitotic neurons (PMN). E12.5, embryonic day 12.5. FGF2, fibroblast growth factor 2. NT-3, neurotrophin-3. FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . (B) Expression profile of an NPC-marker, Nestin, and neuronal markers, Map2 and NeuN, in NPCs and PMNs, assessed by qRT-PCR and normalized to NPC. Bars represent mean ± SEM; points show results from different biological replicates . ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001; unpaired t-test (two-tailed). Nestin p = 0.0005, t = 4.707, n = 7, df = 12; Map2 p = 0.0481, t = 2.201, n = 7, df = 12; NeuN p = 0.0001, t = 14.15, n = 3, df = 4. (C) Expression of Bdnf variants and downstream gene Lin7c during differentiation, assessed by qRT-PCR and normalized to NPC levels. Bars represent mean ± SEM; points show results from different biological replicates (n = 7, df = 12). ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001, ∗∗∗∗p< 0.0001, unpaired t-test (two-tailed). Exon I p = 0.0001, t = 5.627; Exon II p = 0.0007, t = 4.549; Exon III p = 0.0001, t = 5.614; Exon IV p = 0.0019, t = 3.972; Exon V p = 0.0029, t = 3.734; Exon VI p = 0.0021, t = 3.899; Exon VIII p = 0.0397, t = 2.307; Exon IXa p = 0.0023, t = 3.846; Lin7c p = 0.0248, t = 2.565. (D) Relocation of the Bdnf gene during neuronal development assessed by DNA-FISH combined with measurements of the distance of the signal from the closest edge of the nucleus. Left panel; representative confocal sections of DNA-FISH showing nuclear localization of Bdnf loci (green) in NPCs and PMNs. Nuclei were stained with DAPI (gray). For each image, the distance between the center of the FISH signal and the edge of the nucleus is indicated. Scale bars, 5 μm. Right panel; scatter dot plot of the distribution of the distance between Bdnf locus and the edge of the DAPI staining. Solid gray lines denote medians. ∗∗∗∗p = 0.0002, Mann-Whitney test (two-tailed). n = 133 (NPC), 123 (PMN) foci across 4 biological replicates. See also <xref ref-type=Figure S1 . " title="... embryonic day 12.5. FGF2, fibroblast growth factor 2. NT-3, neurotrophin-3. FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . ..." property="contentUrl" width="100%" height="100%"/>
    Figure Legend Snippet: Expression of Bdnf isoforms increases over embryonic cortical neuron differentiation concomitant with movement of the gene locus away from the nuclear periphery (A) Schematic of neuronal precursor cell (NPC) differentiation into post-mitotic neurons (PMN). E12.5, embryonic day 12.5. FGF2, fibroblast growth factor 2. NT-3, neurotrophin-3. FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . (B) Expression profile of an NPC-marker, Nestin, and neuronal markers, Map2 and NeuN, in NPCs and PMNs, assessed by qRT-PCR and normalized to NPC. Bars represent mean ± SEM; points show results from different biological replicates . ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001; unpaired t-test (two-tailed). Nestin p = 0.0005, t = 4.707, n = 7, df = 12; Map2 p = 0.0481, t = 2.201, n = 7, df = 12; NeuN p = 0.0001, t = 14.15, n = 3, df = 4. (C) Expression of Bdnf variants and downstream gene Lin7c during differentiation, assessed by qRT-PCR and normalized to NPC levels. Bars represent mean ± SEM; points show results from different biological replicates (n = 7, df = 12). ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001, ∗∗∗∗p< 0.0001, unpaired t-test (two-tailed). Exon I p = 0.0001, t = 5.627; Exon II p = 0.0007, t = 4.549; Exon III p = 0.0001, t = 5.614; Exon IV p = 0.0019, t = 3.972; Exon V p = 0.0029, t = 3.734; Exon VI p = 0.0021, t = 3.899; Exon VIII p = 0.0397, t = 2.307; Exon IXa p = 0.0023, t = 3.846; Lin7c p = 0.0248, t = 2.565. (D) Relocation of the Bdnf gene during neuronal development assessed by DNA-FISH combined with measurements of the distance of the signal from the closest edge of the nucleus. Left panel; representative confocal sections of DNA-FISH showing nuclear localization of Bdnf loci (green) in NPCs and PMNs. Nuclei were stained with DAPI (gray). For each image, the distance between the center of the FISH signal and the edge of the nucleus is indicated. Scale bars, 5 μm. Right panel; scatter dot plot of the distribution of the distance between Bdnf locus and the edge of the DAPI staining. Solid gray lines denote medians. ∗∗∗∗p = 0.0002, Mann-Whitney test (two-tailed). n = 133 (NPC), 123 (PMN) foci across 4 biological replicates. See also Figure S1 .

    Techniques Used: Expressing, In Vitro, Marker, Quantitative RT-PCR, Two Tailed Test, Staining, MANN-WHITNEY

    nt 3  (Alomone Labs)


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    Alomone Labs nt 3
    Expression of Bdnf isoforms increases over embryonic cortical neuron differentiation concomitant with movement of the gene locus away from the nuclear periphery (A) Schematic of neuronal precursor cell (NPC) differentiation into post-mitotic neurons (PMN). E12.5, embryonic day 12.5. FGF2, fibroblast growth factor 2. <t>NT-3,</t> <t>neurotrophin-3.</t> FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . (B) Expression profile of an NPC-marker, Nestin, and neuronal markers, Map2 and NeuN, in NPCs and PMNs, assessed by qRT-PCR and normalized to NPC. Bars represent mean ± SEM; points show results from different biological replicates . ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001; unpaired t-test (two-tailed). Nestin p = 0.0005, t = 4.707, n = 7, df = 12; Map2 p = 0.0481, t = 2.201, n = 7, df = 12; NeuN p = 0.0001, t = 14.15, n = 3, df = 4. (C) Expression of Bdnf variants and downstream gene Lin7c during differentiation, assessed by qRT-PCR and normalized to NPC levels. Bars represent mean ± SEM; points show results from different biological replicates (n = 7, df = 12). ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001, ∗∗∗∗p< 0.0001, unpaired t-test (two-tailed). Exon I p = 0.0001, t = 5.627; Exon II p = 0.0007, t = 4.549; Exon III p = 0.0001, t = 5.614; Exon IV p = 0.0019, t = 3.972; Exon V p = 0.0029, t = 3.734; Exon VI p = 0.0021, t = 3.899; Exon VIII p = 0.0397, t = 2.307; Exon IXa p = 0.0023, t = 3.846; Lin7c p = 0.0248, t = 2.565. (D) Relocation of the Bdnf gene during neuronal development assessed by DNA-FISH combined with measurements of the distance of the signal from the closest edge of the nucleus. Left panel; representative confocal sections of DNA-FISH showing nuclear localization of Bdnf loci (green) in NPCs and PMNs. Nuclei were stained with DAPI (gray). For each image, the distance between the center of the FISH signal and the edge of the nucleus is indicated. Scale bars, 5 μm. Right panel; scatter dot plot of the distribution of the distance between Bdnf locus and the edge of the DAPI staining. Solid gray lines denote medians. ∗∗∗∗p = 0.0002, Mann-Whitney test (two-tailed). n = 133 (NPC), 123 (PMN) foci across 4 biological replicates. See also <xref ref-type=Figure S1 . " width="250" height="auto" />
    Nt 3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "A novel intergenic enhancer that regulates Bdnf expression in developing cortical neurons"

    Article Title: A novel intergenic enhancer that regulates Bdnf expression in developing cortical neurons

    Journal: iScience

    doi: 10.1016/j.isci.2022.105695

    Expression of Bdnf isoforms increases over embryonic cortical neuron differentiation concomitant with movement of the gene locus away from the nuclear periphery (A) Schematic of neuronal precursor cell (NPC) differentiation into post-mitotic neurons (PMN). E12.5, embryonic day 12.5. FGF2, fibroblast growth factor 2. NT-3, neurotrophin-3. FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . (B) Expression profile of an NPC-marker, Nestin, and neuronal markers, Map2 and NeuN, in NPCs and PMNs, assessed by qRT-PCR and normalized to NPC. Bars represent mean ± SEM; points show results from different biological replicates . ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001; unpaired t-test (two-tailed). Nestin p = 0.0005, t = 4.707, n = 7, df = 12; Map2 p = 0.0481, t = 2.201, n = 7, df = 12; NeuN p = 0.0001, t = 14.15, n = 3, df = 4. (C) Expression of Bdnf variants and downstream gene Lin7c during differentiation, assessed by qRT-PCR and normalized to NPC levels. Bars represent mean ± SEM; points show results from different biological replicates (n = 7, df = 12). ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001, ∗∗∗∗p< 0.0001, unpaired t-test (two-tailed). Exon I p = 0.0001, t = 5.627; Exon II p = 0.0007, t = 4.549; Exon III p = 0.0001, t = 5.614; Exon IV p = 0.0019, t = 3.972; Exon V p = 0.0029, t = 3.734; Exon VI p = 0.0021, t = 3.899; Exon VIII p = 0.0397, t = 2.307; Exon IXa p = 0.0023, t = 3.846; Lin7c p = 0.0248, t = 2.565. (D) Relocation of the Bdnf gene during neuronal development assessed by DNA-FISH combined with measurements of the distance of the signal from the closest edge of the nucleus. Left panel; representative confocal sections of DNA-FISH showing nuclear localization of Bdnf loci (green) in NPCs and PMNs. Nuclei were stained with DAPI (gray). For each image, the distance between the center of the FISH signal and the edge of the nucleus is indicated. Scale bars, 5 μm. Right panel; scatter dot plot of the distribution of the distance between Bdnf locus and the edge of the DAPI staining. Solid gray lines denote medians. ∗∗∗∗p = 0.0002, Mann-Whitney test (two-tailed). n = 133 (NPC), 123 (PMN) foci across 4 biological replicates. See also <xref ref-type=Figure S1 . " title="... embryonic day 12.5. FGF2, fibroblast growth factor 2. NT-3, neurotrophin-3. FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . ..." property="contentUrl" width="100%" height="100%"/>
    Figure Legend Snippet: Expression of Bdnf isoforms increases over embryonic cortical neuron differentiation concomitant with movement of the gene locus away from the nuclear periphery (A) Schematic of neuronal precursor cell (NPC) differentiation into post-mitotic neurons (PMN). E12.5, embryonic day 12.5. FGF2, fibroblast growth factor 2. NT-3, neurotrophin-3. FdU, 5-fluoro-2′-deoxyuridine. DIV, days in vitro . (B) Expression profile of an NPC-marker, Nestin, and neuronal markers, Map2 and NeuN, in NPCs and PMNs, assessed by qRT-PCR and normalized to NPC. Bars represent mean ± SEM; points show results from different biological replicates . ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001; unpaired t-test (two-tailed). Nestin p = 0.0005, t = 4.707, n = 7, df = 12; Map2 p = 0.0481, t = 2.201, n = 7, df = 12; NeuN p = 0.0001, t = 14.15, n = 3, df = 4. (C) Expression of Bdnf variants and downstream gene Lin7c during differentiation, assessed by qRT-PCR and normalized to NPC levels. Bars represent mean ± SEM; points show results from different biological replicates (n = 7, df = 12). ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001, ∗∗∗∗p< 0.0001, unpaired t-test (two-tailed). Exon I p = 0.0001, t = 5.627; Exon II p = 0.0007, t = 4.549; Exon III p = 0.0001, t = 5.614; Exon IV p = 0.0019, t = 3.972; Exon V p = 0.0029, t = 3.734; Exon VI p = 0.0021, t = 3.899; Exon VIII p = 0.0397, t = 2.307; Exon IXa p = 0.0023, t = 3.846; Lin7c p = 0.0248, t = 2.565. (D) Relocation of the Bdnf gene during neuronal development assessed by DNA-FISH combined with measurements of the distance of the signal from the closest edge of the nucleus. Left panel; representative confocal sections of DNA-FISH showing nuclear localization of Bdnf loci (green) in NPCs and PMNs. Nuclei were stained with DAPI (gray). For each image, the distance between the center of the FISH signal and the edge of the nucleus is indicated. Scale bars, 5 μm. Right panel; scatter dot plot of the distribution of the distance between Bdnf locus and the edge of the DAPI staining. Solid gray lines denote medians. ∗∗∗∗p = 0.0002, Mann-Whitney test (two-tailed). n = 133 (NPC), 123 (PMN) foci across 4 biological replicates. See also Figure S1 .

    Techniques Used: Expressing, In Vitro, Marker, Quantitative RT-PCR, Two Tailed Test, Staining, MANN-WHITNEY

    pro nt3  (Alomone Labs)


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    Alomone Labs pro nt3
    Differential expression of mature <t>NT3,</t> pro-NT3, rod and cone phototransduction proteins 7d after Müller cell ablation. (A, F) Western blots using antibodies to detect mature NT3, pro-NT3, rhodopsin, guanine nucleotide-binding protein subunit alpha-1 (GNAT1) and Gα protein transducin (Gαt). GNAT1 and Gαt are proteins essential for rod and cone phototransduction [ - ]. (B-D) Quantitative analysis of protein densitometry showed significant reduction in mature NT3 (B) and upregulation of pro-NT3 (C) , which resulted in a decreased ratio of NT3:pro-NT3 (D) 7d after Müller cell ablation. (E, G, H) The reduced expression of rhodopsin (E) , GNAT1 (G) and Gαt (H) in transgenic (TG) mice indicates that selective Müller cell ablation causes damage to both rod and cone photoreceptors. * P < 0.05 and † P < 0.01, TG vs. control ( Ctl ), n = 8/group in (B, D and E) and n = 11 − 13/group in (C, G and H) , respectively.
    Pro Nt3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Involvement of NT3 and P75 NTR in photoreceptor degeneration following selective Müller cell ablation"

    Article Title: Involvement of NT3 and P75 NTR in photoreceptor degeneration following selective Müller cell ablation

    Journal: Journal of Neuroinflammation

    doi: 10.1186/1742-2094-10-137

    Differential expression of mature NT3, pro-NT3, rod and cone phototransduction proteins 7d after Müller cell ablation. (A, F) Western blots using antibodies to detect mature NT3, pro-NT3, rhodopsin, guanine nucleotide-binding protein subunit alpha-1 (GNAT1) and Gα protein transducin (Gαt). GNAT1 and Gαt are proteins essential for rod and cone phototransduction [ - ]. (B-D) Quantitative analysis of protein densitometry showed significant reduction in mature NT3 (B) and upregulation of pro-NT3 (C) , which resulted in a decreased ratio of NT3:pro-NT3 (D) 7d after Müller cell ablation. (E, G, H) The reduced expression of rhodopsin (E) , GNAT1 (G) and Gαt (H) in transgenic (TG) mice indicates that selective Müller cell ablation causes damage to both rod and cone photoreceptors. * P < 0.05 and † P < 0.01, TG vs. control ( Ctl ), n = 8/group in (B, D and E) and n = 11 − 13/group in (C, G and H) , respectively.
    Figure Legend Snippet: Differential expression of mature NT3, pro-NT3, rod and cone phototransduction proteins 7d after Müller cell ablation. (A, F) Western blots using antibodies to detect mature NT3, pro-NT3, rhodopsin, guanine nucleotide-binding protein subunit alpha-1 (GNAT1) and Gα protein transducin (Gαt). GNAT1 and Gαt are proteins essential for rod and cone phototransduction [ - ]. (B-D) Quantitative analysis of protein densitometry showed significant reduction in mature NT3 (B) and upregulation of pro-NT3 (C) , which resulted in a decreased ratio of NT3:pro-NT3 (D) 7d after Müller cell ablation. (E, G, H) The reduced expression of rhodopsin (E) , GNAT1 (G) and Gαt (H) in transgenic (TG) mice indicates that selective Müller cell ablation causes damage to both rod and cone photoreceptors. * P < 0.05 and † P < 0.01, TG vs. control ( Ctl ), n = 8/group in (B, D and E) and n = 11 − 13/group in (C, G and H) , respectively.

    Techniques Used: Expressing, Western Blot, Binding Assay, Transgenic Assay

    Intravitreal injection of recombinant NT3 and blockage of P75 NTR protected photoreceptor degeneration. Intravitreal injection was performed in transgenic mice 3d and 6d after tamoxifen induced Müller cell ablation, with one eye receiving the testing reagent and the contralateral eye receiving balanced salt solution (BSS). Doses of injection: (1) NT3, 0.4 μg; (2) P75 NTR rabbit polyclonal Ab (1:1 dilution) and (3) 0.4 μg NT3 + 1:1 dilution of P75 NTR Ab. Eyes were enucleated 10d after TMX treatment for flat-mount staining using fluorescence-conjugated peanut-agglutinin (PNA). (A and B) PNA-labeled cone photoreceptor outer segments ( green ) and nuclear counterstaining in a control ( Ctl ) retina. (C-F) Images from transgenic ( TG ) mice showing changes in cone photoreceptor outer segments and protrusion of photoreceptor nuclei into the subretinal space ( red , indicated by asterisks ) in eyes receiving BSS (C) , NT3 (D) , P75 NTR Ab (E) and a combination of NT3 and P75 NTR Ab (F) . (G) Quantitative analysis of PNA staining showed that intravitreal injection of NT3 and P75 Ab protected the loss of cone photoreceptor outer segments, with a combined treatment more effective than either alone. N = 9 − 11/group. Scale bars in A-F : 100 μm.
    Figure Legend Snippet: Intravitreal injection of recombinant NT3 and blockage of P75 NTR protected photoreceptor degeneration. Intravitreal injection was performed in transgenic mice 3d and 6d after tamoxifen induced Müller cell ablation, with one eye receiving the testing reagent and the contralateral eye receiving balanced salt solution (BSS). Doses of injection: (1) NT3, 0.4 μg; (2) P75 NTR rabbit polyclonal Ab (1:1 dilution) and (3) 0.4 μg NT3 + 1:1 dilution of P75 NTR Ab. Eyes were enucleated 10d after TMX treatment for flat-mount staining using fluorescence-conjugated peanut-agglutinin (PNA). (A and B) PNA-labeled cone photoreceptor outer segments ( green ) and nuclear counterstaining in a control ( Ctl ) retina. (C-F) Images from transgenic ( TG ) mice showing changes in cone photoreceptor outer segments and protrusion of photoreceptor nuclei into the subretinal space ( red , indicated by asterisks ) in eyes receiving BSS (C) , NT3 (D) , P75 NTR Ab (E) and a combination of NT3 and P75 NTR Ab (F) . (G) Quantitative analysis of PNA staining showed that intravitreal injection of NT3 and P75 Ab protected the loss of cone photoreceptor outer segments, with a combined treatment more effective than either alone. N = 9 − 11/group. Scale bars in A-F : 100 μm.

    Techniques Used: Injection, Recombinant, Transgenic Assay, Staining, Fluorescence, Labeling

    Inhibition of microglial activation after intravitreal injection of recombinant mature NT3 and blockage of P75 NTR . Transgenic mice received twice intravitreal injections of NT3 (0.4 μg), P75 NTR rabbit polyclonal a polyclonal Ab to P75 NTR (1:1 dilution), a combination of both or balanced salt solution ( BSS ) 3d and 6d after tamoxifen (TMX)-induced Muller cell ablation. Eyes were enucleated 10d after TMX treatment for retinal flat-mount staining using an Ab against ionized calcium binding adaptor molecule 1 ( IBA-1 ) for microglia ( A, D, G and J , green ) and peanut-agglutinin ( PNA )-conjugated with Alexa Fluor-594 for cone photoreceptor outer segments ( B, E, H and K , red ). (C, F, I and L) Merged images. (M) Quantitative analysis of IBA-1 stained retinal whole mounts shows intravitreal injection of NT3 and P75 NTR blockage, either alone or in combination, protected photoreceptors concurrently with inhibition of microglial activation. * P < 0.05 and † P < 0.01, all vs. BSS injected group, n = 9 − 11/group. Scale bars in A-L : 100 μm.
    Figure Legend Snippet: Inhibition of microglial activation after intravitreal injection of recombinant mature NT3 and blockage of P75 NTR . Transgenic mice received twice intravitreal injections of NT3 (0.4 μg), P75 NTR rabbit polyclonal a polyclonal Ab to P75 NTR (1:1 dilution), a combination of both or balanced salt solution ( BSS ) 3d and 6d after tamoxifen (TMX)-induced Muller cell ablation. Eyes were enucleated 10d after TMX treatment for retinal flat-mount staining using an Ab against ionized calcium binding adaptor molecule 1 ( IBA-1 ) for microglia ( A, D, G and J , green ) and peanut-agglutinin ( PNA )-conjugated with Alexa Fluor-594 for cone photoreceptor outer segments ( B, E, H and K , red ). (C, F, I and L) Merged images. (M) Quantitative analysis of IBA-1 stained retinal whole mounts shows intravitreal injection of NT3 and P75 NTR blockage, either alone or in combination, protected photoreceptors concurrently with inhibition of microglial activation. * P < 0.05 and † P < 0.01, all vs. BSS injected group, n = 9 − 11/group. Scale bars in A-L : 100 μm.

    Techniques Used: Inhibition, Activation Assay, Injection, Recombinant, Transgenic Assay, Staining, Binding Assay

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    Alomone Labs pro nt3
    Differential expression of mature <t>NT3,</t> pro-NT3, rod and cone phototransduction proteins 7d after Müller cell ablation. (A, F) Western blots using antibodies to detect mature NT3, pro-NT3, rhodopsin, guanine nucleotide-binding protein subunit alpha-1 (GNAT1) and Gα protein transducin (Gαt). GNAT1 and Gαt are proteins essential for rod and cone phototransduction [ - ]. (B-D) Quantitative analysis of protein densitometry showed significant reduction in mature NT3 (B) and upregulation of pro-NT3 (C) , which resulted in a decreased ratio of NT3:pro-NT3 (D) 7d after Müller cell ablation. (E, G, H) The reduced expression of rhodopsin (E) , GNAT1 (G) and Gαt (H) in transgenic (TG) mice indicates that selective Müller cell ablation causes damage to both rod and cone photoreceptors. * P < 0.05 and † P < 0.01, TG vs. control ( Ctl ), n = 8/group in (B, D and E) and n = 11 − 13/group in (C, G and H) , respectively.
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    (A, D, G) Representative STED (A), TREx (D) and dual-color SMLM (G) images of the ER and ribosomes in axons from neurons expressing GFP-Sec61β and stained for RpS12. (B, E, H) Magnifications and intensity profile lines from merged images for each microscopy method. (C, F, I) Quantification of RpS12 intensity in ER mask, enlarged ER mask, and one-color flipped images, for each microscopy method. (J) Representative STED images and intensity profile line for an axon segment of a neuron transfected as in (A) and co-labeled for puromycilated peptides. (K) Quantification of RpS12 intensity in ER mask and enlarged ER mask with or without high puromycin treatment using dual-color SMLM. (L-N) Schematic representation of split APEX system used to detect ER-ribosome contacts. When the ER protein Sec61β fused to AP module and the ribosomal protein RpL10A fused to EX module interact with each other, APEX is reconstituted and contact sites can be visualized as a biotinylation radius around the interactions (L). Representative images of split APEX assay in distal axons from neurons expressing RpL10A-3xHA-EX and V5-AP-Sec61β (left), or V5-AP-RTN4A as a negative control (right). Expression of constructs are visualized with V5 and HA antibodies, and biotinylation is detected with conjugated Strep-555 (M). Quantification of Strep signal in distal axons from neurons as in (M), and without H2O2 as a negative control for the biotinylation reaction (N). (O) Quantification of axonal ER-bound ribosomes using split APEX assay with RpL10A-3xHA-EX and V5-AP-Sec61β, in neurons stimulated for 30-minutes with BSA (control), BDNF, NT-3 or NGF. Individual data points each represent a neuron in (C, F, I, K, N and O). Boxplots show 25/75-percentiles, the median, and whiskers represent min to max in (C, F, I and K). Data are presented as mean values ± SEM in (N, O). ns = not significant, *p < 0.05, **p < 0.01, ***p<0.001 comparing conditions to control using unpaired t-tests or ordinary one-way ANOVA tests. Scale bars represent 1μm (A, D, G, J) and 5μm (M).

    Journal: bioRxiv

    Article Title: Axonal ER tubules regulate local translation via P180/RRBP1-mediated ribosome interactions

    doi: 10.1101/2022.11.30.518484

    Figure Lengend Snippet: (A, D, G) Representative STED (A), TREx (D) and dual-color SMLM (G) images of the ER and ribosomes in axons from neurons expressing GFP-Sec61β and stained for RpS12. (B, E, H) Magnifications and intensity profile lines from merged images for each microscopy method. (C, F, I) Quantification of RpS12 intensity in ER mask, enlarged ER mask, and one-color flipped images, for each microscopy method. (J) Representative STED images and intensity profile line for an axon segment of a neuron transfected as in (A) and co-labeled for puromycilated peptides. (K) Quantification of RpS12 intensity in ER mask and enlarged ER mask with or without high puromycin treatment using dual-color SMLM. (L-N) Schematic representation of split APEX system used to detect ER-ribosome contacts. When the ER protein Sec61β fused to AP module and the ribosomal protein RpL10A fused to EX module interact with each other, APEX is reconstituted and contact sites can be visualized as a biotinylation radius around the interactions (L). Representative images of split APEX assay in distal axons from neurons expressing RpL10A-3xHA-EX and V5-AP-Sec61β (left), or V5-AP-RTN4A as a negative control (right). Expression of constructs are visualized with V5 and HA antibodies, and biotinylation is detected with conjugated Strep-555 (M). Quantification of Strep signal in distal axons from neurons as in (M), and without H2O2 as a negative control for the biotinylation reaction (N). (O) Quantification of axonal ER-bound ribosomes using split APEX assay with RpL10A-3xHA-EX and V5-AP-Sec61β, in neurons stimulated for 30-minutes with BSA (control), BDNF, NT-3 or NGF. Individual data points each represent a neuron in (C, F, I, K, N and O). Boxplots show 25/75-percentiles, the median, and whiskers represent min to max in (C, F, I and K). Data are presented as mean values ± SEM in (N, O). ns = not significant, *p < 0.05, **p < 0.01, ***p<0.001 comparing conditions to control using unpaired t-tests or ordinary one-way ANOVA tests. Scale bars represent 1μm (A, D, G, J) and 5μm (M).

    Article Snippet: Other reagents used in this study were: Puromycin dihydrochloride (Sigma-Aldrich, Cat# P8833), Anisomycin (Sigma-Aldrich, Cat#9789), recombinant human Neurotrophin-3 (NT-3) protein (50ng/ml, Alomone labs, Cat#N-260), recombinant human BDNF protein (50ng/ml, Alomone labs, Cat#B-250), recombinant rat beta-NGF Protein (50ng/ml, R&D systems, Cat# 556-NG).

    Techniques: Expressing, Staining, Microscopy, Transfection, Labeling, Negative Control, Construct

    Differential expression of mature NT3, pro-NT3, rod and cone phototransduction proteins 7d after Müller cell ablation. (A, F) Western blots using antibodies to detect mature NT3, pro-NT3, rhodopsin, guanine nucleotide-binding protein subunit alpha-1 (GNAT1) and Gα protein transducin (Gαt). GNAT1 and Gαt are proteins essential for rod and cone phototransduction [ - ]. (B-D) Quantitative analysis of protein densitometry showed significant reduction in mature NT3 (B) and upregulation of pro-NT3 (C) , which resulted in a decreased ratio of NT3:pro-NT3 (D) 7d after Müller cell ablation. (E, G, H) The reduced expression of rhodopsin (E) , GNAT1 (G) and Gαt (H) in transgenic (TG) mice indicates that selective Müller cell ablation causes damage to both rod and cone photoreceptors. * P < 0.05 and † P < 0.01, TG vs. control ( Ctl ), n = 8/group in (B, D and E) and n = 11 − 13/group in (C, G and H) , respectively.

    Journal: Journal of Neuroinflammation

    Article Title: Involvement of NT3 and P75 NTR in photoreceptor degeneration following selective Müller cell ablation

    doi: 10.1186/1742-2094-10-137

    Figure Lengend Snippet: Differential expression of mature NT3, pro-NT3, rod and cone phototransduction proteins 7d after Müller cell ablation. (A, F) Western blots using antibodies to detect mature NT3, pro-NT3, rhodopsin, guanine nucleotide-binding protein subunit alpha-1 (GNAT1) and Gα protein transducin (Gαt). GNAT1 and Gαt are proteins essential for rod and cone phototransduction [ - ]. (B-D) Quantitative analysis of protein densitometry showed significant reduction in mature NT3 (B) and upregulation of pro-NT3 (C) , which resulted in a decreased ratio of NT3:pro-NT3 (D) 7d after Müller cell ablation. (E, G, H) The reduced expression of rhodopsin (E) , GNAT1 (G) and Gαt (H) in transgenic (TG) mice indicates that selective Müller cell ablation causes damage to both rod and cone photoreceptors. * P < 0.05 and † P < 0.01, TG vs. control ( Ctl ), n = 8/group in (B, D and E) and n = 11 − 13/group in (C, G and H) , respectively.

    Article Snippet: Membranes were probed with antibodies to GS (mouse monoclonal, 1:1,000; Millipore, no. MAB302), GFAP (mouse monoclonal, 1:5,000; Neomarker, no. MS-280-P), P75 NTR (rabbit polyclonal, a gift from Dr. Moses V. Chao, no. 9651), NT3 (rabbit polyclonal, 1:1,000, Alomone Laboratory, no. ANT-003), pro-NT3 (rabbit polyclonal, 1:500, Alomone Laboratory, no. ANT-012) and rhodopsin (mouse monoclonal, 1:500, Millipore no. MAB5356), guanine nucleotide-binding protein subunit alpha-1 (GNAT1, rabbit polyclonal, 1:500, Santa Cruz no. sc-389) and Gα protein transducin (Gαt, mouse monoclonal, 1:2,000, BD Transduction Laboratories no. 610589).

    Techniques: Expressing, Western Blot, Binding Assay, Transgenic Assay

    Intravitreal injection of recombinant NT3 and blockage of P75 NTR protected photoreceptor degeneration. Intravitreal injection was performed in transgenic mice 3d and 6d after tamoxifen induced Müller cell ablation, with one eye receiving the testing reagent and the contralateral eye receiving balanced salt solution (BSS). Doses of injection: (1) NT3, 0.4 μg; (2) P75 NTR rabbit polyclonal Ab (1:1 dilution) and (3) 0.4 μg NT3 + 1:1 dilution of P75 NTR Ab. Eyes were enucleated 10d after TMX treatment for flat-mount staining using fluorescence-conjugated peanut-agglutinin (PNA). (A and B) PNA-labeled cone photoreceptor outer segments ( green ) and nuclear counterstaining in a control ( Ctl ) retina. (C-F) Images from transgenic ( TG ) mice showing changes in cone photoreceptor outer segments and protrusion of photoreceptor nuclei into the subretinal space ( red , indicated by asterisks ) in eyes receiving BSS (C) , NT3 (D) , P75 NTR Ab (E) and a combination of NT3 and P75 NTR Ab (F) . (G) Quantitative analysis of PNA staining showed that intravitreal injection of NT3 and P75 Ab protected the loss of cone photoreceptor outer segments, with a combined treatment more effective than either alone. N = 9 − 11/group. Scale bars in A-F : 100 μm.

    Journal: Journal of Neuroinflammation

    Article Title: Involvement of NT3 and P75 NTR in photoreceptor degeneration following selective Müller cell ablation

    doi: 10.1186/1742-2094-10-137

    Figure Lengend Snippet: Intravitreal injection of recombinant NT3 and blockage of P75 NTR protected photoreceptor degeneration. Intravitreal injection was performed in transgenic mice 3d and 6d after tamoxifen induced Müller cell ablation, with one eye receiving the testing reagent and the contralateral eye receiving balanced salt solution (BSS). Doses of injection: (1) NT3, 0.4 μg; (2) P75 NTR rabbit polyclonal Ab (1:1 dilution) and (3) 0.4 μg NT3 + 1:1 dilution of P75 NTR Ab. Eyes were enucleated 10d after TMX treatment for flat-mount staining using fluorescence-conjugated peanut-agglutinin (PNA). (A and B) PNA-labeled cone photoreceptor outer segments ( green ) and nuclear counterstaining in a control ( Ctl ) retina. (C-F) Images from transgenic ( TG ) mice showing changes in cone photoreceptor outer segments and protrusion of photoreceptor nuclei into the subretinal space ( red , indicated by asterisks ) in eyes receiving BSS (C) , NT3 (D) , P75 NTR Ab (E) and a combination of NT3 and P75 NTR Ab (F) . (G) Quantitative analysis of PNA staining showed that intravitreal injection of NT3 and P75 Ab protected the loss of cone photoreceptor outer segments, with a combined treatment more effective than either alone. N = 9 − 11/group. Scale bars in A-F : 100 μm.

    Article Snippet: Membranes were probed with antibodies to GS (mouse monoclonal, 1:1,000; Millipore, no. MAB302), GFAP (mouse monoclonal, 1:5,000; Neomarker, no. MS-280-P), P75 NTR (rabbit polyclonal, a gift from Dr. Moses V. Chao, no. 9651), NT3 (rabbit polyclonal, 1:1,000, Alomone Laboratory, no. ANT-003), pro-NT3 (rabbit polyclonal, 1:500, Alomone Laboratory, no. ANT-012) and rhodopsin (mouse monoclonal, 1:500, Millipore no. MAB5356), guanine nucleotide-binding protein subunit alpha-1 (GNAT1, rabbit polyclonal, 1:500, Santa Cruz no. sc-389) and Gα protein transducin (Gαt, mouse monoclonal, 1:2,000, BD Transduction Laboratories no. 610589).

    Techniques: Injection, Recombinant, Transgenic Assay, Staining, Fluorescence, Labeling

    Inhibition of microglial activation after intravitreal injection of recombinant mature NT3 and blockage of P75 NTR . Transgenic mice received twice intravitreal injections of NT3 (0.4 μg), P75 NTR rabbit polyclonal a polyclonal Ab to P75 NTR (1:1 dilution), a combination of both or balanced salt solution ( BSS ) 3d and 6d after tamoxifen (TMX)-induced Muller cell ablation. Eyes were enucleated 10d after TMX treatment for retinal flat-mount staining using an Ab against ionized calcium binding adaptor molecule 1 ( IBA-1 ) for microglia ( A, D, G and J , green ) and peanut-agglutinin ( PNA )-conjugated with Alexa Fluor-594 for cone photoreceptor outer segments ( B, E, H and K , red ). (C, F, I and L) Merged images. (M) Quantitative analysis of IBA-1 stained retinal whole mounts shows intravitreal injection of NT3 and P75 NTR blockage, either alone or in combination, protected photoreceptors concurrently with inhibition of microglial activation. * P < 0.05 and † P < 0.01, all vs. BSS injected group, n = 9 − 11/group. Scale bars in A-L : 100 μm.

    Journal: Journal of Neuroinflammation

    Article Title: Involvement of NT3 and P75 NTR in photoreceptor degeneration following selective Müller cell ablation

    doi: 10.1186/1742-2094-10-137

    Figure Lengend Snippet: Inhibition of microglial activation after intravitreal injection of recombinant mature NT3 and blockage of P75 NTR . Transgenic mice received twice intravitreal injections of NT3 (0.4 μg), P75 NTR rabbit polyclonal a polyclonal Ab to P75 NTR (1:1 dilution), a combination of both or balanced salt solution ( BSS ) 3d and 6d after tamoxifen (TMX)-induced Muller cell ablation. Eyes were enucleated 10d after TMX treatment for retinal flat-mount staining using an Ab against ionized calcium binding adaptor molecule 1 ( IBA-1 ) for microglia ( A, D, G and J , green ) and peanut-agglutinin ( PNA )-conjugated with Alexa Fluor-594 for cone photoreceptor outer segments ( B, E, H and K , red ). (C, F, I and L) Merged images. (M) Quantitative analysis of IBA-1 stained retinal whole mounts shows intravitreal injection of NT3 and P75 NTR blockage, either alone or in combination, protected photoreceptors concurrently with inhibition of microglial activation. * P < 0.05 and † P < 0.01, all vs. BSS injected group, n = 9 − 11/group. Scale bars in A-L : 100 μm.

    Article Snippet: Membranes were probed with antibodies to GS (mouse monoclonal, 1:1,000; Millipore, no. MAB302), GFAP (mouse monoclonal, 1:5,000; Neomarker, no. MS-280-P), P75 NTR (rabbit polyclonal, a gift from Dr. Moses V. Chao, no. 9651), NT3 (rabbit polyclonal, 1:1,000, Alomone Laboratory, no. ANT-003), pro-NT3 (rabbit polyclonal, 1:500, Alomone Laboratory, no. ANT-012) and rhodopsin (mouse monoclonal, 1:500, Millipore no. MAB5356), guanine nucleotide-binding protein subunit alpha-1 (GNAT1, rabbit polyclonal, 1:500, Santa Cruz no. sc-389) and Gα protein transducin (Gαt, mouse monoclonal, 1:2,000, BD Transduction Laboratories no. 610589).

    Techniques: Inhibition, Activation Assay, Injection, Recombinant, Transgenic Assay, Staining, Binding Assay