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Hes5-GFP expression in developing retina. (A) Hes5-GFP in the E12.5 mouse retina. (B) in situ hybridization for Hes5 mRNA at E13.5. (C,D) Hes5-GFP expressing cells in the E12.5 day retina co-express progenitor markers, PH3 (C) and <t>BrdU</t> (D). (E–G) Hes5-GFP expression at postnatal day 7 is confined to the inner nuclear layer in the maturing Müller glia (Id1+; G)), and is not expressed in the Pax6+ or <t>TuJ1+</t> inner retinal neurons (E and F, respectively).
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1) Product Images from "Genome-Wide Analysis of M?ller Glial Differentiation Reveals a Requirement for Notch Signaling in Postmitotic Cells to Maintain the Glial Fate"

Article Title: Genome-Wide Analysis of M?ller Glial Differentiation Reveals a Requirement for Notch Signaling in Postmitotic Cells to Maintain the Glial Fate

Journal: PLoS ONE

doi: 10.1371/journal.pone.0022817

Hes5-GFP expression in developing retina. (A) Hes5-GFP in the E12.5 mouse retina. (B) in situ hybridization for Hes5 mRNA at E13.5. (C,D) Hes5-GFP expressing cells in the E12.5 day retina co-express progenitor markers, PH3 (C) and BrdU (D). (E–G) Hes5-GFP expression at postnatal day 7 is confined to the inner nuclear layer in the maturing Müller glia (Id1+; G)), and is not expressed in the Pax6+ or TuJ1+ inner retinal neurons (E and F, respectively).
Figure Legend Snippet: Hes5-GFP expression in developing retina. (A) Hes5-GFP in the E12.5 mouse retina. (B) in situ hybridization for Hes5 mRNA at E13.5. (C,D) Hes5-GFP expressing cells in the E12.5 day retina co-express progenitor markers, PH3 (C) and BrdU (D). (E–G) Hes5-GFP expression at postnatal day 7 is confined to the inner nuclear layer in the maturing Müller glia (Id1+; G)), and is not expressed in the Pax6+ or TuJ1+ inner retinal neurons (E and F, respectively).

Techniques Used: Expressing, In Situ Hybridization

A combination of BrdU labeling and immunohistochemistry for active Notch signaling shows Notch remains active after the Muller glia have become postmitotic and begun their differentiation. Immunolebeling for active Notch (Notch-ICD; A,C,E, F, F′, F″″, F″″″). An injection of BrdU at postnatal day 5, followed by sacrifice 2 hours later, results in BrdU incorporation in S-phase cells in the peripheral fourth to one third of the retina (Green, C–F). (C,F) The Müller glia in the central retina have already begun their differentiation, as indicated by the strong labeling for Glast, while the progenitor cells in the peripheral retina have only a low level of Glast (C,D). The Notch pathway is active both in the peripheral retinal progenitors (BrdU+;A, C), but also in the central Müller glia (BrdU−/Glast+; A, E, and F-F′″). (G) In situ hybridization at P7 for Notch mRNA expression (H) Hes1 immunohistochemistry localizes to the Muller glia in the INL. All panels except G and H are from the same section, with the arrows and arrowheads in panels A and B pointing out the regions at higher magnification in panels C/D and E/F, respectively. Panels F-F″″″ show the region identified by the arrow in panel F, whereas the arrows in F-F″″″ point to Muller cells (Glast+) that also express Notch-ICD.
Figure Legend Snippet: A combination of BrdU labeling and immunohistochemistry for active Notch signaling shows Notch remains active after the Muller glia have become postmitotic and begun their differentiation. Immunolebeling for active Notch (Notch-ICD; A,C,E, F, F′, F″″, F″″″). An injection of BrdU at postnatal day 5, followed by sacrifice 2 hours later, results in BrdU incorporation in S-phase cells in the peripheral fourth to one third of the retina (Green, C–F). (C,F) The Müller glia in the central retina have already begun their differentiation, as indicated by the strong labeling for Glast, while the progenitor cells in the peripheral retina have only a low level of Glast (C,D). The Notch pathway is active both in the peripheral retinal progenitors (BrdU+;A, C), but also in the central Müller glia (BrdU−/Glast+; A, E, and F-F′″). (G) In situ hybridization at P7 for Notch mRNA expression (H) Hes1 immunohistochemistry localizes to the Muller glia in the INL. All panels except G and H are from the same section, with the arrows and arrowheads in panels A and B pointing out the regions at higher magnification in panels C/D and E/F, respectively. Panels F-F″″″ show the region identified by the arrow in panel F, whereas the arrows in F-F″″″ point to Muller cells (Glast+) that also express Notch-ICD.

Techniques Used: Labeling, Immunohistochemistry, Injection, BrdU Incorporation Assay, In Situ Hybridization, Expressing

2) Product Images from "E-Cadherin Marks a Subset of Inflammatory Dendritic Cells that Promote T Cell-Mediated Colitis"

Article Title: E-Cadherin Marks a Subset of Inflammatory Dendritic Cells that Promote T Cell-Mediated Colitis

Journal: Immunity

doi: 10.1016/j.immuni.2010.03.017

Gr1 + Monocytes Differentiate into E-Cadherin + DCs (A) Colitic BALB/c Rag2 −/− mice were injected i.p. with 2 mg of BrdU. At various time points, whole colon cells were stained for E-cadherin, CD103 and CD11c surface markers, and for intracellular BrdU. Cells were gated on the CD11c hi population and then examined for the percentage of BrdU + cells among each DC subset. Each symbol represents three mice assayed at each time point. (B) Representative staining of E-cadherin, Ki-67, and the relevant isotype control of Ki-67 on gated CD11c hi cells from MLN cell preparations from colitic mice (n = 4). (C) Expression of E-cadherin by CD115 + Gr1 + and Gr1 − monocytes taken from BM of WT and colitic Rag2 −/− mice. Gates applied shown on left panel. Black solid line, E-cadherin expression by Gr1 + CD115 + cells; gray solid line, E-cadherin expression by Gr1 − CD115 + cells; dashed line, E-cadherin expression by Gr1 + CD115 − control cells. Representative plots from three to five individual analyses are shown. (D) Representative staining of E-cadherin and CD11c on flow cytometry-sorted Gr1 + and Gr1 − CD115 + monocytes from pooled BM of colitic BALB/c Rag2 −/− mice (n = 8). (E) The sorted monocytes were cultured overnight in media with or without GM-CSF, and the next day, the cells were harvested and analyzed for E-cadherin and CD11c expression. (F and G) 3 × 10 6 sorted Gr1 + or Gr1 − CD11b + ckit − CD11c − CD115 + monocytes from pooled BM and blood of naive B6 CD45.1 mice were injected i.v. into colitic B6 CD45.2 Rag1 −/− mice, and 36 hr later, various tissues were harvested. Cells were gated on the CD45.1 population, and E-cadherin expression among the CD45.1 + CD11c hi cells was examined. Cells from nontransferred control mice were isolated in parallel. (F) Representative CD45.1 staining on MLN cells. (G) Staining of E-cadherin and CD11c gated on CD45.1 + cells from colitic mice that received Gr1 + (top panel) or Gr1 − (bottom panel) CD115 + monocytes (n = 3 per group). Numbers represent mean ± SD of CD11c hi cells expressing E-cadherin. Representative data from one of three experiments; similar experiments gave the same results. See also Figure S3 .
Figure Legend Snippet: Gr1 + Monocytes Differentiate into E-Cadherin + DCs (A) Colitic BALB/c Rag2 −/− mice were injected i.p. with 2 mg of BrdU. At various time points, whole colon cells were stained for E-cadherin, CD103 and CD11c surface markers, and for intracellular BrdU. Cells were gated on the CD11c hi population and then examined for the percentage of BrdU + cells among each DC subset. Each symbol represents three mice assayed at each time point. (B) Representative staining of E-cadherin, Ki-67, and the relevant isotype control of Ki-67 on gated CD11c hi cells from MLN cell preparations from colitic mice (n = 4). (C) Expression of E-cadherin by CD115 + Gr1 + and Gr1 − monocytes taken from BM of WT and colitic Rag2 −/− mice. Gates applied shown on left panel. Black solid line, E-cadherin expression by Gr1 + CD115 + cells; gray solid line, E-cadherin expression by Gr1 − CD115 + cells; dashed line, E-cadherin expression by Gr1 + CD115 − control cells. Representative plots from three to five individual analyses are shown. (D) Representative staining of E-cadherin and CD11c on flow cytometry-sorted Gr1 + and Gr1 − CD115 + monocytes from pooled BM of colitic BALB/c Rag2 −/− mice (n = 8). (E) The sorted monocytes were cultured overnight in media with or without GM-CSF, and the next day, the cells were harvested and analyzed for E-cadherin and CD11c expression. (F and G) 3 × 10 6 sorted Gr1 + or Gr1 − CD11b + ckit − CD11c − CD115 + monocytes from pooled BM and blood of naive B6 CD45.1 mice were injected i.v. into colitic B6 CD45.2 Rag1 −/− mice, and 36 hr later, various tissues were harvested. Cells were gated on the CD45.1 population, and E-cadherin expression among the CD45.1 + CD11c hi cells was examined. Cells from nontransferred control mice were isolated in parallel. (F) Representative CD45.1 staining on MLN cells. (G) Staining of E-cadherin and CD11c gated on CD45.1 + cells from colitic mice that received Gr1 + (top panel) or Gr1 − (bottom panel) CD115 + monocytes (n = 3 per group). Numbers represent mean ± SD of CD11c hi cells expressing E-cadherin. Representative data from one of three experiments; similar experiments gave the same results. See also Figure S3 .

Techniques Used: Mouse Assay, Injection, Staining, Expressing, Flow Cytometry, Cytometry, Cell Culture, Isolation

3) Product Images from "Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice"

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice

Journal: Journal of Neurotrauma

doi: 10.1089/neu.2010.1563

Progenitor cells differentiate into mature neurons and astrocytes in the hippocampus following traumatic brain injury (TBI). Cells were double-labeled with bromodeoxyuridine (BrdU), and either the neuronal marker NeuN or glial fibrillary acidic protein
Figure Legend Snippet: Progenitor cells differentiate into mature neurons and astrocytes in the hippocampus following traumatic brain injury (TBI). Cells were double-labeled with bromodeoxyuridine (BrdU), and either the neuronal marker NeuN or glial fibrillary acidic protein

Techniques Used: Labeling, Marker

4) Product Images from "Cited3 activates Mef2c to control muscle cell differentiation and survival"

Article Title: Cited3 activates Mef2c to control muscle cell differentiation and survival

Journal: Biology Open

doi: 10.1242/bio.20132550

Knockdown of cited3 results in increased cell death but it does not affect proliferation. Embryos that are injected with the control MO ( A–C ), cited3 MO ( D–F ), cited3 MO + cited3 RNA ( G–I ) were fixed at 2 dpf and immunostained with anti-Mef2 antibody (A,D,G) and TMR red in situ cell death detection kit (B,E,H). Tunel positive cells were counted in non-muscle cells as well as in muscle cells in each group of embryos from 50–100 sections obtained from trunk and tail somites. More apoptotic muscle cells were detected in embryos that are injected with the cited3 MO and fixed at 2 dpf and 3 dpf ( M ). Values were normalized compared to cited3 morphant values. Similarly, embryos injected with the control MO ( J ), cited3 MO ( K ), cited3 MO + cited3 RNA ( L ) was pulsed with BrdU from 30 hpf to 42 hpf, and were fixed and immunostained with anti-BrdU and MF20 antibodies (images of 16 th –18 th somites). There is no significant difference between the three treated groups (J,K,L) when the BrdU-positive muscle cells were counted (data not shown). Scale bars: 25 µm in A and 50 µm in K.
Figure Legend Snippet: Knockdown of cited3 results in increased cell death but it does not affect proliferation. Embryos that are injected with the control MO ( A–C ), cited3 MO ( D–F ), cited3 MO + cited3 RNA ( G–I ) were fixed at 2 dpf and immunostained with anti-Mef2 antibody (A,D,G) and TMR red in situ cell death detection kit (B,E,H). Tunel positive cells were counted in non-muscle cells as well as in muscle cells in each group of embryos from 50–100 sections obtained from trunk and tail somites. More apoptotic muscle cells were detected in embryos that are injected with the cited3 MO and fixed at 2 dpf and 3 dpf ( M ). Values were normalized compared to cited3 morphant values. Similarly, embryos injected with the control MO ( J ), cited3 MO ( K ), cited3 MO + cited3 RNA ( L ) was pulsed with BrdU from 30 hpf to 42 hpf, and were fixed and immunostained with anti-BrdU and MF20 antibodies (images of 16 th –18 th somites). There is no significant difference between the three treated groups (J,K,L) when the BrdU-positive muscle cells were counted (data not shown). Scale bars: 25 µm in A and 50 µm in K.

Techniques Used: Injection, In Situ, TUNEL Assay

5) Product Images from "Regeneration of the zebrafish retinal pigment epithelium after widespread genetic ablation"

Article Title: Regeneration of the zebrafish retinal pigment epithelium after widespread genetic ablation

Journal: PLoS Genetics

doi: 10.1371/journal.pgen.1007939

Proliferative RPE contributes to the regenerated RPE monolayer. (A-A”) Transverse sections from unablated larvae exposed to BrdU from 5-6dpf and pulsed with EdU for 2 hours before fixation at 8dpf. (B-B”) Transverse sections of ablated larvae exposed to BrdU from 0-1dpi and pulsed with EdU for 2 hours before fixation at 3dpi. (A’,B’) Magnified inset of BrdU/EdU. (A”,B”) Magnified inset of BrdU/EdU and DIC. Arrowheads in (B) highlight BrdU + PRs that have integrated into the ONL. Arrow in (B’,B”) highlights a proliferative RPE cell, and arrowheads highlight unpigmented, previously-proliferative RPE-like cell in the injury site. (C) Quantification of BrdU/EdU + and BrdU + nuclei in the injury site. (D,E) Larvae exposed to BrdU 0-1dpi and fixed at 7dpi. (F-G) Larvae exposed to BrdU 3-4dpi and fixed at 7dpi. (H,I) Larvae exposed to BrdU 5-6dpi and fixed at 7dpi. (J) Quantification of the average number of BrdU + cells per section. (K) Quantification of the location of individual BrdU + cells relative to the center of the RPE. The line indicates the average location of BrdU + cells, and the whiskers indicate standard deviation. (L,M) Quantification of BrdU + cells that were labeled 0-1dpi within the RPE at 7dpi in ablated larvae. Analysis of eGFP + BrdU + and GFP - BrdU + cells in the RPE reveal that most BrdU cells in the RPE are eGFP + at 7dpi. (C) Quantification of the location of individual BrdU + cells relative to the center of the RPE indicates that eGFP + BrdU + cells tend to be located toward the center and eGFP - BrdU + localize toward the periphery. Mann-Whitney U Test, * p
Figure Legend Snippet: Proliferative RPE contributes to the regenerated RPE monolayer. (A-A”) Transverse sections from unablated larvae exposed to BrdU from 5-6dpf and pulsed with EdU for 2 hours before fixation at 8dpf. (B-B”) Transverse sections of ablated larvae exposed to BrdU from 0-1dpi and pulsed with EdU for 2 hours before fixation at 3dpi. (A’,B’) Magnified inset of BrdU/EdU. (A”,B”) Magnified inset of BrdU/EdU and DIC. Arrowheads in (B) highlight BrdU + PRs that have integrated into the ONL. Arrow in (B’,B”) highlights a proliferative RPE cell, and arrowheads highlight unpigmented, previously-proliferative RPE-like cell in the injury site. (C) Quantification of BrdU/EdU + and BrdU + nuclei in the injury site. (D,E) Larvae exposed to BrdU 0-1dpi and fixed at 7dpi. (F-G) Larvae exposed to BrdU 3-4dpi and fixed at 7dpi. (H,I) Larvae exposed to BrdU 5-6dpi and fixed at 7dpi. (J) Quantification of the average number of BrdU + cells per section. (K) Quantification of the location of individual BrdU + cells relative to the center of the RPE. The line indicates the average location of BrdU + cells, and the whiskers indicate standard deviation. (L,M) Quantification of BrdU + cells that were labeled 0-1dpi within the RPE at 7dpi in ablated larvae. Analysis of eGFP + BrdU + and GFP - BrdU + cells in the RPE reveal that most BrdU cells in the RPE are eGFP + at 7dpi. (C) Quantification of the location of individual BrdU + cells relative to the center of the RPE indicates that eGFP + BrdU + cells tend to be located toward the center and eGFP - BrdU + localize toward the periphery. Mann-Whitney U Test, * p

Techniques Used: Standard Deviation, Labeling, MANN-WHITNEY

6) Product Images from "Nestin-positive mesenchymal stem cells favour the astroglial lineage in neural progenitors and stem cells by releasing active BMP4"

Article Title: Nestin-positive mesenchymal stem cells favour the astroglial lineage in neural progenitors and stem cells by releasing active BMP4

Journal: BMC Neuroscience

doi: 10.1186/1471-2202-5-33

Nestin-positive MSC conditioned medium act on the GFAP-positive cell death but not on cell proliferation. (A) Characterization of committed cell types in proliferating neurospheres in presence of EGF. We observed that 14.4 ± 7.1%, 7.1± 4.1% and 3.4 ± 1.6% of cells were respectively GFAP-, Tuj1- and O4-positive (these data were obtained by absolute counts on 1576 cells). (B-C) The proliferative capacity of the differentiating neurosphere-derived cells placed in npMSC conditioned medium, nnMSC conditioned medium and in control medium (DEM/F12 + B27) was compared. BrdU incorporation was performed after 48 hours and 4 days in differentiating conditions. Double labelling GFAP-, Tuj1- and O4 with BrdU were performed. The BrdU incorporation by differentiating neurosphere-derived cells placed in the various conditions did not shown significant differences (Statistical test ANOVA, P > 0.05). (D-E) The cell death quantified by propidium iodide incorporation and counting was analysed after 48 hours and 4 days in differentiating conditions and in GFAP-, Tuj1- and O4-positive cells. After 48 hours, a significant decrease of the number of GFAP-positive cells which have incorporated the propidium iodide is observed (***Student T test, p
Figure Legend Snippet: Nestin-positive MSC conditioned medium act on the GFAP-positive cell death but not on cell proliferation. (A) Characterization of committed cell types in proliferating neurospheres in presence of EGF. We observed that 14.4 ± 7.1%, 7.1± 4.1% and 3.4 ± 1.6% of cells were respectively GFAP-, Tuj1- and O4-positive (these data were obtained by absolute counts on 1576 cells). (B-C) The proliferative capacity of the differentiating neurosphere-derived cells placed in npMSC conditioned medium, nnMSC conditioned medium and in control medium (DEM/F12 + B27) was compared. BrdU incorporation was performed after 48 hours and 4 days in differentiating conditions. Double labelling GFAP-, Tuj1- and O4 with BrdU were performed. The BrdU incorporation by differentiating neurosphere-derived cells placed in the various conditions did not shown significant differences (Statistical test ANOVA, P > 0.05). (D-E) The cell death quantified by propidium iodide incorporation and counting was analysed after 48 hours and 4 days in differentiating conditions and in GFAP-, Tuj1- and O4-positive cells. After 48 hours, a significant decrease of the number of GFAP-positive cells which have incorporated the propidium iodide is observed (***Student T test, p

Techniques Used: Activated Clotting Time Assay, Derivative Assay, BrdU Incorporation Assay

7) Product Images from "Null effect of antidepressants on the astrocytes-mediated proliferation of hippocampal progenitor cells in vitro"

Article Title: Null effect of antidepressants on the astrocytes-mediated proliferation of hippocampal progenitor cells in vitro

Journal: Molecular Pain

doi: 10.1186/1744-8069-3-16

Effects of antidepressant-treated astrocytes on the proliferation of NPCs in a contact-dependent culture system . (A) Proliferation of NPCs co-cultured with astrocytes using an overlay co-culture system. After treating astrocytes with antidepressants, NPCs were overlaid on the astrocytes. NPCs were labeled with GFP using a retrovirus. After BrdU immunocytochemical analysis, the nuclei were stained with DAPI (blue). An arrowhead (pink) indicates BrdU and GFP double-positive NPCs, and the arrow (pink) shows BrdU-positive cells. These BrdU-only-positive cells represent astrocytes or NPCs that were not infected with the GFP-expressing retrovirus. Scale bar, 50 μm. (B) Quantification of the proliferation of NPCs. BrdU and GFP double-positive cells were counted. In comparison with naïve astrocytes, those treated with 10 μM of desipramine or fluoxetine did not further increase the proliferation of NPCs. Data shown are mean values ± SEM. One-way ANOVA and Tukey's multiple comparison test were used for data analysis.
Figure Legend Snippet: Effects of antidepressant-treated astrocytes on the proliferation of NPCs in a contact-dependent culture system . (A) Proliferation of NPCs co-cultured with astrocytes using an overlay co-culture system. After treating astrocytes with antidepressants, NPCs were overlaid on the astrocytes. NPCs were labeled with GFP using a retrovirus. After BrdU immunocytochemical analysis, the nuclei were stained with DAPI (blue). An arrowhead (pink) indicates BrdU and GFP double-positive NPCs, and the arrow (pink) shows BrdU-positive cells. These BrdU-only-positive cells represent astrocytes or NPCs that were not infected with the GFP-expressing retrovirus. Scale bar, 50 μm. (B) Quantification of the proliferation of NPCs. BrdU and GFP double-positive cells were counted. In comparison with naïve astrocytes, those treated with 10 μM of desipramine or fluoxetine did not further increase the proliferation of NPCs. Data shown are mean values ± SEM. One-way ANOVA and Tukey's multiple comparison test were used for data analysis.

Techniques Used: Cell Culture, Co-Culture Assay, Labeling, Staining, Infection, Expressing

8) Product Images from "Soluble N-cadherin: A novel inhibitor of VSMC proliferation and intimal thickening"

Article Title: Soluble N-cadherin: A novel inhibitor of VSMC proliferation and intimal thickening

Journal: Vascular Pharmacology

doi: 10.1016/j.vph.2015.11.040

SNC-Fc had no effect on β-catenin signalling. a: Representative Western blots of N-cadherin and GAPDH proteins following 24 h treatment with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF. Chart shows densitometry analysis of N-cadherin normalised by GAPDH, n = 3. b: qPCR analysis of β-catenin target genes, cyclin D1 and p21 levels following 6 h treatment with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF, normalised to 18S, n = 3. c: Galactolight assay (to measure β-galactosidase activity, and thereby β-catenin signalling) measured in arbitrary units in TOPgal VSMCs treated with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF, n = 3. d: Representative images of dual immunocytochemistry for β-galactosidase (green) and BrdU (red) on TOPgal VSMCs treated with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF. Chart shows the percentage of proliferating cells (BrdU positive) that were positive for β-galactosidase, n = 3. Arrowheads indicate some positive cells (red nuclei and green cytoplasm). Scale bars represent 25 μm.
Figure Legend Snippet: SNC-Fc had no effect on β-catenin signalling. a: Representative Western blots of N-cadherin and GAPDH proteins following 24 h treatment with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF. Chart shows densitometry analysis of N-cadherin normalised by GAPDH, n = 3. b: qPCR analysis of β-catenin target genes, cyclin D1 and p21 levels following 6 h treatment with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF, normalised to 18S, n = 3. c: Galactolight assay (to measure β-galactosidase activity, and thereby β-catenin signalling) measured in arbitrary units in TOPgal VSMCs treated with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF, n = 3. d: Representative images of dual immunocytochemistry for β-galactosidase (green) and BrdU (red) on TOPgal VSMCs treated with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF. Chart shows the percentage of proliferating cells (BrdU positive) that were positive for β-galactosidase, n = 3. Arrowheads indicate some positive cells (red nuclei and green cytoplasm). Scale bars represent 25 μm.

Techniques Used: Western Blot, Real-time Polymerase Chain Reaction, Activity Assay, Immunocytochemistry

SNC-Fc had no effect on β-catenin signalling. a: Representative Western blots of N-cadherin and GAPDH proteins following 24 h treatment with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF. Chart shows densitometry analysis of N-cadherin normalised by GAPDH, n = 3. b: qPCR analysis of β-catenin target genes, cyclin D1 and p21 levels following 6 h treatment with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF, normalised to 18S, n = 3. c: Galactolight assay (to measure β-galactosidase activity, and thereby β-catenin signalling) measured in arbitrary units in TOPgal VSMCs treated with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF, n = 3. d: Representative images of dual immunocytochemistry for β-galactosidase (green) and BrdU (red) on TOPgal VSMCs treated with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF. Chart shows the percentage of proliferating cells (BrdU positive) that were positive for β-galactosidase, n = 3. Arrowheads indicate some positive cells (red nuclei and green cytoplasm). Scale bars represent 25 μm.
Figure Legend Snippet: SNC-Fc had no effect on β-catenin signalling. a: Representative Western blots of N-cadherin and GAPDH proteins following 24 h treatment with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF. Chart shows densitometry analysis of N-cadherin normalised by GAPDH, n = 3. b: qPCR analysis of β-catenin target genes, cyclin D1 and p21 levels following 6 h treatment with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF, normalised to 18S, n = 3. c: Galactolight assay (to measure β-galactosidase activity, and thereby β-catenin signalling) measured in arbitrary units in TOPgal VSMCs treated with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF, n = 3. d: Representative images of dual immunocytochemistry for β-galactosidase (green) and BrdU (red) on TOPgal VSMCs treated with 200 pM Fc or SNC-Fc in the presence of bFGF and PDGF. Chart shows the percentage of proliferating cells (BrdU positive) that were positive for β-galactosidase, n = 3. Arrowheads indicate some positive cells (red nuclei and green cytoplasm). Scale bars represent 25 μm.

Techniques Used: Western Blot, Real-time Polymerase Chain Reaction, Activity Assay, Immunocytochemistry

SNC-Fc reduced intimal thickening in an ex vivo organ culture model. Representative images of: a and b: Elastin van Gieson staining to show intimal size. c and d: Immunohistochemistry for BrdU incorporation (brown nuclei: proliferation). e and f: ISEL (brown nuclei: apoptosis). g and h: Immunohistochemistry for QBend10 (brown colour: endothelial cells). i and j: Dual immunohistochemistry for QBend10 (pink, endothelial cells) and ISEL (brown nuclei, apoptosis). k and l: Higher power images of the dual QBend10 (pink) and ISEL (brown) to clearly show the double staining in i. Dotted line delineates the intimal:medial boundary. Arrowheads indicate some of the positive cells. Scale bars in a-d and g and h represent 20 μm, scale bars in e, f, i, j, k and l represent 40 μm, n = 5.
Figure Legend Snippet: SNC-Fc reduced intimal thickening in an ex vivo organ culture model. Representative images of: a and b: Elastin van Gieson staining to show intimal size. c and d: Immunohistochemistry for BrdU incorporation (brown nuclei: proliferation). e and f: ISEL (brown nuclei: apoptosis). g and h: Immunohistochemistry for QBend10 (brown colour: endothelial cells). i and j: Dual immunohistochemistry for QBend10 (pink, endothelial cells) and ISEL (brown nuclei, apoptosis). k and l: Higher power images of the dual QBend10 (pink) and ISEL (brown) to clearly show the double staining in i. Dotted line delineates the intimal:medial boundary. Arrowheads indicate some of the positive cells. Scale bars in a-d and g and h represent 20 μm, scale bars in e, f, i, j, k and l represent 40 μm, n = 5.

Techniques Used: Ex Vivo, Organ Culture, Staining, Immunohistochemistry, BrdU Incorporation Assay, Double Staining

9) Product Images from "Support for the immortal strand hypothesis"

Article Title: Support for the immortal strand hypothesis

Journal: The Journal of Cell Biology

doi: 10.1083/jcb.200502073

After a 2-d exposure to BrdU, cells from adult murine forebrain ventricles demonstrate heterogenous BrdU labeling in vitro. (A) BrdU retention strategy: (1) Each double strand (1 chromosome) represents 10 chromosomes of a mouse cell. Cells are unlabeled for BrdU (black). (2) During multiple rounds of DNA synthesis, BrdU (green) is taken up and distributed in both symmetric and asymmetric divisions in the presence of BrdU. (3) BrdU is removed and the daughter cells now undergo DNA synthesis in the absence of BrdU. (4) BrdU should be retained if labeled chromosomes are cosegregated as immortal strands into SCs. (B) BrdU–neurosphere assay. (1) Cells from adult forebrain lateral ventricles are cultured for 7 d at clonal density. (2) After dissociation, cells are pulsed with BrdU for 2 d, at 3 DIV. (3) BrdU is removed and cells are passaged at clonal density for an additional 7 d. (4) Cells are examined (A), passaged (B), or differentiated (C). (C) 10 d after BrdU exposure, cell clones still contained heavily BrdU(+) cells (arrows) and BrdU(−) cells. The retention of BrdU(+), in cells seeded at clonal density, suggests that BrdU(+) cells give rise to both labeled and unlabeled progeny. (i) Bright field shows 3 d cell clumps; (ii) histone-labeled nuclei are red, BrdU-labeled cells are green; merge shows overlap as yellow.
Figure Legend Snippet: After a 2-d exposure to BrdU, cells from adult murine forebrain ventricles demonstrate heterogenous BrdU labeling in vitro. (A) BrdU retention strategy: (1) Each double strand (1 chromosome) represents 10 chromosomes of a mouse cell. Cells are unlabeled for BrdU (black). (2) During multiple rounds of DNA synthesis, BrdU (green) is taken up and distributed in both symmetric and asymmetric divisions in the presence of BrdU. (3) BrdU is removed and the daughter cells now undergo DNA synthesis in the absence of BrdU. (4) BrdU should be retained if labeled chromosomes are cosegregated as immortal strands into SCs. (B) BrdU–neurosphere assay. (1) Cells from adult forebrain lateral ventricles are cultured for 7 d at clonal density. (2) After dissociation, cells are pulsed with BrdU for 2 d, at 3 DIV. (3) BrdU is removed and cells are passaged at clonal density for an additional 7 d. (4) Cells are examined (A), passaged (B), or differentiated (C). (C) 10 d after BrdU exposure, cell clones still contained heavily BrdU(+) cells (arrows) and BrdU(−) cells. The retention of BrdU(+), in cells seeded at clonal density, suggests that BrdU(+) cells give rise to both labeled and unlabeled progeny. (i) Bright field shows 3 d cell clumps; (ii) histone-labeled nuclei are red, BrdU-labeled cells are green; merge shows overlap as yellow.

Techniques Used: Labeling, In Vitro, DNA Synthesis, Neurosphere Assay, Cell Culture, Clone Assay

NSCs are fast dividing cells in vitro, and show BrdU retention. (A) Distribution of DiI, initially and after 1 wk in vitro. DiI signal decreases as a result of DiI dilution via cell proliferation. (DiI pos) DiI(HI+) fraction; (DiI neg) DiI(LOW+) fraction. (B) DiI(HI+) fraction of slowly dividing neurosphere cells where DiI signal is vivid. (i) Dissociated cells in bright field; (ii) DiI signal in red. (C) DiI(LOW+) fraction of rapidly dividing neurosphere cells where DiI signal is noticeably lower than in DiI(HI+) fraction. (i) Dissociated cells in bright field; (ii) DiI signal in red. (D) Data shows DiI(HI+) population (10% of total). As expected, slowly cycling cells do not greatly attenuate BrdU or DiI. The BrdU(−) population may be the same 1% of cells that are BrdU(−) immediately after BrdU exposure. (E) Data shows DiI(LOW+) population (63% of total). A subset of BrdU(+) cells are DiI(LOW+) after extended cell proliferation in vitro. BrdU retention in rapidly cycling cells suggests these are cosegregating their DNA. (F) Comparison in clonal sphere formation between DiI(HI+) and DiI(LOW+) fractions. The majority of neurospheres arise from the fast-cycling DiI(LOW+) population (7.5-fold increase over DiI[HI+]). This suggests SCs are in this actively proliferating fraction.
Figure Legend Snippet: NSCs are fast dividing cells in vitro, and show BrdU retention. (A) Distribution of DiI, initially and after 1 wk in vitro. DiI signal decreases as a result of DiI dilution via cell proliferation. (DiI pos) DiI(HI+) fraction; (DiI neg) DiI(LOW+) fraction. (B) DiI(HI+) fraction of slowly dividing neurosphere cells where DiI signal is vivid. (i) Dissociated cells in bright field; (ii) DiI signal in red. (C) DiI(LOW+) fraction of rapidly dividing neurosphere cells where DiI signal is noticeably lower than in DiI(HI+) fraction. (i) Dissociated cells in bright field; (ii) DiI signal in red. (D) Data shows DiI(HI+) population (10% of total). As expected, slowly cycling cells do not greatly attenuate BrdU or DiI. The BrdU(−) population may be the same 1% of cells that are BrdU(−) immediately after BrdU exposure. (E) Data shows DiI(LOW+) population (63% of total). A subset of BrdU(+) cells are DiI(LOW+) after extended cell proliferation in vitro. BrdU retention in rapidly cycling cells suggests these are cosegregating their DNA. (F) Comparison in clonal sphere formation between DiI(HI+) and DiI(LOW+) fractions. The majority of neurospheres arise from the fast-cycling DiI(LOW+) population (7.5-fold increase over DiI[HI+]). This suggests SCs are in this actively proliferating fraction.

Techniques Used: In Vitro

Live cell imaging supports nonrandom segregation of DNA. (A) Schematic showing BrdU imaging strategy. (1) Each double strand (1 chromosome) represents 10 chromosomes of a mouse cell. Cells are unlabeled for BrdU (black). (2) During DNA synthesis, BrdU (green) is taken up for exactly one division in the presence of BrdU. (3) BrdU is removed and the daughters enter a second round of DNA synthesis in the absence of BrdU. (4) Division events after the second round of DNA synthesis should show BrdU asymmetry if groups of unlabeled chromosomes are cosegregated as immortal strands into SCs. (B) A clone imaged in real time. After one division event, BrdU was removed, and colony was fixed after two further cell divisions in the absence of BrdU. Arrow indicates a cell that has cleared all BrdU signal. (i) Bright field shows clone; (ii) histone-labeled nuclei are red; (iii) BrdU is indicated by green; (iv) merge of histone and BrdU. (C) Lineage diagrams from four clones traced (ii–iv show asymmetric DNA partitioning). Clone (ii) is the same shown in B. Each lineage represents divisions of one single cell, plated in the presence of BrdU, which is taken up during the first division initially labeling daughter nuclei (green), as demonstrated in clone (i). BrdU was removed after this one division, and cells continued proliferating until analysis. Note, the presence of BrdU(+) is inferred in parental cells from their offspring. Dead cells were observed to disintegrate while imaging, before analysis. (D) Summary of clones traced. 6 out of 15 clones demonstrated asymmetric partitioning of new and old DNA.
Figure Legend Snippet: Live cell imaging supports nonrandom segregation of DNA. (A) Schematic showing BrdU imaging strategy. (1) Each double strand (1 chromosome) represents 10 chromosomes of a mouse cell. Cells are unlabeled for BrdU (black). (2) During DNA synthesis, BrdU (green) is taken up for exactly one division in the presence of BrdU. (3) BrdU is removed and the daughters enter a second round of DNA synthesis in the absence of BrdU. (4) Division events after the second round of DNA synthesis should show BrdU asymmetry if groups of unlabeled chromosomes are cosegregated as immortal strands into SCs. (B) A clone imaged in real time. After one division event, BrdU was removed, and colony was fixed after two further cell divisions in the absence of BrdU. Arrow indicates a cell that has cleared all BrdU signal. (i) Bright field shows clone; (ii) histone-labeled nuclei are red; (iii) BrdU is indicated by green; (iv) merge of histone and BrdU. (C) Lineage diagrams from four clones traced (ii–iv show asymmetric DNA partitioning). Clone (ii) is the same shown in B. Each lineage represents divisions of one single cell, plated in the presence of BrdU, which is taken up during the first division initially labeling daughter nuclei (green), as demonstrated in clone (i). BrdU was removed after this one division, and cells continued proliferating until analysis. Note, the presence of BrdU(+) is inferred in parental cells from their offspring. Dead cells were observed to disintegrate while imaging, before analysis. (D) Summary of clones traced. 6 out of 15 clones demonstrated asymmetric partitioning of new and old DNA.

Techniques Used: Live Cell Imaging, Imaging, DNA Synthesis, Labeling, Clone Assay

Cell division inhibition of BrdU-retaining cells suggests asymmetric DNA partitioning. (A) BrdU distribution in a cell arrested during cytokinesis, 10 d after BrdU. Arrows indicate symmetric BrdU(+) nuclei in same cell. (i) Bright field shows binucleate cell; (ii) histone-labeled nuclei are red; (iii) BrdU is green; (iv) merge of histone and BrdU. (B) BrdU distribution in a cell arrested during cytokinesis, 10 d after BrdU. Arrows indicate BrdU(+) nucleus, adjacent to BrdU(−) nucleus, in same cell. (i) Bright field shows binucleate cell; (ii) histone-labeled nuclei are red; (iii) BrdU is green; (iv) shows merge of histone and BrdU. (C) BrdU distribution in binucleate cell population treated with cytochalasin D. Uneven segregation of labeled DNA to daughter nuclei occurs in 10% of the binucleate cell population. (D) Confocal microscopy of BrdU-exposed cells arrested during karyokinesis, 10 d after BrdU exposure. Upon removal of inhibitor, cells were timed for fixation at late anaphase or telophase. Mitotic cells were observed segregating labeled DNA nonrandomly to one daughter in top row (arrows), as opposed to the even segregation of BrdU in bottom examples (arrowheads). BrdU labeling was confirmed at all focal planes. (i) Bright field shows mitotic cells; (ii) histone-labeled nuclei are red; (iii) BrdU is green; (iv) merge of histone and BrdU. (E) Cell doublets arising from 10 d after BrdU cells inhibited during karyokinesis. Cells released from inhibition were allowed to complete mitosis. Uneven labeling of BrdU(+/−) daughter nuclei was again apparent (arrows). (i) Bright field shows two cells; (ii) histone-labeled nuclei are red; (iii) BrdU is indicated by green; (iv) merge of histone and BrdU. (F) Cell doublets arising from 10 d after BrdU cells inhibited during karyokinesis. Cells released from inhibition were allowed to complete mitosis. Some doublets displayed evenly labeled BrdU(+) daughters (arrows) or unlabeled, BrdU(−) doublets (arrowheads). (i) Bright field shows binucleate cells; (ii) merge of histone-labeled nuclei (red) and BrdU (green).
Figure Legend Snippet: Cell division inhibition of BrdU-retaining cells suggests asymmetric DNA partitioning. (A) BrdU distribution in a cell arrested during cytokinesis, 10 d after BrdU. Arrows indicate symmetric BrdU(+) nuclei in same cell. (i) Bright field shows binucleate cell; (ii) histone-labeled nuclei are red; (iii) BrdU is green; (iv) merge of histone and BrdU. (B) BrdU distribution in a cell arrested during cytokinesis, 10 d after BrdU. Arrows indicate BrdU(+) nucleus, adjacent to BrdU(−) nucleus, in same cell. (i) Bright field shows binucleate cell; (ii) histone-labeled nuclei are red; (iii) BrdU is green; (iv) shows merge of histone and BrdU. (C) BrdU distribution in binucleate cell population treated with cytochalasin D. Uneven segregation of labeled DNA to daughter nuclei occurs in 10% of the binucleate cell population. (D) Confocal microscopy of BrdU-exposed cells arrested during karyokinesis, 10 d after BrdU exposure. Upon removal of inhibitor, cells were timed for fixation at late anaphase or telophase. Mitotic cells were observed segregating labeled DNA nonrandomly to one daughter in top row (arrows), as opposed to the even segregation of BrdU in bottom examples (arrowheads). BrdU labeling was confirmed at all focal planes. (i) Bright field shows mitotic cells; (ii) histone-labeled nuclei are red; (iii) BrdU is green; (iv) merge of histone and BrdU. (E) Cell doublets arising from 10 d after BrdU cells inhibited during karyokinesis. Cells released from inhibition were allowed to complete mitosis. Uneven labeling of BrdU(+/−) daughter nuclei was again apparent (arrows). (i) Bright field shows two cells; (ii) histone-labeled nuclei are red; (iii) BrdU is indicated by green; (iv) merge of histone and BrdU. (F) Cell doublets arising from 10 d after BrdU cells inhibited during karyokinesis. Cells released from inhibition were allowed to complete mitosis. Some doublets displayed evenly labeled BrdU(+) daughters (arrows) or unlabeled, BrdU(−) doublets (arrowheads). (i) Bright field shows binucleate cells; (ii) merge of histone-labeled nuclei (red) and BrdU (green).

Techniques Used: Inhibition, Labeling, Confocal Microscopy

10) Product Images from "Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice"

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice

Journal: Journal of Neurotrauma

doi: 10.1089/neu.2010.1563

Progenitor cells differentiate into mature neurons and astrocytes in the hippocampus following traumatic brain injury (TBI). Cells were double-labeled with bromodeoxyuridine (BrdU), and either the neuronal marker NeuN or glial fibrillary acidic protein
Figure Legend Snippet: Progenitor cells differentiate into mature neurons and astrocytes in the hippocampus following traumatic brain injury (TBI). Cells were double-labeled with bromodeoxyuridine (BrdU), and either the neuronal marker NeuN or glial fibrillary acidic protein

Techniques Used: Labeling, Marker

11) Product Images from "TGF-? Signaling in T cells is Essential for CD8 T Cell Suppression and Viral Persistence In Vivo"

Article Title: TGF-? Signaling in T cells is Essential for CD8 T Cell Suppression and Viral Persistence In Vivo

Journal: Immunity

doi: 10.1016/j.immuni.2009.06.015

dnTGFBRII mice show increased survival and reduced levels of Bim in virus-specific CD8 T cells A-B) WT (black bars) or dnTGFBRII (white bars) mice were infected with LCMV Cl 13. Spleen cells were isolated at day 7 and 9 pi and BrdU incorporation (A) and Annexin V staining (B) of H2D b GP 33-41 tetramer + CD8 + T cells were determined. Histograms display a representative mouse per group and numbers indicate the frequency of cells within regions. C-D) Virus-specific CD8 T cells were FACS-isolated from WT mice infected with LCMV ARM or Cl 13 (C) or dnTGFBRII mice infected with Cl 13 (D) at days 8-10 pi. Cells were processed by immunoblot and the levels of Bim EL and Bim L isoforms are shown. Phospholipase-γ (PLC-γ) was used as loading control. Results are representative of 2-3 independent experiments with 3-5 mice per group (WT vs dnTGFBRII, * p
Figure Legend Snippet: dnTGFBRII mice show increased survival and reduced levels of Bim in virus-specific CD8 T cells A-B) WT (black bars) or dnTGFBRII (white bars) mice were infected with LCMV Cl 13. Spleen cells were isolated at day 7 and 9 pi and BrdU incorporation (A) and Annexin V staining (B) of H2D b GP 33-41 tetramer + CD8 + T cells were determined. Histograms display a representative mouse per group and numbers indicate the frequency of cells within regions. C-D) Virus-specific CD8 T cells were FACS-isolated from WT mice infected with LCMV ARM or Cl 13 (C) or dnTGFBRII mice infected with Cl 13 (D) at days 8-10 pi. Cells were processed by immunoblot and the levels of Bim EL and Bim L isoforms are shown. Phospholipase-γ (PLC-γ) was used as loading control. Results are representative of 2-3 independent experiments with 3-5 mice per group (WT vs dnTGFBRII, * p

Techniques Used: Mouse Assay, Infection, Isolation, BrdU Incorporation Assay, Staining, FACS, Planar Chromatography

Direct and indirect TGF-β effects on virus-specific CD8 T cells A-B) P14-WT (black bars) and P14-dnTGFBRII (white bars) CD8 + T cells were co-transfered into WT mice 1 day before LCMV Cl 13 infection. BrdU incorporation (A) and Annexin V staining (B) of P14 cells were determined at day 8 pi. Bar graphs depict the average frequency of positive cells ± sd. Histograms display a representative mouse and numbers indicate the frequency of cells within regions. C-F) WT:dnTGFBRII mixed BM chimeras were processed to analyze CD45.1 + WT and CD45.2 + dnTGFBRII CD8 T cells before (pre-infection) and at day 8 after Cl 13 infection. C) Total CD8 T cells in blood before infection and within spleen D b /NP 396-404 and D b /GP 33-41 tetramer + cells at day 8 pi. D) Total numbers of tetramer + CD8 + cells per spleen at day 8 pi. E) Production of IFN-γ and TNF-α after GP 33-41 and GP 276-286 LCMV peptide-stimulation at day 8 pi. F) PD-1 expression within D b /GP 33-41 tetramer + cells at day 8 pi. Dot plots display a representative mouse and numbers indicate the frequency of cells within regions. Results are representative of two independent experiments with 4-8 mice per group.
Figure Legend Snippet: Direct and indirect TGF-β effects on virus-specific CD8 T cells A-B) P14-WT (black bars) and P14-dnTGFBRII (white bars) CD8 + T cells were co-transfered into WT mice 1 day before LCMV Cl 13 infection. BrdU incorporation (A) and Annexin V staining (B) of P14 cells were determined at day 8 pi. Bar graphs depict the average frequency of positive cells ± sd. Histograms display a representative mouse and numbers indicate the frequency of cells within regions. C-F) WT:dnTGFBRII mixed BM chimeras were processed to analyze CD45.1 + WT and CD45.2 + dnTGFBRII CD8 T cells before (pre-infection) and at day 8 after Cl 13 infection. C) Total CD8 T cells in blood before infection and within spleen D b /NP 396-404 and D b /GP 33-41 tetramer + cells at day 8 pi. D) Total numbers of tetramer + CD8 + cells per spleen at day 8 pi. E) Production of IFN-γ and TNF-α after GP 33-41 and GP 276-286 LCMV peptide-stimulation at day 8 pi. F) PD-1 expression within D b /GP 33-41 tetramer + cells at day 8 pi. Dot plots display a representative mouse and numbers indicate the frequency of cells within regions. Results are representative of two independent experiments with 4-8 mice per group.

Techniques Used: Mouse Assay, Infection, BrdU Incorporation Assay, Staining, Expressing

12) Product Images from "Intracellular Viral Processing, Not Single-Stranded DNA Accumulation, Is Crucial for Recombinant Adeno-Associated Virus Transduction"

Article Title: Intracellular Viral Processing, Not Single-Stranded DNA Accumulation, Is Crucial for Recombinant Adeno-Associated Virus Transduction

Journal: Journal of Virology

doi: 10.1128/JVI.78.24.13678-13686.2004

Detection of BrdU-labeled viral ss DNA by immunofluorescence staining. HeLa cells were infected with BrdU-labeled AAV2 vector at an MOI of 10,000, and the incorporated BrdU in the viral genome was detected at (A) 10 min, (B) 6 h, (C) 12 h, (D) 24 h, and (E) 48 h postinfection by using a BrdU antibody. Under nondenaturing (Non D) conditions, no signal is detectable. Under denaturing (Denatured) conditions, BrdU signal (red) can be detected in the nucleus, which is stained with Hoechst 33342 (blue) at 6 h postinfection or later on. The strongest signal is detected 24 h postinfection, which is reduced following that time point. As a negative (neg) control, the same time course experiment was performed by using a nonlabeled AAV2-CMV-GFP vector which is shown at 12 h postinfection (F). (G, H, and I) SS plasmid DNA was prepared from bacteria that were grown in Luria-Bertani medium supplemented with BrdU. HeLa cells were transfected with 10 ng (G) or 100 pg (H) of labeled ss DNA or 200 ng of unlabeled ss DNA (I). Cells were fixed at 6 h posttransfection and stained with BrdU antibody (red) under nondenaturing conditions. The nucleus was stained with Hoechst 33342 (blue). The scale bar is 20 μm for all panels.
Figure Legend Snippet: Detection of BrdU-labeled viral ss DNA by immunofluorescence staining. HeLa cells were infected with BrdU-labeled AAV2 vector at an MOI of 10,000, and the incorporated BrdU in the viral genome was detected at (A) 10 min, (B) 6 h, (C) 12 h, (D) 24 h, and (E) 48 h postinfection by using a BrdU antibody. Under nondenaturing (Non D) conditions, no signal is detectable. Under denaturing (Denatured) conditions, BrdU signal (red) can be detected in the nucleus, which is stained with Hoechst 33342 (blue) at 6 h postinfection or later on. The strongest signal is detected 24 h postinfection, which is reduced following that time point. As a negative (neg) control, the same time course experiment was performed by using a nonlabeled AAV2-CMV-GFP vector which is shown at 12 h postinfection (F). (G, H, and I) SS plasmid DNA was prepared from bacteria that were grown in Luria-Bertani medium supplemented with BrdU. HeLa cells were transfected with 10 ng (G) or 100 pg (H) of labeled ss DNA or 200 ng of unlabeled ss DNA (I). Cells were fixed at 6 h posttransfection and stained with BrdU antibody (red) under nondenaturing conditions. The nucleus was stained with Hoechst 33342 (blue). The scale bar is 20 μm for all panels.

Techniques Used: Labeling, Immunofluorescence, Staining, Infection, Plasmid Preparation, Transfection

13) Product Images from "Metformin Acts on Two Different Molecular Pathways to Enhance Adult Neural Precursor Proliferation/Self-Renewal and Differentiation"

Article Title: Metformin Acts on Two Different Molecular Pathways to Enhance Adult Neural Precursor Proliferation/Self-Renewal and Differentiation

Journal: Stem Cell Reports

doi: 10.1016/j.stemcr.2015.10.014

TAp73 Is Required for Metformin-Induced Self-Renewal and Proliferation of Adult Neural Precursors (A and B) Primary neurospheres (NSs) were cultured from the SVZ of adult mice (3 months old) in the presence or absence of metformin (Met) and treated with the pan-PKC inhibitor, chelerythrine. (B) Quantification of primary neurospheres following 6 days in culture was done based on representative micrographs shown in (A). Scale bar, 200 μm. ∗∗ p ≤ 0.01 (pooled data from four independent experiments). (C) The SVZ of 3-month-old WT and CBPS436A-KI mice was dissected and cultured, and primary neurosphere formation was quantified 6 days later in the presence and absence of metformin. ∗∗ p ≤ 0.01. (pooled data from three independent experiments). (D) Metformin- or PBS-treated primary neurospheres (as in A) from both genotypes were passaged into untreated media, and the number of secondary neurospheres was quantified 4 days later. ∗∗ p ≤ 0.01. (pooled data from three independent experiments). (E) qRT-PCR for TAp73 mRNA performed on RNA extracted from primary neurospheres grown in the presence or absence of metformin. ∗∗ p ≤ 0.01 (n = 4 for each group). (F–H) Primary neurospheres were cultured from the SVZ of 3-month-old TAp73 +/+ and TAp73 −/− mice in either the presence or absence of 500 nM metformin. Scale bar, 200 μm. (G) Quantification of primary neurospheres from both genotypes following metformin exposure was done based on representative micrographs shown in (F). ∗∗ p ≤ 0.01 (pooled data from four independent experiments). (H) Primary neurospheres cultured with or without metformin (as in A) from both genotypes were passaged into untreated media, and secondary neurospheres were counted 4 days later. ∗∗ p ≤ 0.01 (pooled data from four independent experiments). (I) Confocal micrographs of representative coronal sections through the lateral ventricles of metformin- or PBS-injected TAp73 +/+ and TAp73 −/− mice, stained for BrdU (green) 24 hr after BrdU injection. Sections are counterstained for Hoechst 33258 (blue). Scale bar, 100 μm. (J and K) Quantification of the total number of BrdU-positive (BrdU+ve) cells in the SVZ (J) and SGZ (K) of metformin- or PBS-treated mice of both genotypes, as pictured in (I). ∗∗∗ p ≤ 0.001; ∗∗ p ≤ 0.01 (pooled data from three independent experiments). Error bars indicate SEM.
Figure Legend Snippet: TAp73 Is Required for Metformin-Induced Self-Renewal and Proliferation of Adult Neural Precursors (A and B) Primary neurospheres (NSs) were cultured from the SVZ of adult mice (3 months old) in the presence or absence of metformin (Met) and treated with the pan-PKC inhibitor, chelerythrine. (B) Quantification of primary neurospheres following 6 days in culture was done based on representative micrographs shown in (A). Scale bar, 200 μm. ∗∗ p ≤ 0.01 (pooled data from four independent experiments). (C) The SVZ of 3-month-old WT and CBPS436A-KI mice was dissected and cultured, and primary neurosphere formation was quantified 6 days later in the presence and absence of metformin. ∗∗ p ≤ 0.01. (pooled data from three independent experiments). (D) Metformin- or PBS-treated primary neurospheres (as in A) from both genotypes were passaged into untreated media, and the number of secondary neurospheres was quantified 4 days later. ∗∗ p ≤ 0.01. (pooled data from three independent experiments). (E) qRT-PCR for TAp73 mRNA performed on RNA extracted from primary neurospheres grown in the presence or absence of metformin. ∗∗ p ≤ 0.01 (n = 4 for each group). (F–H) Primary neurospheres were cultured from the SVZ of 3-month-old TAp73 +/+ and TAp73 −/− mice in either the presence or absence of 500 nM metformin. Scale bar, 200 μm. (G) Quantification of primary neurospheres from both genotypes following metformin exposure was done based on representative micrographs shown in (F). ∗∗ p ≤ 0.01 (pooled data from four independent experiments). (H) Primary neurospheres cultured with or without metformin (as in A) from both genotypes were passaged into untreated media, and secondary neurospheres were counted 4 days later. ∗∗ p ≤ 0.01 (pooled data from four independent experiments). (I) Confocal micrographs of representative coronal sections through the lateral ventricles of metformin- or PBS-injected TAp73 +/+ and TAp73 −/− mice, stained for BrdU (green) 24 hr after BrdU injection. Sections are counterstained for Hoechst 33258 (blue). Scale bar, 100 μm. (J and K) Quantification of the total number of BrdU-positive (BrdU+ve) cells in the SVZ (J) and SGZ (K) of metformin- or PBS-treated mice of both genotypes, as pictured in (I). ∗∗∗ p ≤ 0.001; ∗∗ p ≤ 0.01 (pooled data from three independent experiments). Error bars indicate SEM.

Techniques Used: Cell Culture, Mouse Assay, Quantitative RT-PCR, Injection, Staining

14) Product Images from "Mitochondrial DNA damage and subsequent activation of Z-DNA binding protein 1 links oxidative stress to inflammation in epithelial cells"

Article Title: Mitochondrial DNA damage and subsequent activation of Z-DNA binding protein 1 links oxidative stress to inflammation in epithelial cells

Journal: Scientific Reports

doi: 10.1038/s41598-018-19216-1

Damaged mitochondrial DNA activates ZBP1/TBK1/IRF3 signaling pathway. Interaction between BrDU-labelled mtDNA and TFAM ( A ) and BrDU-labelled mtDNA and ZBP1 ( B ) in control and GOx-treated cells at 1 h. ( C ) Interaction between ZBP1/TBK1, IRF3/TBK1, ZBP1/P-Tyr and ZBP1/P-Ser in control and GOx-treated cells at 1 h. ( D ) Interaction between IRF3/TBK1 in pre-treated with 3 μM of CsA in GOx-treated BEAS 2B cells. ( E ) Level of ZBP1 depletion, shown by Western blotting. ( F ) Expression of IL-6 and IL-8 in unstressed and GOx-treated (0.006 U/ml for 1 h) in control and ZBP1-depleted BEAS2B cells. Representative images of n = 3 independent experiments are shown. Data represent average ± SEM of n = 5 biological replicates. *p
Figure Legend Snippet: Damaged mitochondrial DNA activates ZBP1/TBK1/IRF3 signaling pathway. Interaction between BrDU-labelled mtDNA and TFAM ( A ) and BrDU-labelled mtDNA and ZBP1 ( B ) in control and GOx-treated cells at 1 h. ( C ) Interaction between ZBP1/TBK1, IRF3/TBK1, ZBP1/P-Tyr and ZBP1/P-Ser in control and GOx-treated cells at 1 h. ( D ) Interaction between IRF3/TBK1 in pre-treated with 3 μM of CsA in GOx-treated BEAS 2B cells. ( E ) Level of ZBP1 depletion, shown by Western blotting. ( F ) Expression of IL-6 and IL-8 in unstressed and GOx-treated (0.006 U/ml for 1 h) in control and ZBP1-depleted BEAS2B cells. Representative images of n = 3 independent experiments are shown. Data represent average ± SEM of n = 5 biological replicates. *p

Techniques Used: Western Blot, Expressing

15) Product Images from "Active Dentate Granule Cells Encode Experience to Promote the Addition of Adult-Born Hippocampal Neurons"

Article Title: Active Dentate Granule Cells Encode Experience to Promote the Addition of Adult-Born Hippocampal Neurons

Journal: The Journal of Neuroscience

doi: 10.1523/JNEUROSCI.3417-16.2017

Multiple virtual Enr environments cumulatively promoted hippocampal neuronal addition. A , Experimental timeline. Mice were exposed to either a single Enr environment or multiple Enr environments (2/d) during each exposure session. B , Left, Representative images of BrdU+ cells in the GCL of single- and multiple enrichment-exposed mice. Scale bar, 30 μm. Right, Plots of the number of BrdU+ cells in the GCL of mice exposed to the multiple enrichment and single enrichment groups (single enrichment: 1256.7 ± 68.5 cells, multiple enrichment: 1982.4 ± 214.7 cells, 2-tailed unpaired t test p = 0.007; n = 6,5). C , Left, Representative images of GFP+ cell clusters in the GCL of single- and multiple enrichment-exposed mice. Scale bar, 30 μm. Right, Plots of the average cell cluster size for mice exposed to multiple enrichment compared with those exposed to single enrichment (single enrichment: 1.64 ± 0.14 cells/cluster; multiple enrichment: 2.15 ± 0.16 cells/cluster; 2-tailed unpaired t test p = 0.019; n = 3,3). Scale bars, 30 μm. Dots represent individual animals. Statistical significance was defined as * p
Figure Legend Snippet: Multiple virtual Enr environments cumulatively promoted hippocampal neuronal addition. A , Experimental timeline. Mice were exposed to either a single Enr environment or multiple Enr environments (2/d) during each exposure session. B , Left, Representative images of BrdU+ cells in the GCL of single- and multiple enrichment-exposed mice. Scale bar, 30 μm. Right, Plots of the number of BrdU+ cells in the GCL of mice exposed to the multiple enrichment and single enrichment groups (single enrichment: 1256.7 ± 68.5 cells, multiple enrichment: 1982.4 ± 214.7 cells, 2-tailed unpaired t test p = 0.007; n = 6,5). C , Left, Representative images of GFP+ cell clusters in the GCL of single- and multiple enrichment-exposed mice. Scale bar, 30 μm. Right, Plots of the average cell cluster size for mice exposed to multiple enrichment compared with those exposed to single enrichment (single enrichment: 1.64 ± 0.14 cells/cluster; multiple enrichment: 2.15 ± 0.16 cells/cluster; 2-tailed unpaired t test p = 0.019; n = 3,3). Scale bars, 30 μm. Dots represent individual animals. Statistical significance was defined as * p

Techniques Used: Mouse Assay

Optical silencing of DGCs abolished enrichment-induced hippocampal neurogenesis. A , Experimental timeline. Mice were injected with AAV-CaMKII-GFP or AAV-CaMKII-Halo and implanted with head mounts for optical stimulation, given 2 weeks to recover, and then given a single intraperitoneal BrdU injection and exposed to either an Enr or Std environment for 12 d (2 h/d) with light stimulation (1 Hz, 5 ms in duration). B , Left, Representative images of the GCL of the DG from mice in the GFP and Halo groups under Std and Enr environments, respectively. Scale bar, 30 μm. Right, Plots of BrdU+ cells in the GCL of mice in the GFP-Enr group compared with those in the GFP-Std group (relative fold change in Brdu: GFP-Std: 1.00 ± 0.17, GFP-Enr: 1.90 ± 0.32, 2-tailed unpaired t test p = 0.025),and BrdU+ cells in the GCL of mice in the Halo-Enr compared with those in the Halo-Std group (Halo-Std: 1.00 ± 0.20, Halo-Enr: 1.07 ± 0.20, 2-tailed unpaired t test p = 0.818). C , Left, Representative images of c-fos staining in the GCL of the DG of AAV-CaMKII-GFP and AAV-CaMKII-Halo injected mice exposed to an Enr or Std environment. Right, Plots of c-Fos+ cells in the GCL of mice in the GFP-Enr condition and GFP-Std condition (relative fold change c-Fos+ cells/field: GFP-Std: 1.00 ± 0.19, GFP-Enr: 3.70 ± 0.46; 2-tailed unpaired t test p
Figure Legend Snippet: Optical silencing of DGCs abolished enrichment-induced hippocampal neurogenesis. A , Experimental timeline. Mice were injected with AAV-CaMKII-GFP or AAV-CaMKII-Halo and implanted with head mounts for optical stimulation, given 2 weeks to recover, and then given a single intraperitoneal BrdU injection and exposed to either an Enr or Std environment for 12 d (2 h/d) with light stimulation (1 Hz, 5 ms in duration). B , Left, Representative images of the GCL of the DG from mice in the GFP and Halo groups under Std and Enr environments, respectively. Scale bar, 30 μm. Right, Plots of BrdU+ cells in the GCL of mice in the GFP-Enr group compared with those in the GFP-Std group (relative fold change in Brdu: GFP-Std: 1.00 ± 0.17, GFP-Enr: 1.90 ± 0.32, 2-tailed unpaired t test p = 0.025),and BrdU+ cells in the GCL of mice in the Halo-Enr compared with those in the Halo-Std group (Halo-Std: 1.00 ± 0.20, Halo-Enr: 1.07 ± 0.20, 2-tailed unpaired t test p = 0.818). C , Left, Representative images of c-fos staining in the GCL of the DG of AAV-CaMKII-GFP and AAV-CaMKII-Halo injected mice exposed to an Enr or Std environment. Right, Plots of c-Fos+ cells in the GCL of mice in the GFP-Enr condition and GFP-Std condition (relative fold change c-Fos+ cells/field: GFP-Std: 1.00 ± 0.19, GFP-Enr: 3.70 ± 0.46; 2-tailed unpaired t test p

Techniques Used: Mouse Assay, Injection, Mass Spectrometry, Staining

16) Product Images from "A Unique Role of GATA1s in Down Syndrome Acute Megakaryocytic Leukemia Biology and Therapy"

Article Title: A Unique Role of GATA1s in Down Syndrome Acute Megakaryocytic Leukemia Biology and Therapy

Journal: PLoS ONE

doi: 10.1371/journal.pone.0027486

Down-regulation of GATA1s in CMK cells results in impaired cell proliferation. Expression of GATA1s in two selected subclones, CMK-5a and -5b, in comparison to the negative control (CMK-neg) was verified by Western blotting ( panel A ) and real-time RT-PCR ( panel B ). The real-time RT-PCR results were expressed as mean values ± standard errors from 3 independent experiments using the same cDNA preparation and normalized to GAPDH. To establish the doubling times for each shRNA subclone, the CMK-5a, -5b, and –neg sublines were seeded at 2.5×10 4 cells/ml and counted every 24 h with trypan blue staining ( panel C ). Cell cycle progression in the CMK-5a, -5b, and –neg sublines was assessed by PI staining and flow cytometry analysis, as described in the Materials and Methods ( panel D ). DNA content was also assessed in the CMK-5a, -5b, and –neg sublines by incorporating BrdU into DNA and flow cytometry analysis as described in the Materials and Methods ( panel E ).
Figure Legend Snippet: Down-regulation of GATA1s in CMK cells results in impaired cell proliferation. Expression of GATA1s in two selected subclones, CMK-5a and -5b, in comparison to the negative control (CMK-neg) was verified by Western blotting ( panel A ) and real-time RT-PCR ( panel B ). The real-time RT-PCR results were expressed as mean values ± standard errors from 3 independent experiments using the same cDNA preparation and normalized to GAPDH. To establish the doubling times for each shRNA subclone, the CMK-5a, -5b, and –neg sublines were seeded at 2.5×10 4 cells/ml and counted every 24 h with trypan blue staining ( panel C ). Cell cycle progression in the CMK-5a, -5b, and –neg sublines was assessed by PI staining and flow cytometry analysis, as described in the Materials and Methods ( panel D ). DNA content was also assessed in the CMK-5a, -5b, and –neg sublines by incorporating BrdU into DNA and flow cytometry analysis as described in the Materials and Methods ( panel E ).

Techniques Used: Expressing, Negative Control, Western Blot, Quantitative RT-PCR, shRNA, Staining, Flow Cytometry, Cytometry

17) Product Images from "Mitochondrial degeneration and not apoptosis is the primary cause of embryonic lethality in ceramide transfer protein mutant mice"

Article Title: Mitochondrial degeneration and not apoptosis is the primary cause of embryonic lethality in ceramide transfer protein mutant mice

Journal: The Journal of Cell Biology

doi: 10.1083/jcb.200807176

Growth and cell cycle defects without increased apoptosis in Cert gt/gt mutant embryos. (A) Pregnant mice were injected with BrdU 1 h before dissection, and E10.5 embryos were dissected, genotyped, and stained for BrdU uptake as described in Materials and methods. Four of the areas chosen for counting the fraction of dividing cells are shown in the panels to the right. (B) Cert gt/gt embryos showed a lower fraction of BrdU-positive cells, indicating a slower rate of cell division in these mutant embryos. Error bars indicate standard deviation; n = 4. (C) Western blot analysis for some of the components of the cell cycle in Cert +/+ (+/+) and Cert gt/gt (−/−) embryos. (D) Phosphorylated Akt levels are decreased in Cert gt/gt embryos. (E) TUNEL staining of E10.5 wild-type and mutant embryos. Insets show boxed areas at higher magnification. There were no significant differences in the proportion of dying cells between the wild-type and mutant embryos. Note that the wild-type embryos are large compared with the mutant and, therefore, were photographed twice to include the upper and the lower halves of the embryo. The composites are shown in A and E.
Figure Legend Snippet: Growth and cell cycle defects without increased apoptosis in Cert gt/gt mutant embryos. (A) Pregnant mice were injected with BrdU 1 h before dissection, and E10.5 embryos were dissected, genotyped, and stained for BrdU uptake as described in Materials and methods. Four of the areas chosen for counting the fraction of dividing cells are shown in the panels to the right. (B) Cert gt/gt embryos showed a lower fraction of BrdU-positive cells, indicating a slower rate of cell division in these mutant embryos. Error bars indicate standard deviation; n = 4. (C) Western blot analysis for some of the components of the cell cycle in Cert +/+ (+/+) and Cert gt/gt (−/−) embryos. (D) Phosphorylated Akt levels are decreased in Cert gt/gt embryos. (E) TUNEL staining of E10.5 wild-type and mutant embryos. Insets show boxed areas at higher magnification. There were no significant differences in the proportion of dying cells between the wild-type and mutant embryos. Note that the wild-type embryos are large compared with the mutant and, therefore, were photographed twice to include the upper and the lower halves of the embryo. The composites are shown in A and E.

Techniques Used: Mutagenesis, Mouse Assay, Injection, Dissection, Staining, Standard Deviation, Western Blot, TUNEL Assay

18) Product Images from "Mitochondrial DNA damage and subsequent activation of Z-DNA binding protein 1 links oxidative stress to inflammation in epithelial cells"

Article Title: Mitochondrial DNA damage and subsequent activation of Z-DNA binding protein 1 links oxidative stress to inflammation in epithelial cells

Journal: Scientific Reports

doi: 10.1038/s41598-018-19216-1

Damaged mitochondrial DNA activates ZBP1/TBK1/IRF3 signaling pathway. Interaction between BrDU-labelled mtDNA and TFAM ( A ) and BrDU-labelled mtDNA and ZBP1 ( B ) in control and GOx-treated cells at 1 h. ( C ) Interaction between ZBP1/TBK1, IRF3/TBK1, ZBP1/P-Tyr and ZBP1/P-Ser in control and GOx-treated cells at 1 h. ( D ) Interaction between IRF3/TBK1 in pre-treated with 3 μM of CsA in GOx-treated BEAS 2B cells. ( E ) Level of ZBP1 depletion, shown by Western blotting. ( F ) Expression of IL-6 and IL-8 in unstressed and GOx-treated (0.006 U/ml for 1 h) in control and ZBP1-depleted BEAS2B cells. Representative images of n = 3 independent experiments are shown. Data represent average ± SEM of n = 5 biological replicates. *p
Figure Legend Snippet: Damaged mitochondrial DNA activates ZBP1/TBK1/IRF3 signaling pathway. Interaction between BrDU-labelled mtDNA and TFAM ( A ) and BrDU-labelled mtDNA and ZBP1 ( B ) in control and GOx-treated cells at 1 h. ( C ) Interaction between ZBP1/TBK1, IRF3/TBK1, ZBP1/P-Tyr and ZBP1/P-Ser in control and GOx-treated cells at 1 h. ( D ) Interaction between IRF3/TBK1 in pre-treated with 3 μM of CsA in GOx-treated BEAS 2B cells. ( E ) Level of ZBP1 depletion, shown by Western blotting. ( F ) Expression of IL-6 and IL-8 in unstressed and GOx-treated (0.006 U/ml for 1 h) in control and ZBP1-depleted BEAS2B cells. Representative images of n = 3 independent experiments are shown. Data represent average ± SEM of n = 5 biological replicates. *p

Techniques Used: Western Blot, Expressing

19) Product Images from "The aPKC-CBP Pathway Regulates Adult Hippocampal Neurogenesis in an Age-Dependent Manner"

Article Title: The aPKC-CBP Pathway Regulates Adult Hippocampal Neurogenesis in an Age-Dependent Manner

Journal: Stem Cell Reports

doi: 10.1016/j.stemcr.2016.08.007

Mature Adult Mice Lacking CBPS436 Phosphorylation Show Reduced Hippocampal Neuronal Differentiation Adult mice (3 and 6 months old) received BrdU (100 mg/kg) injections for 3 consecutive days, and were euthanized for immunostaining analysis 12 days after the first BrdU injection. (A, C, E, and G) Fluorescence images of hippocampal sections from 6-month-old (A,C,E) and 3-month-old (G) WT and Cbp S436A-KI mice, stained for: (A) BrdU (green) and NEUN (red); (C) BrdU (green) and SOX2 (red); (E) BrdU (green), TBR2 (purple), and SOX2 (red); and (G) BrdU (green) and DCX (red). Arrows denote double-labeled cells. The boxed areas are shown at higher magnification on the right. Scale bars, 25 μm. (B) Quantitative analysis of the percentage of BrdU-positive cells that express a mature neuron marker, NEUN, in the hippocampi from 3- to 6-month-old WT and Cbp S436A-KI mice. ∗∗ p
Figure Legend Snippet: Mature Adult Mice Lacking CBPS436 Phosphorylation Show Reduced Hippocampal Neuronal Differentiation Adult mice (3 and 6 months old) received BrdU (100 mg/kg) injections for 3 consecutive days, and were euthanized for immunostaining analysis 12 days after the first BrdU injection. (A, C, E, and G) Fluorescence images of hippocampal sections from 6-month-old (A,C,E) and 3-month-old (G) WT and Cbp S436A-KI mice, stained for: (A) BrdU (green) and NEUN (red); (C) BrdU (green) and SOX2 (red); (E) BrdU (green), TBR2 (purple), and SOX2 (red); and (G) BrdU (green) and DCX (red). Arrows denote double-labeled cells. The boxed areas are shown at higher magnification on the right. Scale bars, 25 μm. (B) Quantitative analysis of the percentage of BrdU-positive cells that express a mature neuron marker, NEUN, in the hippocampi from 3- to 6-month-old WT and Cbp S436A-KI mice. ∗∗ p

Techniques Used: Mouse Assay, Immunostaining, Injection, Fluorescence, Staining, Labeling, Marker

Mature Adult Mice Lacking CBPS436 Phosphorylation Show Impaired Hippocampal Neuronal Maturation Adult mice (6 months old) received BrdU (100 mg/kg) injections for 3 consecutive days, and were euthanized 12 days after the first BrdU injection. (A) Confocal Images of coronal hippocampal dentate gyrus sections from 6-month-old WT and Cbp S436A-KI mice, stained for BrdU (green), DCX (red), and NEUN (purple). Arrows denote triple-labeled BrdU + /DCX + /NEUN + cells; arrowheads denote double-labeled BrdU + /DCX + /NEUN − cells; asterisks denote single-labeled BrdU + /DCX − /NEUN − cells. Scale bars, 20 μm. (B–F) Quantitative analysis of the percentage of BrdU/DCX-positive cells that also express NEUN (B); the percentage of BrdU-positive cells that were positive for both NEUN and DCX (C); the percentage of BrdU-positive cells that were positive for NEUN but negative for DCX (D); the percentage of BrdU-positive cells that express DCX but not NEUN (E); and the percentage of BrdU-positive cells that were negative for both NEUN and DCX (F) in the hippocampi from 3- to 6-month-old WT and Cbp S436A-KI mice. ∗ p
Figure Legend Snippet: Mature Adult Mice Lacking CBPS436 Phosphorylation Show Impaired Hippocampal Neuronal Maturation Adult mice (6 months old) received BrdU (100 mg/kg) injections for 3 consecutive days, and were euthanized 12 days after the first BrdU injection. (A) Confocal Images of coronal hippocampal dentate gyrus sections from 6-month-old WT and Cbp S436A-KI mice, stained for BrdU (green), DCX (red), and NEUN (purple). Arrows denote triple-labeled BrdU + /DCX + /NEUN + cells; arrowheads denote double-labeled BrdU + /DCX + /NEUN − cells; asterisks denote single-labeled BrdU + /DCX − /NEUN − cells. Scale bars, 20 μm. (B–F) Quantitative analysis of the percentage of BrdU/DCX-positive cells that also express NEUN (B); the percentage of BrdU-positive cells that were positive for both NEUN and DCX (C); the percentage of BrdU-positive cells that were positive for NEUN but negative for DCX (D); the percentage of BrdU-positive cells that express DCX but not NEUN (E); and the percentage of BrdU-positive cells that were negative for both NEUN and DCX (F) in the hippocampi from 3- to 6-month-old WT and Cbp S436A-KI mice. ∗ p

Techniques Used: Mouse Assay, Injection, Staining, Labeling

Adult Mice Lacking CBPS436 Phosphorylation Show a Reduction in Adult Neurogenesis (A) Fluorescence images of hippocampal sections from 3-month-old Cbp S436A ( Cbp S436A-KI) and their wild-type littermates (WT), euthanized 12 days after BrdU injections and stained for BrdU (green) and NEUN (red). The insets show the boxed areas at higher magnification. Arrows represent double-labeled BrdU/NEUN-positive neurons. Scale bar, 100 μm. (B) Quantitative analysis of the total number of BrdU/NEUN-positive newborn neurons in the hippocampi from 3- to 6-month-old WT and Cbp S436A-KI mice. ∗ p
Figure Legend Snippet: Adult Mice Lacking CBPS436 Phosphorylation Show a Reduction in Adult Neurogenesis (A) Fluorescence images of hippocampal sections from 3-month-old Cbp S436A ( Cbp S436A-KI) and their wild-type littermates (WT), euthanized 12 days after BrdU injections and stained for BrdU (green) and NEUN (red). The insets show the boxed areas at higher magnification. Arrows represent double-labeled BrdU/NEUN-positive neurons. Scale bar, 100 μm. (B) Quantitative analysis of the total number of BrdU/NEUN-positive newborn neurons in the hippocampi from 3- to 6-month-old WT and Cbp S436A-KI mice. ∗ p

Techniques Used: Mouse Assay, Fluorescence, Staining, Labeling

20) Product Images from "Ilexonin A Promotes Neuronal Proliferation and Regeneration via Activation of the Canonical Wnt Signaling Pathway after Cerebral Ischemia Reperfusion in Rats"

Article Title: Ilexonin A Promotes Neuronal Proliferation and Regeneration via Activation of the Canonical Wnt Signaling Pathway after Cerebral Ischemia Reperfusion in Rats

Journal: Evidence-based Complementary and Alternative Medicine : eCAM

doi: 10.1155/2016/9753189

IA treatment group's corresponding trend of Brdu/NeuN double positive cell and β -catenin positive cell at corresponding time point after ischemia reperfusion in rats.
Figure Legend Snippet: IA treatment group's corresponding trend of Brdu/NeuN double positive cell and β -catenin positive cell at corresponding time point after ischemia reperfusion in rats.

Techniques Used: IA

21) Product Images from "Early Life Stress Enhancement of Limbic Epileptogenesis in Adult Rats: Mechanistic Insights"

Article Title: Early Life Stress Enhancement of Limbic Epileptogenesis in Adult Rats: Mechanistic Insights

Journal: PLoS ONE

doi: 10.1371/journal.pone.0024033

BrdU-labelled cells predominantly adopt a neuronal fate. Images show representative images from a MS-exposed female kindled rat brain (left panels) and an EH-exposed kindled female rat brain (right panels). Panels (i) and (ii) display NeuN+ve cells in green, indicative of mature neurons. Panels (iii) and (iv) display BrdU+ve cells in red, indicative of newly synthesised cells. Panels (v) and (vi) represent GFAP+ve glial cells in blue. Panels (vii) and (viii) represent the merged images of the three panels above, and panels (ix) and (x) are enlarged sections of the boxed areas in panels (vii) and (viii), respectively. Note the pink arrows indicating cells yellow cells in panels (vii) – (x), the product of double-labelling with NeuN (green) and BrdU (red), and indicative of newly generated neurons.
Figure Legend Snippet: BrdU-labelled cells predominantly adopt a neuronal fate. Images show representative images from a MS-exposed female kindled rat brain (left panels) and an EH-exposed kindled female rat brain (right panels). Panels (i) and (ii) display NeuN+ve cells in green, indicative of mature neurons. Panels (iii) and (iv) display BrdU+ve cells in red, indicative of newly synthesised cells. Panels (v) and (vi) represent GFAP+ve glial cells in blue. Panels (vii) and (viii) represent the merged images of the three panels above, and panels (ix) and (x) are enlarged sections of the boxed areas in panels (vii) and (viii), respectively. Note the pink arrows indicating cells yellow cells in panels (vii) – (x), the product of double-labelling with NeuN (green) and BrdU (red), and indicative of newly generated neurons.

Techniques Used: Mass Spectrometry, Generated

Phenotypes of BrdU positive cells labelled with NeuN, GFAP or neither in kindled MS and EH rats (p
Figure Legend Snippet: Phenotypes of BrdU positive cells labelled with NeuN, GFAP or neither in kindled MS and EH rats (p

Techniques Used: Mass Spectrometry

22) Product Images from "Tracking of quiescence in Leishmania by quantifying the expression of GFP in the ribosomal DNA locus"

Article Title: Tracking of quiescence in Leishmania by quantifying the expression of GFP in the ribosomal DNA locus

Journal: Scientific Reports

doi: 10.1038/s41598-019-55486-z

Relationship between BrdU incorporation (as a measure of replication) and rGFP expression in promastigotes and amastigotes of L. braziliensis . ( A) Confocal microscopy pictures of the immuno-detection of GFP and BrdU, 8 hrs after exposure of the cells to BrdU. The images for Pro Log , Pro Sta and Ama Axe were taken with the same microscopical settings to allow a comparison between the samples. The images for Ama Int were taken with increased settings of the Gain Master to have a higher sensitivity for the detection of GFP. The new settings became the rGFP expression detectable in a small proportion of them. The scale bar in each picture represents 5 µM. ( B) Quantification by flow cytometry of the percentage of cells in a proliferative stage as evidenced by the incorporation of BrdU. The bars and the dots represent the mean ± SEM of three biological replicates. ( C) Density plot of rGFP expression (Y-axis) and level of BrdU incorporation (X-axis), by flow cytometry in each stage of the parasite over 24 hrs of incubation with BrdU. Each plot in the left top corner is labelled with an ID. Moreover, each plot is divided in four quadrants: UL, UR, LL and LR. The proliferative cells (BrdU + ) are within the quadrants UR if they are also GFP + and in the quadrant LR if they are GFP − . ( D ) Quantification of rGFP expression in proliferative (BrdU + ) and non-proliferative (BrdU − ) populations of each stage, after 8 hrs of exposure to BrdU. The bars represent the mean ± SEM of three biological replicates.
Figure Legend Snippet: Relationship between BrdU incorporation (as a measure of replication) and rGFP expression in promastigotes and amastigotes of L. braziliensis . ( A) Confocal microscopy pictures of the immuno-detection of GFP and BrdU, 8 hrs after exposure of the cells to BrdU. The images for Pro Log , Pro Sta and Ama Axe were taken with the same microscopical settings to allow a comparison between the samples. The images for Ama Int were taken with increased settings of the Gain Master to have a higher sensitivity for the detection of GFP. The new settings became the rGFP expression detectable in a small proportion of them. The scale bar in each picture represents 5 µM. ( B) Quantification by flow cytometry of the percentage of cells in a proliferative stage as evidenced by the incorporation of BrdU. The bars and the dots represent the mean ± SEM of three biological replicates. ( C) Density plot of rGFP expression (Y-axis) and level of BrdU incorporation (X-axis), by flow cytometry in each stage of the parasite over 24 hrs of incubation with BrdU. Each plot in the left top corner is labelled with an ID. Moreover, each plot is divided in four quadrants: UL, UR, LL and LR. The proliferative cells (BrdU + ) are within the quadrants UR if they are also GFP + and in the quadrant LR if they are GFP − . ( D ) Quantification of rGFP expression in proliferative (BrdU + ) and non-proliferative (BrdU − ) populations of each stage, after 8 hrs of exposure to BrdU. The bars represent the mean ± SEM of three biological replicates.

Techniques Used: BrdU Incorporation Assay, Expressing, Confocal Microscopy, Flow Cytometry, Cytometry, Incubation

Relationship between BrdU incorporation (as a measure of replication) and rGFP expression in promastigotes and amastigotes of L. braziliensis . ( A) Confocal microscopy pictures of the immuno-detection of GFP and BrdU, 8 hrs after exposure of the cells to BrdU. The images for Pro Log , Pro Sta and Ama Axe were taken with the same microscopical settings to allow a comparison between the samples. The images for Ama Int were taken with increased settings of the Gain Master to have a higher sensitivity for the detection of GFP. The new settings became the rGFP expression detectable in a small proportion of them. The scale bar in each picture represents 5 µM. ( B) Quantification by flow cytometry of the percentage of cells in a proliferative stage as evidenced by the incorporation of BrdU. The bars and the dots represent the mean ± SEM of three biological replicates. ( C) Density plot of rGFP expression (Y-axis) and level of BrdU incorporation (X-axis), by flow cytometry in each stage of the parasite over 24 hrs of incubation with BrdU. Each plot in the left top corner is labelled with an ID. Moreover, each plot is divided in four quadrants: UL, UR, LL and LR. The proliferative cells (BrdU + ) are within the quadrants UR if they are also GFP + and in the quadrant LR if they are GFP − . ( D ) Quantification of rGFP expression in proliferative (BrdU + ) and non-proliferative (BrdU − ) populations of each stage, after 8 hrs of exposure to BrdU. The bars represent the mean ± SEM of three biological replicates.
Figure Legend Snippet: Relationship between BrdU incorporation (as a measure of replication) and rGFP expression in promastigotes and amastigotes of L. braziliensis . ( A) Confocal microscopy pictures of the immuno-detection of GFP and BrdU, 8 hrs after exposure of the cells to BrdU. The images for Pro Log , Pro Sta and Ama Axe were taken with the same microscopical settings to allow a comparison between the samples. The images for Ama Int were taken with increased settings of the Gain Master to have a higher sensitivity for the detection of GFP. The new settings became the rGFP expression detectable in a small proportion of them. The scale bar in each picture represents 5 µM. ( B) Quantification by flow cytometry of the percentage of cells in a proliferative stage as evidenced by the incorporation of BrdU. The bars and the dots represent the mean ± SEM of three biological replicates. ( C) Density plot of rGFP expression (Y-axis) and level of BrdU incorporation (X-axis), by flow cytometry in each stage of the parasite over 24 hrs of incubation with BrdU. Each plot in the left top corner is labelled with an ID. Moreover, each plot is divided in four quadrants: UL, UR, LL and LR. The proliferative cells (BrdU + ) are within the quadrants UR if they are also GFP + and in the quadrant LR if they are GFP − . ( D ) Quantification of rGFP expression in proliferative (BrdU + ) and non-proliferative (BrdU − ) populations of each stage, after 8 hrs of exposure to BrdU. The bars represent the mean ± SEM of three biological replicates.

Techniques Used: BrdU Incorporation Assay, Expressing, Confocal Microscopy, Flow Cytometry, Cytometry, Incubation

23) Product Images from "Necdin Controls Proliferation of White Adipocyte Progenitor Cells"

Article Title: Necdin Controls Proliferation of White Adipocyte Progenitor Cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0030948

Necdin deficiency enhances preadipocyte proliferation in adipose SV cells. ( A ) Co-immunostaining for necdin and PPARγ. SV cells were treated with (+) or without (−) adipogenic inducers, and fixed 72 hr after induction. Cells were double-immunostained for necdin (red) and PPARγ (green), and confocal laser microscopic images are merged (Merge). Arrowheads point to the nucleus. ( B ) Co-staining for BrdU and PPARγ. SV cells were treated with adipogenic inducers, pulse-labeled with BrdU, triple-stained for BrdU, PPARγ and nuclear DNA with Hoechst 33342, and observed by fluorescence microscopy. Arrowheads (Merge) point to BrdU + PPARγ + cells. ( C, D ) Flow cytometry for BrdU incorporation. SV cells were prepared from Ndn +/+ and Ndn +m/−p mice, treated with adipogenic inducers, and analyzed by flow cytometry for BrdU incorporation. BrdU + cells in the colored area (blue for Ndn +/+ , red for Ndn +m/−p )( C ) were counted ( D ). The threshold (broken line) was set using negative control cells without BrdU treatment (Ctl). ( E, F ) Flow cytometry for apoptosis. SV cells were treated with adipogenic inducers and analyzed 20 hr later by flow cytometry using FITC-labeled Annexin V. * P
Figure Legend Snippet: Necdin deficiency enhances preadipocyte proliferation in adipose SV cells. ( A ) Co-immunostaining for necdin and PPARγ. SV cells were treated with (+) or without (−) adipogenic inducers, and fixed 72 hr after induction. Cells were double-immunostained for necdin (red) and PPARγ (green), and confocal laser microscopic images are merged (Merge). Arrowheads point to the nucleus. ( B ) Co-staining for BrdU and PPARγ. SV cells were treated with adipogenic inducers, pulse-labeled with BrdU, triple-stained for BrdU, PPARγ and nuclear DNA with Hoechst 33342, and observed by fluorescence microscopy. Arrowheads (Merge) point to BrdU + PPARγ + cells. ( C, D ) Flow cytometry for BrdU incorporation. SV cells were prepared from Ndn +/+ and Ndn +m/−p mice, treated with adipogenic inducers, and analyzed by flow cytometry for BrdU incorporation. BrdU + cells in the colored area (blue for Ndn +/+ , red for Ndn +m/−p )( C ) were counted ( D ). The threshold (broken line) was set using negative control cells without BrdU treatment (Ctl). ( E, F ) Flow cytometry for apoptosis. SV cells were treated with adipogenic inducers and analyzed 20 hr later by flow cytometry using FITC-labeled Annexin V. * P

Techniques Used: Immunostaining, Staining, Labeling, Fluorescence, Microscopy, Flow Cytometry, Cytometry, BrdU Incorporation Assay, Mouse Assay, Negative Control, CTL Assay

Necdin deficiency enhances preadipocyte proliferation in adipose SV cells. ( A ) Co-immunostaining for necdin and PPARγ. SV cells were treated with (+) or without (−) adipogenic inducers, and fixed 72 hr after induction. Cells were double-immunostained for necdin (red) and PPARγ (green), and confocal laser microscopic images are merged (Merge). Arrowheads point to the nucleus. ( B ) Co-staining for BrdU and PPARγ. SV cells were treated with adipogenic inducers, pulse-labeled with BrdU, triple-stained for BrdU, PPARγ and nuclear DNA with Hoechst 33342, and observed by fluorescence microscopy. Arrowheads (Merge) point to BrdU + PPARγ + cells. ( C, D ) Flow cytometry for BrdU incorporation. SV cells were prepared from Ndn +/+ and Ndn +m/−p mice, treated with adipogenic inducers, and analyzed by flow cytometry for BrdU incorporation. BrdU + cells in the colored area (blue for Ndn +/+ , red for Ndn +m/−p )( C ) were counted ( D ). The threshold (broken line) was set using negative control cells without BrdU treatment (Ctl). ( E, F ) Flow cytometry for apoptosis. SV cells were treated with adipogenic inducers and analyzed 20 hr later by flow cytometry using FITC-labeled Annexin V. * P
Figure Legend Snippet: Necdin deficiency enhances preadipocyte proliferation in adipose SV cells. ( A ) Co-immunostaining for necdin and PPARγ. SV cells were treated with (+) or without (−) adipogenic inducers, and fixed 72 hr after induction. Cells were double-immunostained for necdin (red) and PPARγ (green), and confocal laser microscopic images are merged (Merge). Arrowheads point to the nucleus. ( B ) Co-staining for BrdU and PPARγ. SV cells were treated with adipogenic inducers, pulse-labeled with BrdU, triple-stained for BrdU, PPARγ and nuclear DNA with Hoechst 33342, and observed by fluorescence microscopy. Arrowheads (Merge) point to BrdU + PPARγ + cells. ( C, D ) Flow cytometry for BrdU incorporation. SV cells were prepared from Ndn +/+ and Ndn +m/−p mice, treated with adipogenic inducers, and analyzed by flow cytometry for BrdU incorporation. BrdU + cells in the colored area (blue for Ndn +/+ , red for Ndn +m/−p )( C ) were counted ( D ). The threshold (broken line) was set using negative control cells without BrdU treatment (Ctl). ( E, F ) Flow cytometry for apoptosis. SV cells were treated with adipogenic inducers and analyzed 20 hr later by flow cytometry using FITC-labeled Annexin V. * P

Techniques Used: Immunostaining, Staining, Labeling, Fluorescence, Microscopy, Flow Cytometry, Cytometry, BrdU Incorporation Assay, Mouse Assay, Negative Control, CTL Assay

24) Product Images from "Induction of Functional Hair-Cell-Like Cells from Mouse Cochlear Multipotent Cells"

Article Title: Induction of Functional Hair-Cell-Like Cells from Mouse Cochlear Multipotent Cells

Journal: Stem Cells International

doi: 10.1155/2016/8197279

Characterization of spheres from the cochlear epithelia of newborn mice. (a) The cochlear sensory epithelia of P0 ICR mice were dissociated into single cells and cultured in suspension plates for 5 days to form spheres in presence of EGF, bFGF, and IGF-1, as indicated. (b) A bright-field view of a typical sphere generated after 5 days of in vitro culture. (c) Sphere cells expressed the neuronal stem cell marker Nestin (green). (d) Shortly after dissociation, most sphere cells were immunoreactive for 5-bromo-2′-deoxyuridine (BrdU) incorporation into the genomic DNA (red, left). (e) The nuclei of the sphere cells were stained with DAPI (blue). (f) The merging of BrdU immunostaining (red) and DAPI staining (blue).
Figure Legend Snippet: Characterization of spheres from the cochlear epithelia of newborn mice. (a) The cochlear sensory epithelia of P0 ICR mice were dissociated into single cells and cultured in suspension plates for 5 days to form spheres in presence of EGF, bFGF, and IGF-1, as indicated. (b) A bright-field view of a typical sphere generated after 5 days of in vitro culture. (c) Sphere cells expressed the neuronal stem cell marker Nestin (green). (d) Shortly after dissociation, most sphere cells were immunoreactive for 5-bromo-2′-deoxyuridine (BrdU) incorporation into the genomic DNA (red, left). (e) The nuclei of the sphere cells were stained with DAPI (blue). (f) The merging of BrdU immunostaining (red) and DAPI staining (blue).

Techniques Used: Mouse Assay, Cell Culture, Generated, In Vitro, Marker, BrdU Incorporation Assay, Staining, Immunostaining

25) Product Images from "Bipotent progenitors as embryonic origin of retinal stem cells"

Article Title: Bipotent progenitors as embryonic origin of retinal stem cells

Journal: The Journal of Cell Biology

doi: 10.1083/jcb.201611057

Blood vessels are required for actively proliferative RSCs. (A) Schematic of the design of BAC transgene kdrl:mCherrycaax . The initiation codon ATG was used. (B) 3D illustration (frontal and lateral views) of blood vessels (in magenta) in the eye of the transgenic fish line Tg( rx2:GFPcaax::kdrl:mCherrycaax ). SAVs and AHVs surround the CMZ tip region. (C) A high-resolution image of a four-cell cluster of the CMZ tip region that is associated with the local blood vessels. (D) Quantification, measured by the physical cell positions of the CMZ tip region, showing that the second to fourth cells ( n = 11) often directly associated with the local blood vessel (AHV), whereas dormant CMZ tip cells did not ( n = 12). (E) 3D representation of the blood vessels in control eyes and eyes treated with 50 nM Ki8751 at 36, 48, and 72 hpf. (F) Representative images of the CMZ labeled in the Tg( rx2:GFPcaax ) with the treatment of 50 nM Ki8751 at 5 and 8 dpf showing the reduction of the CMZ. Insets represent the enlarged CMZ. (G) Quantifications of eye diameters in embryos with or without Ki8751 treatment show a significant reduction of the eye size in the treated embryos at 72 hpf onwards. The p-values in groups of 48 hpf, 72 hpf, and 5 dpf were 0.124, 1.42 × 10 −6 , and 3.1 × 10 −5 , respectively. (H–J) BrdU and pH3 staining of the retina in embryos with or without Ki8751 treatment shows a significant reduction of cell proliferation in the CMZ of the retina in the treated embryos at 5 dpf. The p-values in groups of BrdU and pH3 were 0.002 and 0.001, respectively. Students’ t test: **, P
Figure Legend Snippet: Blood vessels are required for actively proliferative RSCs. (A) Schematic of the design of BAC transgene kdrl:mCherrycaax . The initiation codon ATG was used. (B) 3D illustration (frontal and lateral views) of blood vessels (in magenta) in the eye of the transgenic fish line Tg( rx2:GFPcaax::kdrl:mCherrycaax ). SAVs and AHVs surround the CMZ tip region. (C) A high-resolution image of a four-cell cluster of the CMZ tip region that is associated with the local blood vessels. (D) Quantification, measured by the physical cell positions of the CMZ tip region, showing that the second to fourth cells ( n = 11) often directly associated with the local blood vessel (AHV), whereas dormant CMZ tip cells did not ( n = 12). (E) 3D representation of the blood vessels in control eyes and eyes treated with 50 nM Ki8751 at 36, 48, and 72 hpf. (F) Representative images of the CMZ labeled in the Tg( rx2:GFPcaax ) with the treatment of 50 nM Ki8751 at 5 and 8 dpf showing the reduction of the CMZ. Insets represent the enlarged CMZ. (G) Quantifications of eye diameters in embryos with or without Ki8751 treatment show a significant reduction of the eye size in the treated embryos at 72 hpf onwards. The p-values in groups of 48 hpf, 72 hpf, and 5 dpf were 0.124, 1.42 × 10 −6 , and 3.1 × 10 −5 , respectively. (H–J) BrdU and pH3 staining of the retina in embryos with or without Ki8751 treatment shows a significant reduction of cell proliferation in the CMZ of the retina in the treated embryos at 5 dpf. The p-values in groups of BrdU and pH3 were 0.002 and 0.001, respectively. Students’ t test: **, P

Techniques Used: BAC Assay, Transgenic Assay, Fluorescence In Situ Hybridization, Labeling, Staining

Blood vessels are required for actively proliferative RSCs. (A) Schematic of the design of BAC transgene kdrl:mCherrycaax . The initiation codon ATG was used. (B) 3D illustration (frontal and lateral views) of blood vessels (in magenta) in the eye of the transgenic fish line Tg( rx2:GFPcaax::kdrl:mCherrycaax ). SAVs and AHVs surround the CMZ tip region. (C) A high-resolution image of a four-cell cluster of the CMZ tip region that is associated with the local blood vessels. (D) Quantification, measured by the physical cell positions of the CMZ tip region, showing that the second to fourth cells ( n = 11) often directly associated with the local blood vessel (AHV), whereas dormant CMZ tip cells did not ( n = 12). (E) 3D representation of the blood vessels in control eyes and eyes treated with 50 nM Ki8751 at 36, 48, and 72 hpf. (F) Representative images of the CMZ labeled in the Tg( rx2:GFPcaax ) with the treatment of 50 nM Ki8751 at 5 and 8 dpf showing the reduction of the CMZ. Insets represent the enlarged CMZ. (G) Quantifications of eye diameters in embryos with or without Ki8751 treatment show a significant reduction of the eye size in the treated embryos at 72 hpf onwards. The p-values in groups of 48 hpf, 72 hpf, and 5 dpf were 0.124, 1.42 × 10 −6 , and 3.1 × 10 −5 , respectively. (H–J) BrdU and pH3 staining of the retina in embryos with or without Ki8751 treatment shows a significant reduction of cell proliferation in the CMZ of the retina in the treated embryos at 5 dpf. The p-values in groups of BrdU and pH3 were 0.002 and 0.001, respectively. Students’ t test: **, P
Figure Legend Snippet: Blood vessels are required for actively proliferative RSCs. (A) Schematic of the design of BAC transgene kdrl:mCherrycaax . The initiation codon ATG was used. (B) 3D illustration (frontal and lateral views) of blood vessels (in magenta) in the eye of the transgenic fish line Tg( rx2:GFPcaax::kdrl:mCherrycaax ). SAVs and AHVs surround the CMZ tip region. (C) A high-resolution image of a four-cell cluster of the CMZ tip region that is associated with the local blood vessels. (D) Quantification, measured by the physical cell positions of the CMZ tip region, showing that the second to fourth cells ( n = 11) often directly associated with the local blood vessel (AHV), whereas dormant CMZ tip cells did not ( n = 12). (E) 3D representation of the blood vessels in control eyes and eyes treated with 50 nM Ki8751 at 36, 48, and 72 hpf. (F) Representative images of the CMZ labeled in the Tg( rx2:GFPcaax ) with the treatment of 50 nM Ki8751 at 5 and 8 dpf showing the reduction of the CMZ. Insets represent the enlarged CMZ. (G) Quantifications of eye diameters in embryos with or without Ki8751 treatment show a significant reduction of the eye size in the treated embryos at 72 hpf onwards. The p-values in groups of 48 hpf, 72 hpf, and 5 dpf were 0.124, 1.42 × 10 −6 , and 3.1 × 10 −5 , respectively. (H–J) BrdU and pH3 staining of the retina in embryos with or without Ki8751 treatment shows a significant reduction of cell proliferation in the CMZ of the retina in the treated embryos at 5 dpf. The p-values in groups of BrdU and pH3 were 0.002 and 0.001, respectively. Students’ t test: **, P

Techniques Used: BAC Assay, Transgenic Assay, Fluorescence In Situ Hybridization, Labeling, Staining

26) Product Images from "Lysophosphatidic Acid Receptor Is a Functional Marker of Adult Hippocampal Precursor Cells"

Article Title: Lysophosphatidic Acid Receptor Is a Functional Marker of Adult Hippocampal Precursor Cells

Journal: Stem Cell Reports

doi: 10.1016/j.stemcr.2016.03.002

A Running-Induced Increase in Proliferation Can Be Measured Directly in LPA 1 -GFP Mice (A and B) Experimental design (A) and histogram (B) representing the number of BrdU-labeled cells in running (RUN) and standardly housed (STD) LPA 1 -GFP mice. n = 11 mice per group, ∗∗∗ p
Figure Legend Snippet: A Running-Induced Increase in Proliferation Can Be Measured Directly in LPA 1 -GFP Mice (A and B) Experimental design (A) and histogram (B) representing the number of BrdU-labeled cells in running (RUN) and standardly housed (STD) LPA 1 -GFP mice. n = 11 mice per group, ∗∗∗ p

Techniques Used: Mouse Assay, Labeling

27) Product Images from "Malignant Catarrhal Fever Induced by Alcelaphine herpesvirus 1 Is Associated with Proliferation of CD8+ T Cells Supporting a Latent Infection"

Article Title: Malignant Catarrhal Fever Induced by Alcelaphine herpesvirus 1 Is Associated with Proliferation of CD8+ T Cells Supporting a Latent Infection

Journal: PLoS ONE

doi: 10.1371/journal.pone.0001627

Induction of WB-MCF in rabbits. A. Cumulative incidence survival curve of AlHV-1 infected rabbits. On day 0, two groups, each consisting of four rabbits, were inoculated intravenously with 3×10 6 mock infected or AlHV-1 infected EBL cells. At day 10 post-inoculation, BrdU (25 mg/kg) was injected intravenously every day to all rabbits. Rabbit developing WD-MCF were euthanized when rectal temperature remained higher than 40°C for two consecutive days. For each euthanized infected rabbit, a contemporary mock infected rabbit was also sacrificed. Rabbits were identified according to their infected (IR) or mock infected (MR) status and according to the time of death (see numbers). B. Popliteal lymph nodes were dissected during necropsy examination. C. Relative proportions of CD11b + , IgM + , CD4 + and CD8 + cells in PBMC, popliteal lymph node and spleen of AlHV-1 infected (IR) or mock infected rabbits (MR). At time of death, cell suspensions were prepared, stained and analysed by flow cytometry.
Figure Legend Snippet: Induction of WB-MCF in rabbits. A. Cumulative incidence survival curve of AlHV-1 infected rabbits. On day 0, two groups, each consisting of four rabbits, were inoculated intravenously with 3×10 6 mock infected or AlHV-1 infected EBL cells. At day 10 post-inoculation, BrdU (25 mg/kg) was injected intravenously every day to all rabbits. Rabbit developing WD-MCF were euthanized when rectal temperature remained higher than 40°C for two consecutive days. For each euthanized infected rabbit, a contemporary mock infected rabbit was also sacrificed. Rabbits were identified according to their infected (IR) or mock infected (MR) status and according to the time of death (see numbers). B. Popliteal lymph nodes were dissected during necropsy examination. C. Relative proportions of CD11b + , IgM + , CD4 + and CD8 + cells in PBMC, popliteal lymph node and spleen of AlHV-1 infected (IR) or mock infected rabbits (MR). At time of death, cell suspensions were prepared, stained and analysed by flow cytometry.

Techniques Used: Western Blot, Infection, Injection, Staining, Flow Cytometry, Cytometry

Analysis of in vivo BrdU incorporation. Rabbits were treated as described in Fig. 1 . PBMCs were collected at days 11, 15, 17, 20 and 24 post-inoculation, while mononuclear cells were isolated from popliteal lymph node and spleen at the time of death. Cells were labelled with anti-CD11b, IgM, CD5, CD4 and CD8 mAbs as the primary antibodies. Alexa 633-GAM was used as the secondary antibody. In vivo BrdU incorporation was revealed by immunofluorescent staining as described in Methods . After staining, cells were analysed by flow cytometry. A. Representative flow cytometry dot plots are shown for each double staining, they illustrate the data obtained at day 17 post-infection for the PBMC of rabbits MR17/1 and IR17/1. The data represent the percentages of BrdU positive cells ( y -axis) calculated based on the acquisition of 10,000 cells expressing the indicated cell marker ( x -axis). B. The percentage of BrdU positive cells amongst the indicated cellular subset was determined and compared between AlHV-1 infected (left column: bold lines; middle and right columns: hatched bars) and mock infected (left column: dotted lines; middle and right columns: open bars) groups (* P
Figure Legend Snippet: Analysis of in vivo BrdU incorporation. Rabbits were treated as described in Fig. 1 . PBMCs were collected at days 11, 15, 17, 20 and 24 post-inoculation, while mononuclear cells were isolated from popliteal lymph node and spleen at the time of death. Cells were labelled with anti-CD11b, IgM, CD5, CD4 and CD8 mAbs as the primary antibodies. Alexa 633-GAM was used as the secondary antibody. In vivo BrdU incorporation was revealed by immunofluorescent staining as described in Methods . After staining, cells were analysed by flow cytometry. A. Representative flow cytometry dot plots are shown for each double staining, they illustrate the data obtained at day 17 post-infection for the PBMC of rabbits MR17/1 and IR17/1. The data represent the percentages of BrdU positive cells ( y -axis) calculated based on the acquisition of 10,000 cells expressing the indicated cell marker ( x -axis). B. The percentage of BrdU positive cells amongst the indicated cellular subset was determined and compared between AlHV-1 infected (left column: bold lines; middle and right columns: hatched bars) and mock infected (left column: dotted lines; middle and right columns: open bars) groups (* P

Techniques Used: In Vivo, BrdU Incorporation Assay, Isolation, Staining, Flow Cytometry, Cytometry, Double Staining, Infection, Expressing, Marker

28) Product Images from "Ciliary Neurotrophic Factor Mediates Dopamine D2 Receptor-Induced CNS Neurogenesis in Adult Mice"

Article Title: Ciliary Neurotrophic Factor Mediates Dopamine D2 Receptor-Induced CNS Neurogenesis in Adult Mice

Journal: The Journal of Neuroscience

doi: 10.1523/JNEUROSCI.3574-07.2008

Dopamine D 2 receptor promotes neural precursor proliferation in the adult mouse forebrain SVZ. Compared with the number of BrdU-labeled nuclei in coronal sections through the SVZ of C57BL/6 mice injected intraperitoneally with saline ( A , B ), that of mice
Figure Legend Snippet: Dopamine D 2 receptor promotes neural precursor proliferation in the adult mouse forebrain SVZ. Compared with the number of BrdU-labeled nuclei in coronal sections through the SVZ of C57BL/6 mice injected intraperitoneally with saline ( A , B ), that of mice

Techniques Used: Labeling, Mouse Assay, Injection

D 2 -induced SVZ proliferation is dependent on CNTF. A , A 3 d, 18 mg/kg/d quinpirole treatment increases the number of BrdU + nuclei in the SVZ of adult CNTF +/+ mice but not in their CNTF −/− littermates, suggesting endogenous CNTF mediates
Figure Legend Snippet: D 2 -induced SVZ proliferation is dependent on CNTF. A , A 3 d, 18 mg/kg/d quinpirole treatment increases the number of BrdU + nuclei in the SVZ of adult CNTF +/+ mice but not in their CNTF −/− littermates, suggesting endogenous CNTF mediates

Techniques Used: Mouse Assay

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Clone Assay:

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Immunocytochemistry:

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Synthesized:

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Cytometry:

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Immunostaining:

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Article Title: Genome-Wide Analysis of M?ller Glial Differentiation Reveals a Requirement for Notch Signaling in Postmitotic Cells to Maintain the Glial Fate
Article Snippet: For the immunofluorescent labeing, we used the following primary antibodies: chicken anti-GFP (1∶500 dilution, Abcam, USA, Cat. No. AB13970); goat anti-Sox2 (1∶500, Santa Cruz Biotechnology, USA, Sox2 Y-17 Cat. No. SC-17320); Cralbp (rabbit, UW55, kindly provided by J. Saari); Sox9 (rabbit, Chemicon); Id1 (sc488, Santa Cruz; TuJ1 (mouse, Covance); Glast (guinea pig, Chemicon); BrdU (rat, Accurate); mouse anti-S-100 (1∶500, Sigma-Aldrich, St. Louis, MO). .. After immunostaining, slides were coverslipped in Fluoromount G (Southern Biotechnology, Birmingham, AL).

Incubation:

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice
Article Snippet: After rinsing in PBS, the sections were incubated in blocking solution (5% rabbit serum and 0.1% Triton X-100) for 60 min, then incubated overnight at 4°C with BrdU rat monoclonal antibody (1:400; Abcam, Cambridge, MA) diluted in 5% normal rabbit serum and 0.1% Triton X-100. .. The brains underwent double immunofluorescence staining with BrdU (rat, 1:400) and NeuN (1:300, mouse; Chemicon International, Temecula, CA), as well as BrdU with GFAP (1:800; mouse, Sigma-Aldrich, St. Louis, MO).

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Article Snippet: For staining with IAb /GP66-77 , Db /NP396–404 , Db /GP276–286 (provided by NIH Tetramer Core Facility; Atlanta, GA) or Db /GP33–41 (Beckman Coulter; Fullerton, CA) tetramers, cells were incubated for 1 h and 15 min at room temperature. .. To quantify incorporation of BrdU by tetramer+ CD8 T cells, mice were injected with 1 mg of BrdU (Sigma-Aldrich) 16 h before analysis and splenocytes stained with BrdU Flow kit (BD Biosciences) following the manufacturer instructions.

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Article Title: Genome-Wide Analysis of M?ller Glial Differentiation Reveals a Requirement for Notch Signaling in Postmitotic Cells to Maintain the Glial Fate
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Expressing:

Article Title: Null effect of antidepressants on the astrocytes-mediated proliferation of hippocampal progenitor cells in vitro
Article Snippet: After washing out the antidepressants, NPCs expressing GFP were overlaid onto astrocytes in the N2 medium containing 20 ng/ml bFGF. .. After the treatment with HCl, cells were incubated with cold PBT (0.1% Triton X-100 and 0.1% BSA in PBS) for 15 min. After blocking with a preblock solution (2% BSA and 0.08% Triton X-100 in PBS) at room temperature for 2 h, the cells were incubated overnight with the following primary antibodies in a preblock solution: BrdU (1:100, mouse; Sigma), GFP (1:500, rabbit; Abcam), nestin (1:400, mouse; BD biosciences), GFAP (1:400, rabbit; DAKO).

Modification:

Article Title: Cited3 activates Mef2c to control muscle cell differentiation and survival
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Article Title: Genome-Wide Analysis of M?ller Glial Differentiation Reveals a Requirement for Notch Signaling in Postmitotic Cells to Maintain the Glial Fate
Article Snippet: Eyes were fixed overnight at 4°C in modified Carnoy's solution (60% ethanol, 11.1% formaldehyde (30% of 37% stock), 10% glacial acetic acid), embedded in paraffin and sectioned at 6–8 µm. .. For the immunofluorescent labeing, we used the following primary antibodies: chicken anti-GFP (1∶500 dilution, Abcam, USA, Cat. No. AB13970); goat anti-Sox2 (1∶500, Santa Cruz Biotechnology, USA, Sox2 Y-17 Cat. No. SC-17320); Cralbp (rabbit, UW55, kindly provided by J. Saari); Sox9 (rabbit, Chemicon); Id1 (sc488, Santa Cruz; TuJ1 (mouse, Covance); Glast (guinea pig, Chemicon); BrdU (rat, Accurate); mouse anti-S-100 (1∶500, Sigma-Aldrich, St. Louis, MO).

Double Immunofluorescence Staining:

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice
Article Snippet: .. The brains underwent double immunofluorescence staining with BrdU (rat, 1:400) and NeuN (1:300, mouse; Chemicon International, Temecula, CA), as well as BrdU with GFAP (1:800; mouse, Sigma-Aldrich, St. Louis, MO). .. The corresponding secondary antibodies used were Alexa Fluor 488 or 594 rabbit anti-rat (1:1000; Invitrogen, Carlsbad, CA) for BrdU, and Alexa Fluor 488 or 594 goat anti-mouse (1:800, Invitrogen) for NeuN and GFAP.

Transformation Assay:

Article Title: Intracellular Viral Processing, Not Single-Stranded DNA Accumulation, Is Crucial for Recombinant Adeno-Associated Virus Transduction
Article Snippet: Cloning vector pBluescriptII (Stratagene) was transformed into competent bacteria. .. To label the plasmid with BrdU, bacteria were grown overnight in Luria-Bertani medium containing 33 μg of BrdU (Sigma)/ml and 0.55 μg of FrdU (Sigma)/ml.

Blocking Assay:

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice
Article Snippet: After rinsing in PBS, the sections were incubated in blocking solution (5% rabbit serum and 0.1% Triton X-100) for 60 min, then incubated overnight at 4°C with BrdU rat monoclonal antibody (1:400; Abcam, Cambridge, MA) diluted in 5% normal rabbit serum and 0.1% Triton X-100. .. The brains underwent double immunofluorescence staining with BrdU (rat, 1:400) and NeuN (1:300, mouse; Chemicon International, Temecula, CA), as well as BrdU with GFAP (1:800; mouse, Sigma-Aldrich, St. Louis, MO).

Article Title: Null effect of antidepressants on the astrocytes-mediated proliferation of hippocampal progenitor cells in vitro
Article Snippet: .. After the treatment with HCl, cells were incubated with cold PBT (0.1% Triton X-100 and 0.1% BSA in PBS) for 15 min. After blocking with a preblock solution (2% BSA and 0.08% Triton X-100 in PBS) at room temperature for 2 h, the cells were incubated overnight with the following primary antibodies in a preblock solution: BrdU (1:100, mouse; Sigma), GFP (1:500, rabbit; Abcam), nestin (1:400, mouse; BD biosciences), GFAP (1:400, rabbit; DAKO). .. After washing with PBT, the cells were incubated with the fluorescence-labeled secondary antibodies for 2 h. Cells were counterstained for 15 min with 10 ng/ml 4', 6'-diamidino-2-phenylindole (DAPI) (Molecular Probes) or 50 μg/ml propidium iodide (PI).

Hybridization:

Article Title: Genome-Wide Analysis of M?ller Glial Differentiation Reveals a Requirement for Notch Signaling in Postmitotic Cells to Maintain the Glial Fate
Article Snippet: Slides were hybridized with probe overnight hybridization and hybridized probe was detected using anti-Digoxygenin alkaline phosphatase conjugated antibody (1∶2000 dilution, Roche Biochemical, Indianapolis, IN). .. For the immunofluorescent labeing, we used the following primary antibodies: chicken anti-GFP (1∶500 dilution, Abcam, USA, Cat. No. AB13970); goat anti-Sox2 (1∶500, Santa Cruz Biotechnology, USA, Sox2 Y-17 Cat. No. SC-17320); Cralbp (rabbit, UW55, kindly provided by J. Saari); Sox9 (rabbit, Chemicon); Id1 (sc488, Santa Cruz; TuJ1 (mouse, Covance); Glast (guinea pig, Chemicon); BrdU (rat, Accurate); mouse anti-S-100 (1∶500, Sigma-Aldrich, St. Louis, MO).

TUNEL Assay:

Article Title: Cited3 activates Mef2c to control muscle cell differentiation and survival
Article Snippet: .. BrdU and Tunel staining Embryos were pulsed for different time periods in 1 mM BrdU (Sigma), fixed in 4% PFA and stained using anti-BrdU antibody (Sigma) following the protocol of Barresi et al. ( ) with minor modification for whole mount and on sections. .. To identify apoptotic cells, in situ cell death detection kit (Roche) was used.

Flow Cytometry:

Article Title: E-Cadherin Marks a Subset of Inflammatory Dendritic Cells that Promote T Cell-Mediated Colitis
Article Snippet: BrdU Labeling and Analysis BALB/c Rag2 −/− mice were transferred with CD4+ CD45RBhi T cells and after 6 weeks, when the recipients displayed clinical signs of intestinal inflammation, the mice were i.p. injected with 2 mg BrdU (Sigma-Aldrich) resuspended in PBS. .. Cell suspensions were prepared at different time points after BrdU administration and were analyzed for BrdU incorporation with the BrdU Flow Kit (BD Biosciences) according to the manufacturer's instructions.

Article Title: TGF-? Signaling in T cells is Essential for CD8 T Cell Suppression and Viral Persistence In Vivo
Article Snippet: .. To quantify incorporation of BrdU by tetramer+ CD8 T cells, mice were injected with 1 mg of BrdU (Sigma-Aldrich) 16 h before analysis and splenocytes stained with BrdU Flow kit (BD Biosciences) following the manufacturer instructions. .. Cells were acquired using the Digital LSR II flow cytometer (Becton Dickinson, San Jose, CA).

Immunolabeling:

Article Title: Genome-Wide Analysis of M?ller Glial Differentiation Reveals a Requirement for Notch Signaling in Postmitotic Cells to Maintain the Glial Fate
Article Snippet: Paragraph title: In Situ Hybridization and Immunolabeling ... For the immunofluorescent labeing, we used the following primary antibodies: chicken anti-GFP (1∶500 dilution, Abcam, USA, Cat. No. AB13970); goat anti-Sox2 (1∶500, Santa Cruz Biotechnology, USA, Sox2 Y-17 Cat. No. SC-17320); Cralbp (rabbit, UW55, kindly provided by J. Saari); Sox9 (rabbit, Chemicon); Id1 (sc488, Santa Cruz; TuJ1 (mouse, Covance); Glast (guinea pig, Chemicon); BrdU (rat, Accurate); mouse anti-S-100 (1∶500, Sigma-Aldrich, St. Louis, MO).

Pentobarbital Overdose:

Article Title: Metformin Acts on Two Different Molecular Pathways to Enhance Adult Neural Precursor Proliferation/Self-Renewal and Differentiation
Article Snippet: BrdU Labeling To quantify proliferating precursors in the SVZ and SGZ, adult mice were injected i.p. with a single dose of BrdU (Sigma-Aldrich) at 100 mg/kg. .. Animals were sacrificed 24 hr later via sodium pentobarbital overdose and transcardially perfused with PBS followed by 4% paraformaldehyde.

Generated:

Article Title: Intracellular Viral Processing, Not Single-Stranded DNA Accumulation, Is Crucial for Recombinant Adeno-Associated Virus Transduction
Article Snippet: To label the plasmid with BrdU, bacteria were grown overnight in Luria-Bertani medium containing 33 μg of BrdU (Sigma)/ml and 0.55 μg of FrdU (Sigma)/ml. .. Labeled DNA was digested with XbaI and KpnI, and ss DNA was generated by using ExoIII nuclease.

Injection:

Article Title: E-Cadherin Marks a Subset of Inflammatory Dendritic Cells that Promote T Cell-Mediated Colitis
Article Snippet: .. BrdU Labeling and Analysis BALB/c Rag2 −/− mice were transferred with CD4+ CD45RBhi T cells and after 6 weeks, when the recipients displayed clinical signs of intestinal inflammation, the mice were i.p. injected with 2 mg BrdU (Sigma-Aldrich) resuspended in PBS. .. Cell suspensions were prepared at different time points after BrdU administration and were analyzed for BrdU incorporation with the BrdU Flow Kit (BD Biosciences) according to the manufacturer's instructions.

Article Title: Metformin Acts on Two Different Molecular Pathways to Enhance Adult Neural Precursor Proliferation/Self-Renewal and Differentiation
Article Snippet: .. BrdU Labeling To quantify proliferating precursors in the SVZ and SGZ, adult mice were injected i.p. with a single dose of BrdU (Sigma-Aldrich) at 100 mg/kg. .. Animals were sacrificed 24 hr later via sodium pentobarbital overdose and transcardially perfused with PBS followed by 4% paraformaldehyde.

Article Title: TGF-? Signaling in T cells is Essential for CD8 T Cell Suppression and Viral Persistence In Vivo
Article Snippet: .. To quantify incorporation of BrdU by tetramer+ CD8 T cells, mice were injected with 1 mg of BrdU (Sigma-Aldrich) 16 h before analysis and splenocytes stained with BrdU Flow kit (BD Biosciences) following the manufacturer instructions. .. Cells were acquired using the Digital LSR II flow cytometer (Becton Dickinson, San Jose, CA).

Article Title: Effects of Electromagnetic Radiation from Smartphones on Learning Ability and Hippocampal Progenitor Cell Proliferation in Mice
Article Snippet: .. 2.4 Injection of 5-bromo-2′-deoxyuridine and immunohistochemistry To label proliferating cells, mice received an intraperitoneal (i.p.) injection of 5-bromo-2′-deoxyuridine (BrdU; 100 mg/kg, dissolved in saline; Sigma Aldrich, St. Louis, MO, USA) and were sacrificed 1 day later. .. A BrdU immunohistochemistry method was described by Choi et al .

Software:

Article Title: TGF-? Signaling in T cells is Essential for CD8 T Cell Suppression and Viral Persistence In Vivo
Article Snippet: To quantify incorporation of BrdU by tetramer+ CD8 T cells, mice were injected with 1 mg of BrdU (Sigma-Aldrich) 16 h before analysis and splenocytes stained with BrdU Flow kit (BD Biosciences) following the manufacturer instructions. .. Flow cytometric data were analyzed with FlowJo software.

Article Title: Null effect of antidepressants on the astrocytes-mediated proliferation of hippocampal progenitor cells in vitro
Article Snippet: After the treatment with HCl, cells were incubated with cold PBT (0.1% Triton X-100 and 0.1% BSA in PBS) for 15 min. After blocking with a preblock solution (2% BSA and 0.08% Triton X-100 in PBS) at room temperature for 2 h, the cells were incubated overnight with the following primary antibodies in a preblock solution: BrdU (1:100, mouse; Sigma), GFP (1:500, rabbit; Abcam), nestin (1:400, mouse; BD biosciences), GFAP (1:400, rabbit; DAKO). .. Fluorescence-positive cells were quantified in 20 fields (×10) per dish either manually (overlay culture) or by using Image J software (Banker culture).

Immunofluorescence:

Article Title: Adult Hippocampal Neurogenesis Modulation by the Membrane-Associated Progesterone Receptor Family Member Neudesin
Article Snippet: Paragraph title: Immunohistochemistry and Immunofluorescence ... The sections were further incubated with the following primary antibodies: doublecortin (DCX; rabbit polyclonal, Abcam, Cambridge, UK) at a dilution of 1:300, BrdU (rat anti-BrdU, BU1/75 clone, Abcam) at a dilution of 1:100, Calbindin (Calb; rabbit polyclonal, Abcam) at a dilution of 1:300 and neudesin at a dilution of 1:50 (NENF; rabbit polyclonal; Sigma).

Article Title: Null effect of antidepressants on the astrocytes-mediated proliferation of hippocampal progenitor cells in vitro
Article Snippet: After the treatment with HCl, cells were incubated with cold PBT (0.1% Triton X-100 and 0.1% BSA in PBS) for 15 min. After blocking with a preblock solution (2% BSA and 0.08% Triton X-100 in PBS) at room temperature for 2 h, the cells were incubated overnight with the following primary antibodies in a preblock solution: BrdU (1:100, mouse; Sigma), GFP (1:500, rabbit; Abcam), nestin (1:400, mouse; BD biosciences), GFAP (1:400, rabbit; DAKO). .. After the treatment with HCl, cells were incubated with cold PBT (0.1% Triton X-100 and 0.1% BSA in PBS) for 15 min. After blocking with a preblock solution (2% BSA and 0.08% Triton X-100 in PBS) at room temperature for 2 h, the cells were incubated overnight with the following primary antibodies in a preblock solution: BrdU (1:100, mouse; Sigma), GFP (1:500, rabbit; Abcam), nestin (1:400, mouse; BD biosciences), GFAP (1:400, rabbit; DAKO).

Fluorescence:

Article Title: Null effect of antidepressants on the astrocytes-mediated proliferation of hippocampal progenitor cells in vitro
Article Snippet: After the treatment with HCl, cells were incubated with cold PBT (0.1% Triton X-100 and 0.1% BSA in PBS) for 15 min. After blocking with a preblock solution (2% BSA and 0.08% Triton X-100 in PBS) at room temperature for 2 h, the cells were incubated overnight with the following primary antibodies in a preblock solution: BrdU (1:100, mouse; Sigma), GFP (1:500, rabbit; Abcam), nestin (1:400, mouse; BD biosciences), GFAP (1:400, rabbit; DAKO). .. After washing with PBT, the cells were incubated with the fluorescence-labeled secondary antibodies for 2 h. Cells were counterstained for 15 min with 10 ng/ml 4', 6'-diamidino-2-phenylindole (DAPI) (Molecular Probes) or 50 μg/ml propidium iodide (PI).

Isolation:

Article Title: Intracellular Viral Processing, Not Single-Stranded DNA Accumulation, Is Crucial for Recombinant Adeno-Associated Virus Transduction
Article Snippet: To label the plasmid with BrdU, bacteria were grown overnight in Luria-Bertani medium containing 33 μg of BrdU (Sigma)/ml and 0.55 μg of FrdU (Sigma)/ml. .. DNA was isolated with a QIAGEN Plasmid Mega kit.

Immunohistochemistry:

Article Title: Adult Hippocampal Neurogenesis Modulation by the Membrane-Associated Progesterone Receptor Family Member Neudesin
Article Snippet: Paragraph title: Immunohistochemistry and Immunofluorescence ... The sections were further incubated with the following primary antibodies: doublecortin (DCX; rabbit polyclonal, Abcam, Cambridge, UK) at a dilution of 1:300, BrdU (rat anti-BrdU, BU1/75 clone, Abcam) at a dilution of 1:100, Calbindin (Calb; rabbit polyclonal, Abcam) at a dilution of 1:300 and neudesin at a dilution of 1:50 (NENF; rabbit polyclonal; Sigma).

Article Title: Effects of Electromagnetic Radiation from Smartphones on Learning Ability and Hippocampal Progenitor Cell Proliferation in Mice
Article Snippet: .. 2.4 Injection of 5-bromo-2′-deoxyuridine and immunohistochemistry To label proliferating cells, mice received an intraperitoneal (i.p.) injection of 5-bromo-2′-deoxyuridine (BrdU; 100 mg/kg, dissolved in saline; Sigma Aldrich, St. Louis, MO, USA) and were sacrificed 1 day later. .. A BrdU immunohistochemistry method was described by Choi et al .

Microscopy:

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice
Article Snippet: The brains underwent double immunofluorescence staining with BrdU (rat, 1:400) and NeuN (1:300, mouse; Chemicon International, Temecula, CA), as well as BrdU with GFAP (1:800; mouse, Sigma-Aldrich, St. Louis, MO). .. After rinsing in PBS for 5 min, the sections were cover-slipped with Vectashield® (Vector Laboratories) mounting media and visualized using an Olympus BX- microscope.

Article Title: Null effect of antidepressants on the astrocytes-mediated proliferation of hippocampal progenitor cells in vitro
Article Snippet: After the treatment with HCl, cells were incubated with cold PBT (0.1% Triton X-100 and 0.1% BSA in PBS) for 15 min. After blocking with a preblock solution (2% BSA and 0.08% Triton X-100 in PBS) at room temperature for 2 h, the cells were incubated overnight with the following primary antibodies in a preblock solution: BrdU (1:100, mouse; Sigma), GFP (1:500, rabbit; Abcam), nestin (1:400, mouse; BD biosciences), GFAP (1:400, rabbit; DAKO). .. After the treatment with HCl, cells were incubated with cold PBT (0.1% Triton X-100 and 0.1% BSA in PBS) for 15 min. After blocking with a preblock solution (2% BSA and 0.08% Triton X-100 in PBS) at room temperature for 2 h, the cells were incubated overnight with the following primary antibodies in a preblock solution: BrdU (1:100, mouse; Sigma), GFP (1:500, rabbit; Abcam), nestin (1:400, mouse; BD biosciences), GFAP (1:400, rabbit; DAKO).

Avidin-Biotin Assay:

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice
Article Snippet: The following day, the sections were rinsed in PBS and placed in biotinylated rabbit anti-rat antibody (1:200; Vector Laboratories, Burlingame, CA) for 1 h at room temperature, and then in 3% H2 O2 /PBS for 10 min, followed by application of an avidin-biotin complex (ABC) kit (Vector Laboratories), and visualized with diaminobenzidine (DAB) for quantification. .. The brains underwent double immunofluorescence staining with BrdU (rat, 1:400) and NeuN (1:300, mouse; Chemicon International, Temecula, CA), as well as BrdU with GFAP (1:800; mouse, Sigma-Aldrich, St. Louis, MO).

Co-Culture Assay:

Article Title: Null effect of antidepressants on the astrocytes-mediated proliferation of hippocampal progenitor cells in vitro
Article Snippet: Twenty-four hours after the co-culture, NPCs were treated with BrdU (10 μM, Banker culture and 2.5 μM, overlay co-culture, respectively) (Sigma) for 24 h. In case of the overlay co-culture, NPCs and astrocytes were treated with BrdU. .. After the treatment with HCl, cells were incubated with cold PBT (0.1% Triton X-100 and 0.1% BSA in PBS) for 15 min. After blocking with a preblock solution (2% BSA and 0.08% Triton X-100 in PBS) at room temperature for 2 h, the cells were incubated overnight with the following primary antibodies in a preblock solution: BrdU (1:100, mouse; Sigma), GFP (1:500, rabbit; Abcam), nestin (1:400, mouse; BD biosciences), GFAP (1:400, rabbit; DAKO).

Mouse Assay:

Article Title: E-Cadherin Marks a Subset of Inflammatory Dendritic Cells that Promote T Cell-Mediated Colitis
Article Snippet: .. BrdU Labeling and Analysis BALB/c Rag2 −/− mice were transferred with CD4+ CD45RBhi T cells and after 6 weeks, when the recipients displayed clinical signs of intestinal inflammation, the mice were i.p. injected with 2 mg BrdU (Sigma-Aldrich) resuspended in PBS. .. Cell suspensions were prepared at different time points after BrdU administration and were analyzed for BrdU incorporation with the BrdU Flow Kit (BD Biosciences) according to the manufacturer's instructions.

Article Title: Metformin Acts on Two Different Molecular Pathways to Enhance Adult Neural Precursor Proliferation/Self-Renewal and Differentiation
Article Snippet: .. BrdU Labeling To quantify proliferating precursors in the SVZ and SGZ, adult mice were injected i.p. with a single dose of BrdU (Sigma-Aldrich) at 100 mg/kg. .. Animals were sacrificed 24 hr later via sodium pentobarbital overdose and transcardially perfused with PBS followed by 4% paraformaldehyde.

Article Title: TGF-? Signaling in T cells is Essential for CD8 T Cell Suppression and Viral Persistence In Vivo
Article Snippet: .. To quantify incorporation of BrdU by tetramer+ CD8 T cells, mice were injected with 1 mg of BrdU (Sigma-Aldrich) 16 h before analysis and splenocytes stained with BrdU Flow kit (BD Biosciences) following the manufacturer instructions. .. Cells were acquired using the Digital LSR II flow cytometer (Becton Dickinson, San Jose, CA).

Article Title: Adult Hippocampal Neurogenesis Modulation by the Membrane-Associated Progesterone Receptor Family Member Neudesin
Article Snippet: For immunofluorescence analysis, six littermate control and six neudesin-null mice were used. .. The sections were further incubated with the following primary antibodies: doublecortin (DCX; rabbit polyclonal, Abcam, Cambridge, UK) at a dilution of 1:300, BrdU (rat anti-BrdU, BU1/75 clone, Abcam) at a dilution of 1:100, Calbindin (Calb; rabbit polyclonal, Abcam) at a dilution of 1:300 and neudesin at a dilution of 1:50 (NENF; rabbit polyclonal; Sigma).

Article Title: Effects of Electromagnetic Radiation from Smartphones on Learning Ability and Hippocampal Progenitor Cell Proliferation in Mice
Article Snippet: .. 2.4 Injection of 5-bromo-2′-deoxyuridine and immunohistochemistry To label proliferating cells, mice received an intraperitoneal (i.p.) injection of 5-bromo-2′-deoxyuridine (BrdU; 100 mg/kg, dissolved in saline; Sigma Aldrich, St. Louis, MO, USA) and were sacrificed 1 day later. .. A BrdU immunohistochemistry method was described by Choi et al .

In Situ Hybridization:

Article Title: Genome-Wide Analysis of M?ller Glial Differentiation Reveals a Requirement for Notch Signaling in Postmitotic Cells to Maintain the Glial Fate
Article Snippet: Paragraph title: In Situ Hybridization and Immunolabeling ... For the immunofluorescent labeing, we used the following primary antibodies: chicken anti-GFP (1∶500 dilution, Abcam, USA, Cat. No. AB13970); goat anti-Sox2 (1∶500, Santa Cruz Biotechnology, USA, Sox2 Y-17 Cat. No. SC-17320); Cralbp (rabbit, UW55, kindly provided by J. Saari); Sox9 (rabbit, Chemicon); Id1 (sc488, Santa Cruz; TuJ1 (mouse, Covance); Glast (guinea pig, Chemicon); BrdU (rat, Accurate); mouse anti-S-100 (1∶500, Sigma-Aldrich, St. Louis, MO).

Plasmid Preparation:

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice
Article Snippet: The brains underwent double immunofluorescence staining with BrdU (rat, 1:400) and NeuN (1:300, mouse; Chemicon International, Temecula, CA), as well as BrdU with GFAP (1:800; mouse, Sigma-Aldrich, St. Louis, MO). .. After rinsing in PBS for 5 min, the sections were cover-slipped with Vectashield® (Vector Laboratories) mounting media and visualized using an Olympus BX- microscope.

Article Title: Intracellular Viral Processing, Not Single-Stranded DNA Accumulation, Is Crucial for Recombinant Adeno-Associated Virus Transduction
Article Snippet: .. To label the plasmid with BrdU, bacteria were grown overnight in Luria-Bertani medium containing 33 μg of BrdU (Sigma)/ml and 0.55 μg of FrdU (Sigma)/ml. .. DNA was isolated with a QIAGEN Plasmid Mega kit.

BrdU Staining:

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice
Article Snippet: Briefly, for BrdU staining, the brain sections were thawed and air-dried for 30 min, rinsed in PBS (pH 7.4) for 5 min, then incubated at 37°C in 2 M HCl for 40 min to denature the DNA therein. .. The brains underwent double immunofluorescence staining with BrdU (rat, 1:400) and NeuN (1:300, mouse; Chemicon International, Temecula, CA), as well as BrdU with GFAP (1:800; mouse, Sigma-Aldrich, St. Louis, MO).

In Situ:

Article Title: Cited3 activates Mef2c to control muscle cell differentiation and survival
Article Snippet: BrdU and Tunel staining Embryos were pulsed for different time periods in 1 mM BrdU (Sigma), fixed in 4% PFA and stained using anti-BrdU antibody (Sigma) following the protocol of Barresi et al. ( ) with minor modification for whole mount and on sections. .. To identify apoptotic cells, in situ cell death detection kit (Roche) was used.

Labeling:

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice
Article Snippet: For the Ki-67 (1:500 rabbit polyclonal antibody; Abcam) and DCX (1:300 goat polyclonal antibody; Santa Cruz Biotechnology, Santa Cruz, CA) labeling, the protocol employed was identical to that used for BrdU minus the DNA denaturating step. .. The brains underwent double immunofluorescence staining with BrdU (rat, 1:400) and NeuN (1:300, mouse; Chemicon International, Temecula, CA), as well as BrdU with GFAP (1:800; mouse, Sigma-Aldrich, St. Louis, MO).

Article Title: E-Cadherin Marks a Subset of Inflammatory Dendritic Cells that Promote T Cell-Mediated Colitis
Article Snippet: .. BrdU Labeling and Analysis BALB/c Rag2 −/− mice were transferred with CD4+ CD45RBhi T cells and after 6 weeks, when the recipients displayed clinical signs of intestinal inflammation, the mice were i.p. injected with 2 mg BrdU (Sigma-Aldrich) resuspended in PBS. .. Cell suspensions were prepared at different time points after BrdU administration and were analyzed for BrdU incorporation with the BrdU Flow Kit (BD Biosciences) according to the manufacturer's instructions.

Article Title: Metformin Acts on Two Different Molecular Pathways to Enhance Adult Neural Precursor Proliferation/Self-Renewal and Differentiation
Article Snippet: .. BrdU Labeling To quantify proliferating precursors in the SVZ and SGZ, adult mice were injected i.p. with a single dose of BrdU (Sigma-Aldrich) at 100 mg/kg. .. Animals were sacrificed 24 hr later via sodium pentobarbital overdose and transcardially perfused with PBS followed by 4% paraformaldehyde.

Article Title: Support for the immortal strand hypothesis
Article Snippet: .. BrdU and dye labeling 0.6 μM BrdU (Sigma-Aldrich) was used to label synthesized DNA. ..

Article Title: Genome-Wide Analysis of M?ller Glial Differentiation Reveals a Requirement for Notch Signaling in Postmitotic Cells to Maintain the Glial Fate
Article Snippet: After in situ hybridization, sections were post-fixed in 4% PFA and rinsed in PBS prior to immunofluorescent labeling. .. For the immunofluorescent labeing, we used the following primary antibodies: chicken anti-GFP (1∶500 dilution, Abcam, USA, Cat. No. AB13970); goat anti-Sox2 (1∶500, Santa Cruz Biotechnology, USA, Sox2 Y-17 Cat. No. SC-17320); Cralbp (rabbit, UW55, kindly provided by J. Saari); Sox9 (rabbit, Chemicon); Id1 (sc488, Santa Cruz; TuJ1 (mouse, Covance); Glast (guinea pig, Chemicon); BrdU (rat, Accurate); mouse anti-S-100 (1∶500, Sigma-Aldrich, St. Louis, MO).

Article Title: Regeneration of the zebrafish retinal pigment epithelium after widespread genetic ablation
Article Snippet: .. BrdU and EdU exposures For BrdU labeling, larvae were incubated in system water containing 10mM BrdU (Sigma-Aldrich) for 24 hours at various time points. .. After incubation, larvae were either immediately fixed for analysis or rinsed with fresh system water and then allowed to recover until 7dpi.

Article Title: Intracellular Viral Processing, Not Single-Stranded DNA Accumulation, Is Crucial for Recombinant Adeno-Associated Virus Transduction
Article Snippet: To label the plasmid with BrdU, bacteria were grown overnight in Luria-Bertani medium containing 33 μg of BrdU (Sigma)/ml and 0.55 μg of FrdU (Sigma)/ml. .. Labeled DNA was digested with XbaI and KpnI, and ss DNA was generated by using ExoIII nuclease.

BrdU Incorporation Assay:

Article Title: E-Cadherin Marks a Subset of Inflammatory Dendritic Cells that Promote T Cell-Mediated Colitis
Article Snippet: BrdU Labeling and Analysis BALB/c Rag2 −/− mice were transferred with CD4+ CD45RBhi T cells and after 6 weeks, when the recipients displayed clinical signs of intestinal inflammation, the mice were i.p. injected with 2 mg BrdU (Sigma-Aldrich) resuspended in PBS. .. Cell suspensions were prepared at different time points after BrdU administration and were analyzed for BrdU incorporation with the BrdU Flow Kit (BD Biosciences) according to the manufacturer's instructions.

Concentration Assay:

Article Title: Support for the immortal strand hypothesis
Article Snippet: BrdU and dye labeling 0.6 μM BrdU (Sigma-Aldrich) was used to label synthesized DNA. .. BrdU was applied at the same concentration and time interval in ESCs and fibroblasts.

Marker:

Article Title: Nestin-positive mesenchymal stem cells favour the astroglial lineage in neural progenitors and stem cells by releasing active BMP4
Article Snippet: .. BrdU and propidium iodide incorporation After 24 hours or 3 days of culture, BrdU (20 μM, Sigma) which is a S-phase marker, or propidium iodide (400 mg/ml, Sigma) was added to the differentiating neurosphere cultures for 18 hours before fixation and staining. ..

Staining:

Article Title: Imipramine Treatment Improves Cognitive Outcome Associated with Enhanced Hippocampal Neurogenesis after Traumatic Brain Injury in Mice
Article Snippet: Sections were counterstained with Nissl stain to visualize the cell bodies. .. The brains underwent double immunofluorescence staining with BrdU (rat, 1:400) and NeuN (1:300, mouse; Chemicon International, Temecula, CA), as well as BrdU with GFAP (1:800; mouse, Sigma-Aldrich, St. Louis, MO).

Article Title: TGF-? Signaling in T cells is Essential for CD8 T Cell Suppression and Viral Persistence In Vivo
Article Snippet: .. To quantify incorporation of BrdU by tetramer+ CD8 T cells, mice were injected with 1 mg of BrdU (Sigma-Aldrich) 16 h before analysis and splenocytes stained with BrdU Flow kit (BD Biosciences) following the manufacturer instructions. .. Cells were acquired using the Digital LSR II flow cytometer (Becton Dickinson, San Jose, CA).

Article Title: Cited3 activates Mef2c to control muscle cell differentiation and survival
Article Snippet: .. BrdU and Tunel staining Embryos were pulsed for different time periods in 1 mM BrdU (Sigma), fixed in 4% PFA and stained using anti-BrdU antibody (Sigma) following the protocol of Barresi et al. ( ) with minor modification for whole mount and on sections. .. To identify apoptotic cells, in situ cell death detection kit (Roche) was used.

Article Title: Nestin-positive mesenchymal stem cells favour the astroglial lineage in neural progenitors and stem cells by releasing active BMP4
Article Snippet: .. BrdU and propidium iodide incorporation After 24 hours or 3 days of culture, BrdU (20 μM, Sigma) which is a S-phase marker, or propidium iodide (400 mg/ml, Sigma) was added to the differentiating neurosphere cultures for 18 hours before fixation and staining. ..

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  • 82
    Millipore in vivo brdu labeling
    <t>LCs</t> in chimeric DKO* mice originate from the bone marrow A–H Lethally irradiated DKO* mice were reconstituted with CD45.1-expressing bone marrow cells, before psoriatic disease was induced. Mice were analyzed on day 9 after disease induction. (A) Representative image showing epidermal ear sheet of a DKO* mouse stained for Langerin (green) and CD45.1 (red). Scale bar indicates 100 μm. (B) Flow cytometric analysis of ear and tail epidermal LCs (CD45 + Lan + cells) that express CD45.2 (host-derived) or CD45.1 (donor-derived), and quantification of (C) host-derived LCs (CD45 + Lan + cells) and (D) donor-derived LCs (CD45 + Lan + cells) ( n = 6) of indicated mice. (E) Flow cytometric analysis showing <t>BrdU</t> incorporation in host- or donor-derived ear and tail epidermal LCs (CD45 + Langerin + cells) and (F) quantification of BrdU + LCs of host- or donor-derived origin ( n = 16). (G) Flow cytometric analysis showing Ki-67 expression by host- or donor-derived ear and tail epidermal LCs (CD45 + Langerin + cells) and (H) quantification of Ki-67 + LCs of host- or donor-derived origin ( n = 16). I–L C57BL/6J mice were reconstituted with CD45.1-expressing syngeneic bone marrow, and mouse ears were treated daily with Imiquimod (Imi) for 6 days and analyzed on day 8. (I) Representative image of an epidermal ear sheet of a DKO* mouse stained for Langerin (green) and CD45.2 (red). (J) Flow cytometric analysis showing host (CD45.2 + ) and donor (CD45.1 + ) contribution to ear and tail epidermal LCs (CD45 + Lan + cells). (K) Percentage of host-derived (CD45.2 + ) LCs ( n = 2–4) or (L) donor-derived (CD45.1 + ) LCs of total epidermal cells of the indicated mice ( n = 2–4). Data information: Flow cytometric quantifications are depicted as percentage of live cells. Data were analyzed using Mann–Whitney U -test (* P
    In Vivo Brdu Labeling, supplied by Millipore, used in various techniques. Bioz Stars score: 82/100, based on 16 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore brdu proliferation assay proliferation
    Low concentrations of 2-DG and metformin reduce viability of <t>MDA-MB-231</t> cells synergistically. (A) MDA-MB-231 cells were treated with metformin and 600 μM 2-DG for 72 hours in low-glucose (1 g/L) RPMI-1640. Medium was renewed daily. Viability was determined by MTS assay. Results are means±SEM (n = 3–4). * P ≤0.05. (B) MDA-MB-231 cells were treated with metformin and 600 μM 2-DG for 72 hours in low-glucose RPMI-1640. Medium was renewed daily. Cell number was determined by Hoechst assay. Results are means±SEM (n = 3–4). * P ≤0.05. (C) MDA-MB-231 cells were treated metformin and 600 μM 2-DG in low-glucose RPMI-1640 for 24 hours. Proliferation was determined by <t>BrdU</t> assay. Results are means±SEM (n = 3). * P ≤0.05.
    Brdu Proliferation Assay Proliferation, supplied by Millipore, used in various techniques. Bioz Stars score: 78/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore bromodeoxyuridine brdu labeling solution
    NF-κB activation promoted Oli-neu cell survival in response to <t>IFN-γ.</t> A. The cells were treated with 50 U/ml, 100 U/ml, or 150 U/ml IFN-γ for 24 hrs. MTT assay showed that IFN-γ treatment reduced the number of Oli-neu cells in a dose-dependent manner and that enforced expression of IκBαΔN further reduced the cell numbers. B. The cells were treated with 100 U/ml IFN-γ for 24 hrs. Caspase-3 activity assay showed that IFN-γ treatment did not alter the activity of caspase-3 in Oli-neu cells. Nevertheless, IFN-γ treatment significantly increased the activity of caspase-3 in IκBαΔN 1 and 4 cells. C , D. The cells were treated with 100 U/ml IFN-γ for 24 hrs. Active caspase-3 immunostaining showed that IFN-γ treatment did not change the percentage of active caspase-3 positive cells in Oli-neu cells. Nevertheless, IFN-γ treatment significantly increased the percentage of active caspase-3 positive cells in IκBαΔN 1 and 4 cells. E. <t>BrdU</t> cell proliferation assay showed that the treatment with 100 U/ml IFN-γ for 24 hrs significantly suppressed Oli-neu cell proliferation. Nevertheless, suppression of the NF-κB pathway did not significantly alter the sensitivity of Oli-neu cells to the antiproliferative effects of IFN-γ. The experiments were repeated at least three times, error bars represent standard deviation, asterisk p
    Bromodeoxyuridine Brdu Labeling Solution, supplied by Millipore, used in various techniques. Bioz Stars score: 77/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore brdu
    <t>SVZ</t> neurogenesis is not affected by P2X7 antagonist or agonist injections. <t>BrdU-positive</t> cells of are shown in coronal sections through the SVZ of adult C57BL/6 mice treated with saline ( a ), BBG ( b ), and BzATP ( c ). CC corpus callosum, LV lateral ventricle,
    Brdu, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2228 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    LCs in chimeric DKO* mice originate from the bone marrow A–H Lethally irradiated DKO* mice were reconstituted with CD45.1-expressing bone marrow cells, before psoriatic disease was induced. Mice were analyzed on day 9 after disease induction. (A) Representative image showing epidermal ear sheet of a DKO* mouse stained for Langerin (green) and CD45.1 (red). Scale bar indicates 100 μm. (B) Flow cytometric analysis of ear and tail epidermal LCs (CD45 + Lan + cells) that express CD45.2 (host-derived) or CD45.1 (donor-derived), and quantification of (C) host-derived LCs (CD45 + Lan + cells) and (D) donor-derived LCs (CD45 + Lan + cells) ( n = 6) of indicated mice. (E) Flow cytometric analysis showing BrdU incorporation in host- or donor-derived ear and tail epidermal LCs (CD45 + Langerin + cells) and (F) quantification of BrdU + LCs of host- or donor-derived origin ( n = 16). (G) Flow cytometric analysis showing Ki-67 expression by host- or donor-derived ear and tail epidermal LCs (CD45 + Langerin + cells) and (H) quantification of Ki-67 + LCs of host- or donor-derived origin ( n = 16). I–L C57BL/6J mice were reconstituted with CD45.1-expressing syngeneic bone marrow, and mouse ears were treated daily with Imiquimod (Imi) for 6 days and analyzed on day 8. (I) Representative image of an epidermal ear sheet of a DKO* mouse stained for Langerin (green) and CD45.2 (red). (J) Flow cytometric analysis showing host (CD45.2 + ) and donor (CD45.1 + ) contribution to ear and tail epidermal LCs (CD45 + Lan + cells). (K) Percentage of host-derived (CD45.2 + ) LCs ( n = 2–4) or (L) donor-derived (CD45.1 + ) LCs of total epidermal cells of the indicated mice ( n = 2–4). Data information: Flow cytometric quantifications are depicted as percentage of live cells. Data were analyzed using Mann–Whitney U -test (* P

    Journal: EMBO Molecular Medicine

    Article Title: Specific roles for dendritic cell subsets during initiation and progression of psoriasis

    doi: 10.15252/emmm.201404114

    Figure Lengend Snippet: LCs in chimeric DKO* mice originate from the bone marrow A–H Lethally irradiated DKO* mice were reconstituted with CD45.1-expressing bone marrow cells, before psoriatic disease was induced. Mice were analyzed on day 9 after disease induction. (A) Representative image showing epidermal ear sheet of a DKO* mouse stained for Langerin (green) and CD45.1 (red). Scale bar indicates 100 μm. (B) Flow cytometric analysis of ear and tail epidermal LCs (CD45 + Lan + cells) that express CD45.2 (host-derived) or CD45.1 (donor-derived), and quantification of (C) host-derived LCs (CD45 + Lan + cells) and (D) donor-derived LCs (CD45 + Lan + cells) ( n = 6) of indicated mice. (E) Flow cytometric analysis showing BrdU incorporation in host- or donor-derived ear and tail epidermal LCs (CD45 + Langerin + cells) and (F) quantification of BrdU + LCs of host- or donor-derived origin ( n = 16). (G) Flow cytometric analysis showing Ki-67 expression by host- or donor-derived ear and tail epidermal LCs (CD45 + Langerin + cells) and (H) quantification of Ki-67 + LCs of host- or donor-derived origin ( n = 16). I–L C57BL/6J mice were reconstituted with CD45.1-expressing syngeneic bone marrow, and mouse ears were treated daily with Imiquimod (Imi) for 6 days and analyzed on day 8. (I) Representative image of an epidermal ear sheet of a DKO* mouse stained for Langerin (green) and CD45.2 (red). (J) Flow cytometric analysis showing host (CD45.2 + ) and donor (CD45.1 + ) contribution to ear and tail epidermal LCs (CD45 + Lan + cells). (K) Percentage of host-derived (CD45.2 + ) LCs ( n = 2–4) or (L) donor-derived (CD45.1 + ) LCs of total epidermal cells of the indicated mice ( n = 2–4). Data information: Flow cytometric quantifications are depicted as percentage of live cells. Data were analyzed using Mann–Whitney U -test (* P

    Article Snippet: In vivo BrdU labeling For proliferation studies of LCs, mice received one intraperitoneal injection of 1.5 mg 5-Bromo-2′deoxyuridine (BrdU, Calbiochem) followed by 1 week of BrdU application via drinking water (0.8 mg/ml).

    Techniques: Mouse Assay, Irradiation, Expressing, Staining, Flow Cytometry, Derivative Assay, BrdU Incorporation Assay, MANN-WHITNEY

    Low concentrations of 2-DG and metformin reduce viability of MDA-MB-231 cells synergistically. (A) MDA-MB-231 cells were treated with metformin and 600 μM 2-DG for 72 hours in low-glucose (1 g/L) RPMI-1640. Medium was renewed daily. Viability was determined by MTS assay. Results are means±SEM (n = 3–4). * P ≤0.05. (B) MDA-MB-231 cells were treated with metformin and 600 μM 2-DG for 72 hours in low-glucose RPMI-1640. Medium was renewed daily. Cell number was determined by Hoechst assay. Results are means±SEM (n = 3–4). * P ≤0.05. (C) MDA-MB-231 cells were treated metformin and 600 μM 2-DG in low-glucose RPMI-1640 for 24 hours. Proliferation was determined by BrdU assay. Results are means±SEM (n = 3). * P ≤0.05.

    Journal: PLoS ONE

    Article Title: Medium Renewal Blocks Anti-Proliferative Effects of Metformin in Cultured MDA-MB-231 Breast Cancer Cells

    doi: 10.1371/journal.pone.0154747

    Figure Lengend Snippet: Low concentrations of 2-DG and metformin reduce viability of MDA-MB-231 cells synergistically. (A) MDA-MB-231 cells were treated with metformin and 600 μM 2-DG for 72 hours in low-glucose (1 g/L) RPMI-1640. Medium was renewed daily. Viability was determined by MTS assay. Results are means±SEM (n = 3–4). * P ≤0.05. (B) MDA-MB-231 cells were treated with metformin and 600 μM 2-DG for 72 hours in low-glucose RPMI-1640. Medium was renewed daily. Cell number was determined by Hoechst assay. Results are means±SEM (n = 3–4). * P ≤0.05. (C) MDA-MB-231 cells were treated metformin and 600 μM 2-DG in low-glucose RPMI-1640 for 24 hours. Proliferation was determined by BrdU assay. Results are means±SEM (n = 3). * P ≤0.05.

    Article Snippet: BrdU proliferation assay Proliferation of MDA-MB-231 cell was determined by Bromodeoxyuridine (BrdU) Cell Proliferation Assay Kit (Calbiochem) according to the manufacturer’s instructions.

    Techniques: Multiple Displacement Amplification, MTS Assay, BrdU Staining

    Medium renewal blocks anti-proliferative effects of metformin in cultured MDA-MB-231 cells. MDA-MB-231 cells were treated with metformin in (A) high-glucose (4.5 g/L), (B) low-glucose (1 g/L) or (C) glucose-free RPMI-1640 (with 10% FBS) for 48 hours. Medium was renewed after 24 hours. Viability was determined by MTS assay. Results are means±SEM (n = 3). * P ≤0.05. (D) MDA-MB-231 cells were treated with metformin in low-glucose RPMI-1640 (with 10% FBS) for 24 hours. Proliferation was determined by BrdU assay. Results are means±SEM (one experiment, 8 replicates). * P ≤0.05. MDA-MB-231 cells were treated with metformin in (E) low-glucose or (F) high-glucose RPMI-1640 (serum-free) for 48 hours. Medium was renewed after 24 hours. Viability was determined by MTS assay. Results are means±SEM (n = 2). * P ≤0.05. (G) MDA-MB-231 cells were treated with 5 mM metfromin for 72 hours in low-glucose RPMI-1640 (with 10% FBS) without medium renewal. Glucose concentrations in media were determined every 24 hours. Results are means±SEM (n = 3). * P ≤0.05 vs. 0 hours. (H, I) Different numbers of MDA-MB-231 cells were treated with 5 mM metformin in low-glucose RPMI-1640 (with 10% FBS) for 72 hours with (black) or without (white) medium renewal. Medium was renewed every 24 hours. (H) MTS assay and (I) Hoechst staining were used to estimate viability and cell number. Results are means±SEM (n = 3). * P ≤0.05.

    Journal: PLoS ONE

    Article Title: Medium Renewal Blocks Anti-Proliferative Effects of Metformin in Cultured MDA-MB-231 Breast Cancer Cells

    doi: 10.1371/journal.pone.0154747

    Figure Lengend Snippet: Medium renewal blocks anti-proliferative effects of metformin in cultured MDA-MB-231 cells. MDA-MB-231 cells were treated with metformin in (A) high-glucose (4.5 g/L), (B) low-glucose (1 g/L) or (C) glucose-free RPMI-1640 (with 10% FBS) for 48 hours. Medium was renewed after 24 hours. Viability was determined by MTS assay. Results are means±SEM (n = 3). * P ≤0.05. (D) MDA-MB-231 cells were treated with metformin in low-glucose RPMI-1640 (with 10% FBS) for 24 hours. Proliferation was determined by BrdU assay. Results are means±SEM (one experiment, 8 replicates). * P ≤0.05. MDA-MB-231 cells were treated with metformin in (E) low-glucose or (F) high-glucose RPMI-1640 (serum-free) for 48 hours. Medium was renewed after 24 hours. Viability was determined by MTS assay. Results are means±SEM (n = 2). * P ≤0.05. (G) MDA-MB-231 cells were treated with 5 mM metfromin for 72 hours in low-glucose RPMI-1640 (with 10% FBS) without medium renewal. Glucose concentrations in media were determined every 24 hours. Results are means±SEM (n = 3). * P ≤0.05 vs. 0 hours. (H, I) Different numbers of MDA-MB-231 cells were treated with 5 mM metformin in low-glucose RPMI-1640 (with 10% FBS) for 72 hours with (black) or without (white) medium renewal. Medium was renewed every 24 hours. (H) MTS assay and (I) Hoechst staining were used to estimate viability and cell number. Results are means±SEM (n = 3). * P ≤0.05.

    Article Snippet: BrdU proliferation assay Proliferation of MDA-MB-231 cell was determined by Bromodeoxyuridine (BrdU) Cell Proliferation Assay Kit (Calbiochem) according to the manufacturer’s instructions.

    Techniques: Cell Culture, Multiple Displacement Amplification, MTS Assay, BrdU Staining, Staining

    NF-κB activation promoted Oli-neu cell survival in response to IFN-γ. A. The cells were treated with 50 U/ml, 100 U/ml, or 150 U/ml IFN-γ for 24 hrs. MTT assay showed that IFN-γ treatment reduced the number of Oli-neu cells in a dose-dependent manner and that enforced expression of IκBαΔN further reduced the cell numbers. B. The cells were treated with 100 U/ml IFN-γ for 24 hrs. Caspase-3 activity assay showed that IFN-γ treatment did not alter the activity of caspase-3 in Oli-neu cells. Nevertheless, IFN-γ treatment significantly increased the activity of caspase-3 in IκBαΔN 1 and 4 cells. C , D. The cells were treated with 100 U/ml IFN-γ for 24 hrs. Active caspase-3 immunostaining showed that IFN-γ treatment did not change the percentage of active caspase-3 positive cells in Oli-neu cells. Nevertheless, IFN-γ treatment significantly increased the percentage of active caspase-3 positive cells in IκBαΔN 1 and 4 cells. E. BrdU cell proliferation assay showed that the treatment with 100 U/ml IFN-γ for 24 hrs significantly suppressed Oli-neu cell proliferation. Nevertheless, suppression of the NF-κB pathway did not significantly alter the sensitivity of Oli-neu cells to the antiproliferative effects of IFN-γ. The experiments were repeated at least three times, error bars represent standard deviation, asterisk p

    Journal: PLoS ONE

    Article Title: Interferon-? Activates Nuclear Factor-? B in Oligodendrocytes through a Process Mediated by the Unfolded Protein Response

    doi: 10.1371/journal.pone.0036408

    Figure Lengend Snippet: NF-κB activation promoted Oli-neu cell survival in response to IFN-γ. A. The cells were treated with 50 U/ml, 100 U/ml, or 150 U/ml IFN-γ for 24 hrs. MTT assay showed that IFN-γ treatment reduced the number of Oli-neu cells in a dose-dependent manner and that enforced expression of IκBαΔN further reduced the cell numbers. B. The cells were treated with 100 U/ml IFN-γ for 24 hrs. Caspase-3 activity assay showed that IFN-γ treatment did not alter the activity of caspase-3 in Oli-neu cells. Nevertheless, IFN-γ treatment significantly increased the activity of caspase-3 in IκBαΔN 1 and 4 cells. C , D. The cells were treated with 100 U/ml IFN-γ for 24 hrs. Active caspase-3 immunostaining showed that IFN-γ treatment did not change the percentage of active caspase-3 positive cells in Oli-neu cells. Nevertheless, IFN-γ treatment significantly increased the percentage of active caspase-3 positive cells in IκBαΔN 1 and 4 cells. E. BrdU cell proliferation assay showed that the treatment with 100 U/ml IFN-γ for 24 hrs significantly suppressed Oli-neu cell proliferation. Nevertheless, suppression of the NF-κB pathway did not significantly alter the sensitivity of Oli-neu cells to the antiproliferative effects of IFN-γ. The experiments were repeated at least three times, error bars represent standard deviation, asterisk p

    Article Snippet: After 24 hrs, IFN-γ and Bromodeoxyuridine (BrdU) labeling solution (Millipore, Billerica, Massachusetts) were added to the culture media for 24 hrs.

    Techniques: Activation Assay, MTT Assay, Expressing, Caspase-3 Activity Assay, Activity Assay, Immunostaining, BrdU Cell Proliferation Assay, Standard Deviation

    SVZ neurogenesis is not affected by P2X7 antagonist or agonist injections. BrdU-positive cells of are shown in coronal sections through the SVZ of adult C57BL/6 mice treated with saline ( a ), BBG ( b ), and BzATP ( c ). CC corpus callosum, LV lateral ventricle,

    Journal: Translational stroke research

    Article Title: P2X7 Receptor Inhibition Increases CNTF in the Subventricular Zone, But Not Neurogenesis or Neuroprotection After Stroke in Adult Mice

    doi: 10.1007/s12975-013-0265-2

    Figure Lengend Snippet: SVZ neurogenesis is not affected by P2X7 antagonist or agonist injections. BrdU-positive cells of are shown in coronal sections through the SVZ of adult C57BL/6 mice treated with saline ( a ), BBG ( b ), and BzATP ( c ). CC corpus callosum, LV lateral ventricle,

    Article Snippet: Starting at a random point along the rostrocaudal axis of the brain, every sixth section through the SVZ was immunostained for BrdU (MAB3510, mouse IgG, clone BU-1, 1:30,000; Millipore, Billerica, MA, USA).

    Techniques: Mouse Assay