rabbit anti β gal  (Millipore)


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  • 93
    Name:
    Anti Beta Gal antibody
    Description:

    Catalog Number:
    sab2700176
    Price:
    None
    Applications:
    Suggested starting dilutions are as follows: WB: Assay-dependent dilution. Not yet tested in other applications. Optimal working dilutions should be determined experimentally by the end user.
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    Structured Review

    Millipore rabbit anti β gal
    Anti Beta Gal antibody

    https://www.bioz.com/result/rabbit anti β gal/product/Millipore
    Average 93 stars, based on 12 article reviews
    Price from $9.99 to $1999.99
    rabbit anti β gal - by Bioz Stars, 2020-08
    93/100 stars

    Images

    1) Product Images from "Four Classes of Intercellular Channels between Glial Cells in the CNS"

    Article Title: Four Classes of Intercellular Channels between Glial Cells in the CNS

    Journal: The Journal of Neuroscience

    doi: 10.1523/JNEUROSCI.3303-03.2004

    The majority of oligodendrocytes simultaneously transcribe Cx29, Cx32, and Cx47. A , In heterozygote spinal cord, β-gal (red) was observed in every CC1-positive (green) cell. Note the small soma consisting of large nucleus (red) and scant cytoplasm (green), typical of oligodendrocytes. Cx32-positive ( B ) and Cx47-positive ( C ) puncta outline typical β-gal-positive cells and are distributed along proximal process. Scale bars, 10 μm.
    Figure Legend Snippet: The majority of oligodendrocytes simultaneously transcribe Cx29, Cx32, and Cx47. A , In heterozygote spinal cord, β-gal (red) was observed in every CC1-positive (green) cell. Note the small soma consisting of large nucleus (red) and scant cytoplasm (green), typical of oligodendrocytes. Cx32-positive ( B ) and Cx47-positive ( C ) puncta outline typical β-gal-positive cells and are distributed along proximal process. Scale bars, 10 μm.

    Techniques Used:

    2) Product Images from "The receptor tyrosine kinase Pvr promotes tissue closure by coordinating corpse removal and epidermal zippering"

    Article Title: The receptor tyrosine kinase Pvr promotes tissue closure by coordinating corpse removal and epidermal zippering

    Journal: Development (Cambridge, England)

    doi: 10.1242/dev.122226

    JNK signaling is unaffected by changes in Pvr activity. (A-G) Immunostaining of mid-stage dorsal closure embryos for Fas3 (or Pvr) and β-gal protein from the JNK transcriptional reporter puc-lacZ at the LE (arrows). pnr-G4 directs transgene expression
    Figure Legend Snippet: JNK signaling is unaffected by changes in Pvr activity. (A-G) Immunostaining of mid-stage dorsal closure embryos for Fas3 (or Pvr) and β-gal protein from the JNK transcriptional reporter puc-lacZ at the LE (arrows). pnr-G4 directs transgene expression

    Techniques Used: Activity Assay, Immunostaining, Expressing

    3) Product Images from "Activation of Smurf E3 Ligase Promoted by Smoothened Regulates Hedgehog Signaling through Targeting Patched Turnover"

    Article Title: Activation of Smurf E3 Ligase Promoted by Smoothened Regulates Hedgehog Signaling through Targeting Patched Turnover

    Journal: PLoS Biology

    doi: 10.1371/journal.pbio.1001721

    Smurf positively regulates Hh signaling pathway in wing discs. (A) Wild-type (WT) wing discs carrying dpp-LacZ ( dppZ ) reporter were immunostained with anti-β Gal antibody to show the expression of dppZ (red). (B–B″) Wing discs expressing Flag:Smurf driven by MS1096-gal4 were immunostained to show the expression of dppZ (red), Ci (blue), and Flag (green). (C–C′″) Wing discs expressing Flag:Smurf C1029A driven by MS1096-gal4 were immunostained to show the expression of dppZ (red), Ptc (green), and Ci (blue). (D–D′″) Wing discs expressing shmiR smurf driven by MS1096-gal4 were immunostained to show the expression of dppZ (red), Ptc (green), and Ci (blue). (E and F–F″) Low (E) and high (F–F″) magnification views of a smurf 15c clone at A/P boundary. Immunostaining was performed to show the expression of dppZ (red) and GFP (green). Smurf 15c clones are recognized by the lack of GFP. (G and G′) Wing discs expressing Flag:Smurf driven by MS1096-gal4 were immunostained with Ptc antibody to show the expression of Ptc (red) and Flag (green). (H and H′) Wing discs expressing Flag:Smurf driven by MS1096-gal4 were immunostained to show the expression of ptcZ (red) and Flag (green). (I) Wing discs expressing shmiR smurf driven by MS1096-gal4 were immunostained to show the expression of ptcZ (red). (J–L) comparison of adult wing phenotype from wild-type flies (J), UAS-ptc overexpression driven by en-gal4 (K), and UAS-ptc overexpression driven by en-gal4 in smurf 15C homozygous background (L).
    Figure Legend Snippet: Smurf positively regulates Hh signaling pathway in wing discs. (A) Wild-type (WT) wing discs carrying dpp-LacZ ( dppZ ) reporter were immunostained with anti-β Gal antibody to show the expression of dppZ (red). (B–B″) Wing discs expressing Flag:Smurf driven by MS1096-gal4 were immunostained to show the expression of dppZ (red), Ci (blue), and Flag (green). (C–C′″) Wing discs expressing Flag:Smurf C1029A driven by MS1096-gal4 were immunostained to show the expression of dppZ (red), Ptc (green), and Ci (blue). (D–D′″) Wing discs expressing shmiR smurf driven by MS1096-gal4 were immunostained to show the expression of dppZ (red), Ptc (green), and Ci (blue). (E and F–F″) Low (E) and high (F–F″) magnification views of a smurf 15c clone at A/P boundary. Immunostaining was performed to show the expression of dppZ (red) and GFP (green). Smurf 15c clones are recognized by the lack of GFP. (G and G′) Wing discs expressing Flag:Smurf driven by MS1096-gal4 were immunostained with Ptc antibody to show the expression of Ptc (red) and Flag (green). (H and H′) Wing discs expressing Flag:Smurf driven by MS1096-gal4 were immunostained to show the expression of ptcZ (red) and Flag (green). (I) Wing discs expressing shmiR smurf driven by MS1096-gal4 were immunostained to show the expression of ptcZ (red). (J–L) comparison of adult wing phenotype from wild-type flies (J), UAS-ptc overexpression driven by en-gal4 (K), and UAS-ptc overexpression driven by en-gal4 in smurf 15C homozygous background (L).

    Techniques Used: Expressing, Immunostaining, Clone Assay, Over Expression

    Smurf regulates Hh signaling by controlling Ptc turnover. (A–A′ and B–B′) Wing discs expressing Ptc:GFP fusion protein ubiquitously expressed by heat shock promoter, which also expressed Flag-Smurf or Flag-Smurf C1029A driven by ap-gal4 , were immunostained to show the intensity of GFP (green) and Flag (red). (C) Wild-type control discs were immunostained with anti-Ptc antibody to show Ptc protein expression. (D) Wing discs expressing Smo SD12 driven by MS1096-gal4 were immunostained to show Ptc protein expression levels. (E and F) Wing discs expressing Smo SD12 and Flag:Smurf (E) or Flag:Smurf C1029A (F) driven by MS1096-gal4 were immunostained to show Ptc protein expression levels. (G) Wing discs expressing Smo SD123 driven by MS1096-gal4 were immunostained to show Ptc protein expression. (H and I) Wing discs expressing Smo SD123 and Flag:Smurf (H) or Flag:Smurf C1029A (I) driven by MS1096-gal4 were immunostained to show Ptc protein expression. (J) Wing discs with indicated genotypes were used for Western blots to show levels of Ptc protein; α-tubulin was used as loading control. (K) Wing discs expressing Smo SD123 driven by MS1096-gal4 were immunostained with anti-β-Gal antibody to show ptcZ expression. (L) Wing discs expressing Smo SD123 and Flag:Smurf driven by MS1096-gal4 were immunostained to show ptcZ expression. (M) Wing discs expressing Smo SD123 driven by MS1096-gal4 were immunostained with anti-β-Gal antibody to show dppZ expression. (N) Wing discs expressing Smo SD123 and Flag:Smurf driven by MS1096-gal4 were immunostained to show dppZ expression. (O–O′) Wing discs expressing Hh driven by MS1096-gal4 were immunostained to show Ptc expression. (P–P′) Wing discs expressing Hh and Flag:Smurf driven by MS1096-gal4 were immunostained to show Ptc (red) and Flag (green) expression.
    Figure Legend Snippet: Smurf regulates Hh signaling by controlling Ptc turnover. (A–A′ and B–B′) Wing discs expressing Ptc:GFP fusion protein ubiquitously expressed by heat shock promoter, which also expressed Flag-Smurf or Flag-Smurf C1029A driven by ap-gal4 , were immunostained to show the intensity of GFP (green) and Flag (red). (C) Wild-type control discs were immunostained with anti-Ptc antibody to show Ptc protein expression. (D) Wing discs expressing Smo SD12 driven by MS1096-gal4 were immunostained to show Ptc protein expression levels. (E and F) Wing discs expressing Smo SD12 and Flag:Smurf (E) or Flag:Smurf C1029A (F) driven by MS1096-gal4 were immunostained to show Ptc protein expression levels. (G) Wing discs expressing Smo SD123 driven by MS1096-gal4 were immunostained to show Ptc protein expression. (H and I) Wing discs expressing Smo SD123 and Flag:Smurf (H) or Flag:Smurf C1029A (I) driven by MS1096-gal4 were immunostained to show Ptc protein expression. (J) Wing discs with indicated genotypes were used for Western blots to show levels of Ptc protein; α-tubulin was used as loading control. (K) Wing discs expressing Smo SD123 driven by MS1096-gal4 were immunostained with anti-β-Gal antibody to show ptcZ expression. (L) Wing discs expressing Smo SD123 and Flag:Smurf driven by MS1096-gal4 were immunostained to show ptcZ expression. (M) Wing discs expressing Smo SD123 driven by MS1096-gal4 were immunostained with anti-β-Gal antibody to show dppZ expression. (N) Wing discs expressing Smo SD123 and Flag:Smurf driven by MS1096-gal4 were immunostained to show dppZ expression. (O–O′) Wing discs expressing Hh driven by MS1096-gal4 were immunostained to show Ptc expression. (P–P′) Wing discs expressing Hh and Flag:Smurf driven by MS1096-gal4 were immunostained to show Ptc (red) and Flag (green) expression.

    Techniques Used: Expressing, Western Blot

    4) Product Images from "Anxiety Specific Response and Contribution of Active Hippocampal Neural Stem Cells to Chronic Pain Through Wnt/β-Catenin Signaling in Mice"

    Article Title: Anxiety Specific Response and Contribution of Active Hippocampal Neural Stem Cells to Chronic Pain Through Wnt/β-Catenin Signaling in Mice

    Journal: Frontiers in Molecular Neuroscience

    doi: 10.3389/fnmol.2018.00296

    Effects of chronic pain on Wnt activity in ventral hippocampus. (A) Double-immunostaining of Nestin/β-gal and Western-blotting of β-gal in the ventral hippocampus of sham-injured and SNI-treated Topgal mice at 21 dpi. (B) Western-blotting of β-catenin and Axin2 in the ventral hippocampus of sham-injured and SNI-treated mice at 21 dpi. (C) Double-immunostaining and quantification of Nestin/β-gal in the SVZ of sham-injured and SNI-treated Topgal mice at 21 dpi. Inserts in (A) are magnified typical Nestin/β-gal-positive cells in each group, which were pointed by arrows. Bars = 25 μm in (A) and 50 μm in (C) . Values represent mean ± SE. Unpaired, two tailed Student’s t -tests were performed in (B,C) . * P
    Figure Legend Snippet: Effects of chronic pain on Wnt activity in ventral hippocampus. (A) Double-immunostaining of Nestin/β-gal and Western-blotting of β-gal in the ventral hippocampus of sham-injured and SNI-treated Topgal mice at 21 dpi. (B) Western-blotting of β-catenin and Axin2 in the ventral hippocampus of sham-injured and SNI-treated mice at 21 dpi. (C) Double-immunostaining and quantification of Nestin/β-gal in the SVZ of sham-injured and SNI-treated Topgal mice at 21 dpi. Inserts in (A) are magnified typical Nestin/β-gal-positive cells in each group, which were pointed by arrows. Bars = 25 μm in (A) and 50 μm in (C) . Values represent mean ± SE. Unpaired, two tailed Student’s t -tests were performed in (B,C) . * P

    Techniques Used: Activity Assay, Double Immunostaining, Western Blot, Mouse Assay, Two Tailed Test

    Effects of Fluoxetine treatment on the Wnt activity in hippocampus. (A) Double-immunostaining of Nestin/β-gal in sham mice treated with saline (Sham+S), sham mice treated with Fluoxetine (Sham+F), SNI mice treated with saline (SNI+S), and SNI mice treated with Fluoxetine (SNI+F) at 21 dpi. Notice the significant increase of radial Nestin-positive cells in Fluoxetine treated mice. (B) Western-blotting of β-gal and β-catenin in Topgal mice with the following treatments: sham injury plus saline (Sham+S), sham injury plus Fluoxetine (Sham+F), SNI plus saline (SNI+S) and SNI plus Fluoxetine (SNI+F). Notice that Fluoxetine treatment significantly increased the expression level of β-gal and β-catenin in both sham and SNI-treated mice, as compared to corresponding saline controls. S, saline. F, Fluoxetine. Inserts in (A) are typical double-stained cells in each group, which were pointed by arrows. Bars = 50 μm. Values represent mean ± SE. Unpaired, two tailed Student’s t -tests were performed in (A,B) . * P
    Figure Legend Snippet: Effects of Fluoxetine treatment on the Wnt activity in hippocampus. (A) Double-immunostaining of Nestin/β-gal in sham mice treated with saline (Sham+S), sham mice treated with Fluoxetine (Sham+F), SNI mice treated with saline (SNI+S), and SNI mice treated with Fluoxetine (SNI+F) at 21 dpi. Notice the significant increase of radial Nestin-positive cells in Fluoxetine treated mice. (B) Western-blotting of β-gal and β-catenin in Topgal mice with the following treatments: sham injury plus saline (Sham+S), sham injury plus Fluoxetine (Sham+F), SNI plus saline (SNI+S) and SNI plus Fluoxetine (SNI+F). Notice that Fluoxetine treatment significantly increased the expression level of β-gal and β-catenin in both sham and SNI-treated mice, as compared to corresponding saline controls. S, saline. F, Fluoxetine. Inserts in (A) are typical double-stained cells in each group, which were pointed by arrows. Bars = 50 μm. Values represent mean ± SE. Unpaired, two tailed Student’s t -tests were performed in (A,B) . * P

    Techniques Used: Activity Assay, Double Immunostaining, Mouse Assay, Western Blot, Expressing, Staining, Two Tailed Test

    5) Product Images from "Anxiety Specific Response and Contribution of Active Hippocampal Neural Stem Cells to Chronic Pain Through Wnt/β-Catenin Signaling in Mice"

    Article Title: Anxiety Specific Response and Contribution of Active Hippocampal Neural Stem Cells to Chronic Pain Through Wnt/β-Catenin Signaling in Mice

    Journal: Frontiers in Molecular Neuroscience

    doi: 10.3389/fnmol.2018.00296

    Effects of chronic pain on Wnt activity in ventral hippocampus. (A) Double-immunostaining of Nestin/β-gal and Western-blotting of β-gal in the ventral hippocampus of sham-injured and SNI-treated Topgal mice at 21 dpi. (B) Western-blotting of β-catenin and Axin2 in the ventral hippocampus of sham-injured and SNI-treated mice at 21 dpi. (C) Double-immunostaining and quantification of Nestin/β-gal in the SVZ of sham-injured and SNI-treated Topgal mice at 21 dpi. Inserts in (A) are magnified typical Nestin/β-gal-positive cells in each group, which were pointed by arrows. Bars = 25 μm in (A) and 50 μm in (C) . Values represent mean ± SE. Unpaired, two tailed Student’s t -tests were performed in (B,C) . * P
    Figure Legend Snippet: Effects of chronic pain on Wnt activity in ventral hippocampus. (A) Double-immunostaining of Nestin/β-gal and Western-blotting of β-gal in the ventral hippocampus of sham-injured and SNI-treated Topgal mice at 21 dpi. (B) Western-blotting of β-catenin and Axin2 in the ventral hippocampus of sham-injured and SNI-treated mice at 21 dpi. (C) Double-immunostaining and quantification of Nestin/β-gal in the SVZ of sham-injured and SNI-treated Topgal mice at 21 dpi. Inserts in (A) are magnified typical Nestin/β-gal-positive cells in each group, which were pointed by arrows. Bars = 25 μm in (A) and 50 μm in (C) . Values represent mean ± SE. Unpaired, two tailed Student’s t -tests were performed in (B,C) . * P

    Techniques Used: Activity Assay, Double Immunostaining, Western Blot, Mouse Assay, Two Tailed Test

    Effects of Fluoxetine treatment on the Wnt activity in hippocampus. (A) Double-immunostaining of Nestin/β-gal in sham mice treated with saline (Sham+S), sham mice treated with Fluoxetine (Sham+F), SNI mice treated with saline (SNI+S), and SNI mice treated with Fluoxetine (SNI+F) at 21 dpi. Notice the significant increase of radial Nestin-positive cells in Fluoxetine treated mice. (B) Western-blotting of β-gal and β-catenin in Topgal mice with the following treatments: sham injury plus saline (Sham+S), sham injury plus Fluoxetine (Sham+F), SNI plus saline (SNI+S) and SNI plus Fluoxetine (SNI+F). Notice that Fluoxetine treatment significantly increased the expression level of β-gal and β-catenin in both sham and SNI-treated mice, as compared to corresponding saline controls. S, saline. F, Fluoxetine. Inserts in (A) are typical double-stained cells in each group, which were pointed by arrows. Bars = 50 μm. Values represent mean ± SE. Unpaired, two tailed Student’s t -tests were performed in (A,B) . * P
    Figure Legend Snippet: Effects of Fluoxetine treatment on the Wnt activity in hippocampus. (A) Double-immunostaining of Nestin/β-gal in sham mice treated with saline (Sham+S), sham mice treated with Fluoxetine (Sham+F), SNI mice treated with saline (SNI+S), and SNI mice treated with Fluoxetine (SNI+F) at 21 dpi. Notice the significant increase of radial Nestin-positive cells in Fluoxetine treated mice. (B) Western-blotting of β-gal and β-catenin in Topgal mice with the following treatments: sham injury plus saline (Sham+S), sham injury plus Fluoxetine (Sham+F), SNI plus saline (SNI+S) and SNI plus Fluoxetine (SNI+F). Notice that Fluoxetine treatment significantly increased the expression level of β-gal and β-catenin in both sham and SNI-treated mice, as compared to corresponding saline controls. S, saline. F, Fluoxetine. Inserts in (A) are typical double-stained cells in each group, which were pointed by arrows. Bars = 50 μm. Values represent mean ± SE. Unpaired, two tailed Student’s t -tests were performed in (A,B) . * P

    Techniques Used: Activity Assay, Double Immunostaining, Mouse Assay, Western Blot, Expressing, Staining, Two Tailed Test

    6) Product Images from "Pathologic and Phenotypic Alterations in a Mouse Expressing a Connexin47 Missense Mutation That Causes Pelizaeus-Merzbacher-Like Disease in Humans"

    Article Title: Pathologic and Phenotypic Alterations in a Mouse Expressing a Connexin47 Missense Mutation That Causes Pelizaeus-Merzbacher-Like Disease in Humans

    Journal: PLoS Genetics

    doi: 10.1371/journal.pgen.1002146

    mCx47 M282T and β-gal expression in oligodendrocytes of 40-day-old mutant mice. A–C, LacZ staining of 50 µm brain slices obtained from Cx47 +/M282T mice. The β-gal activity is restricted to nuclear localisation. A, β-Gal positive cells in the corpus callosum show typical oligodendrocytic chain-like organization and coexpression of the oligodendrocytic marker CNPase. LacZ expression was not detectable in GFAP-positive astrocytes (B) or NeuN-positive neurons (C). D, Immunostaining with Cx47 antibodies (green) revealed signals in close proximity of Syto-61 stained nuclei in the corpus callosum of Cx47 +/+ mice. Weaker signals were also found more distal to the nuclei. E, Cx47 antibody stainings on heterozygous Cx47 +/M282T mice resulted in weaker, but similarly localized immunosignals compared to those obtained on wildtype brain tissue. Cx47 gap junction immunosignals were predominantly localized close to β-gal positive nuclei indicated by antibody staining. F, Homozygous Cx47 M282T/M282T mice showed apparent β-gal immunoreactivity, but robust Cx47 gap junction immunosignals were not detected in the perikarya. Very weak Cx47 signals were noticed in brain tissue of Cx47 M282T/M282T mice. G, Immunoblot staining against β-gal on whole brain lysates of 40-day-old mice yielded strong signals with Cx47 M282T/M282T , weaker signals with Cx47 +/M282T and no signals with Cx47 +/+ tissue. H, Immunoblot analysis on Cx47 protein resulted in unspecific bands at 50 kDa of whole brain lysates obtained from wildtype , Cx47 M282T/M282T and Cx47 deleted ( Cx47 −/− ) tissue. Immunoprecipitated Cx47 was detected in wildtype and Cx47 M282T/M282T tissue. Lysates of Cx47-eGFP fusion protein expressing HeLa cells yielded signals at 75 kDa as expected and additional weaker signals at approximately 40 kDa after immunoprecipitation of Cx47 and with crude HeLa cell lysate. Additional signals at 40 kDa in these lanes may represent the cleaved C-terminal region of Cx47. Scale bars: A–C, 50 µm; D–F, 20 µm.
    Figure Legend Snippet: mCx47 M282T and β-gal expression in oligodendrocytes of 40-day-old mutant mice. A–C, LacZ staining of 50 µm brain slices obtained from Cx47 +/M282T mice. The β-gal activity is restricted to nuclear localisation. A, β-Gal positive cells in the corpus callosum show typical oligodendrocytic chain-like organization and coexpression of the oligodendrocytic marker CNPase. LacZ expression was not detectable in GFAP-positive astrocytes (B) or NeuN-positive neurons (C). D, Immunostaining with Cx47 antibodies (green) revealed signals in close proximity of Syto-61 stained nuclei in the corpus callosum of Cx47 +/+ mice. Weaker signals were also found more distal to the nuclei. E, Cx47 antibody stainings on heterozygous Cx47 +/M282T mice resulted in weaker, but similarly localized immunosignals compared to those obtained on wildtype brain tissue. Cx47 gap junction immunosignals were predominantly localized close to β-gal positive nuclei indicated by antibody staining. F, Homozygous Cx47 M282T/M282T mice showed apparent β-gal immunoreactivity, but robust Cx47 gap junction immunosignals were not detected in the perikarya. Very weak Cx47 signals were noticed in brain tissue of Cx47 M282T/M282T mice. G, Immunoblot staining against β-gal on whole brain lysates of 40-day-old mice yielded strong signals with Cx47 M282T/M282T , weaker signals with Cx47 +/M282T and no signals with Cx47 +/+ tissue. H, Immunoblot analysis on Cx47 protein resulted in unspecific bands at 50 kDa of whole brain lysates obtained from wildtype , Cx47 M282T/M282T and Cx47 deleted ( Cx47 −/− ) tissue. Immunoprecipitated Cx47 was detected in wildtype and Cx47 M282T/M282T tissue. Lysates of Cx47-eGFP fusion protein expressing HeLa cells yielded signals at 75 kDa as expected and additional weaker signals at approximately 40 kDa after immunoprecipitation of Cx47 and with crude HeLa cell lysate. Additional signals at 40 kDa in these lanes may represent the cleaved C-terminal region of Cx47. Scale bars: A–C, 50 µm; D–F, 20 µm.

    Techniques Used: Expressing, Mutagenesis, Mouse Assay, Staining, Activity Assay, Marker, Immunostaining, Immunoprecipitation

    The number of β-gal positive cells increases during postnatal development of homozygous mCx47 M282T expressing mice. Sagittal 50 µm brain sections were X-Gal stained for LacZ expression and counterstained with eosin. The LacZ reporter gene reflects the expression of Cx47. A, In the cerebellum of 7-day-old Cx47 M282T/M282T mice several X-Gal stained oligodendrocytes were already found in the medullary centre. B, Stainings on cerebella of P10 mice revealed an increase in the number of β-gal positive cells in the white matter compared to P7 mice. Single β-gal positive cells were already found in the granular layer. C and D, Cx47 expressing cells further increased in number during development of the cerebellum. In addition to their localisation in the cerebellar white matter, LacZ expressing cells were found to be widespread among the granular layer in the cerebellum of 105 day old mice. E, Compared to the number of X-Gal stained cells in the white matter of P7 cerebellum, only few cells were stained in P7 cerebral white matter, localized in the corpus callosum and the hippocampal fimbria. Single cells were found in the cortex proximal to the corpus callosum. The hippocampal gray matter was almost free of β-gal positive cells at this developmental stage. F, On P10 the number of X-Gal stained cells was remarkably increased in the corpus callosum and the hippocampal fimbria compared to P7. β-Gal positive cells decreased in number from ventral to dorsal in the corpus callosum to the cortex. G, P16 animals showed robust LacZ expression in the white matter of the cerebrum including corpus callosum, hippocampal fimbria and hippocampal alveus. Furthermore, β-gal positive cells were present at the hippocampal fissure and near the hippocampal regions Cornu ammonis 1 (CA1) and CA3. Compared to P10 the number of LacZ expressing cells increased dramatically in cortical layers VI to IV of P16 mice. H, 105 days after birth Cx47 expressing cells represented by LacZ reporter gene stainings were localized all over the telencephalon with strong accumulation and typical oligodendrocytic chain-like organizaiton in the white matter. In gray matter, β-gal positive cells were scatterly distributed with a decreasing gradient ranging from layers VI to I of the neocortex. Scale bar: A–H, 500 µm.
    Figure Legend Snippet: The number of β-gal positive cells increases during postnatal development of homozygous mCx47 M282T expressing mice. Sagittal 50 µm brain sections were X-Gal stained for LacZ expression and counterstained with eosin. The LacZ reporter gene reflects the expression of Cx47. A, In the cerebellum of 7-day-old Cx47 M282T/M282T mice several X-Gal stained oligodendrocytes were already found in the medullary centre. B, Stainings on cerebella of P10 mice revealed an increase in the number of β-gal positive cells in the white matter compared to P7 mice. Single β-gal positive cells were already found in the granular layer. C and D, Cx47 expressing cells further increased in number during development of the cerebellum. In addition to their localisation in the cerebellar white matter, LacZ expressing cells were found to be widespread among the granular layer in the cerebellum of 105 day old mice. E, Compared to the number of X-Gal stained cells in the white matter of P7 cerebellum, only few cells were stained in P7 cerebral white matter, localized in the corpus callosum and the hippocampal fimbria. Single cells were found in the cortex proximal to the corpus callosum. The hippocampal gray matter was almost free of β-gal positive cells at this developmental stage. F, On P10 the number of X-Gal stained cells was remarkably increased in the corpus callosum and the hippocampal fimbria compared to P7. β-Gal positive cells decreased in number from ventral to dorsal in the corpus callosum to the cortex. G, P16 animals showed robust LacZ expression in the white matter of the cerebrum including corpus callosum, hippocampal fimbria and hippocampal alveus. Furthermore, β-gal positive cells were present at the hippocampal fissure and near the hippocampal regions Cornu ammonis 1 (CA1) and CA3. Compared to P10 the number of LacZ expressing cells increased dramatically in cortical layers VI to IV of P16 mice. H, 105 days after birth Cx47 expressing cells represented by LacZ reporter gene stainings were localized all over the telencephalon with strong accumulation and typical oligodendrocytic chain-like organizaiton in the white matter. In gray matter, β-gal positive cells were scatterly distributed with a decreasing gradient ranging from layers VI to I of the neocortex. Scale bar: A–H, 500 µm.

    Techniques Used: Expressing, Mouse Assay, Staining

    7) Product Images from "Characterization of an Sf-rhabdovirus-negative S. frugiperda cell line as an alternative host for recombinant protein production in the baculovirus-insect cell system"

    Article Title: Characterization of an Sf-rhabdovirus-negative S. frugiperda cell line as an alternative host for recombinant protein production in the baculovirus-insect cell system

    Journal: Protein expression and purification

    doi: 10.1016/j.pep.2016.02.014

    Recombinant β-gal production. Sf-RVN and Sf9 cells were infected with an Sf-rhabdovirus-negative stock of BacPAK6-ΔChi/Cath at an MOI of 5 pfu/cell. Triplicate samples were harvested at various times post-infection, and clarified intracellular extracts were assayed for (A) β-gal activity, as described in Materials and methods. This plot shows the average results with error bars representing the confidence intervals (P
    Figure Legend Snippet: Recombinant β-gal production. Sf-RVN and Sf9 cells were infected with an Sf-rhabdovirus-negative stock of BacPAK6-ΔChi/Cath at an MOI of 5 pfu/cell. Triplicate samples were harvested at various times post-infection, and clarified intracellular extracts were assayed for (A) β-gal activity, as described in Materials and methods. This plot shows the average results with error bars representing the confidence intervals (P

    Techniques Used: Recombinant, Infection, Activity Assay

    8) Product Images from "Sonic hedgehog signaling regulates a novel epithelial progenitor domain of the hindbrain choroid plexus"

    Article Title: Sonic hedgehog signaling regulates a novel epithelial progenitor domain of the hindbrain choroid plexus

    Journal:

    doi: 10.1242/dev.033795

    Genetic removal of Shh signaling impairs hChP biogenesis. ( A-D ) The transgenic Wnt1-Cre line effectively drives gene deletion in hChPe cells, as indicated by a strong β-Gal staining signal in E13.5 Wnt1-Cre; R26R embryos (A,B) and abrogated
    Figure Legend Snippet: Genetic removal of Shh signaling impairs hChP biogenesis. ( A-D ) The transgenic Wnt1-Cre line effectively drives gene deletion in hChPe cells, as indicated by a strong β-Gal staining signal in E13.5 Wnt1-Cre; R26R embryos (A,B) and abrogated

    Techniques Used: Transgenic Assay, Staining

    Related Articles

    Immunohistochemistry:

    Article Title: Polyploidization Without Mitosis Improves In Vivo Liver Transduction With Lentiviral Vectors
    Article Snippet: .. The presence of β-galactosidase- and GFP-positive hepatocytes was assessed by immunohistochemistry on formalin-fixed/paraffin-embedded sections, using anti-GFP (Clontech, Saint-Germain-en-Laye, France) or anti-β-Gal antibodies (Chemicon International, Temecula, CA). .. Endogenous peroxidase activity was blocked by incubation for 10 min in a 3% H2 O2 solution in methyl alcohol.

    Immunocytochemistry:

    Article Title: Four Classes of Intercellular Channels between Glial Cells in the CNS
    Article Snippet: .. For immunocytochemistry, slides were washed with PBS, blocked with 5% goat serum and 0.1% Triton X-100 in PBS, and incubated overnight at 4°C with combinations of the following primary antibodies: mouse anti-Cx32 [7C6.C7, 1:2 dilution ( )], rabbit anti-β-gal (1:2500 dilution; Chemicon, Temecula, CA), rabbit anti-Cx47 [1:1000 dilution ( )], mouse anti-CC1 [APC (Ab-7), 1:20 dilution; Oncogene Research Products, Boston, MA), and mouse anti-parvalbumin (P3088, 1:1000 dilution; Sigma, St. Louis, MO). .. The monoclonal mouse anti-β-gal Ab (clone 40-1a; 1:10 dilution) was developed by Joshua Sanes (Washington University, St. Louis, MO) and was obtained from the Developmental Studies Hybridoma Bank developed under the auspices of the National Institute of Child Health and Human Development and maintained by the University of Iowa (Department of Biological Sciences, Iowa City, IA).

    Immunohistofluorescence:

    Article Title: Recovery of Myelin after Induction of Oligodendrocyte Cell Death in Postnatal Brain
    Article Snippet: .. An immunohistofluorescence using rabbit anti-β-gal antibody (Sigma) was also performed. ..

    Incubation:

    Article Title: Feline Immunodeficiency Virus as a Gene Transfer Vector in the Rat Nucleus Tractus Solitarii
    Article Snippet: .. For double-label immunofluorescent staining for β Gal and NeuN, brain stem sections were washed with PBS and then blocked with 10% donkey normal serum (Jackson ImmunoResearch Labs, USA) in PBS at 25°C for 1 h. Sections were then incubated in a mixture of rabbit anti- β Gal antibody (1:200, Chemicon, USA) and mouse anti-NeuN (1:50, Chemicon) in 10% donkey normal serum for 24 h in a humid chamber at 25°C. ..

    Article Title: Four Classes of Intercellular Channels between Glial Cells in the CNS
    Article Snippet: .. For immunocytochemistry, slides were washed with PBS, blocked with 5% goat serum and 0.1% Triton X-100 in PBS, and incubated overnight at 4°C with combinations of the following primary antibodies: mouse anti-Cx32 [7C6.C7, 1:2 dilution ( )], rabbit anti-β-gal (1:2500 dilution; Chemicon, Temecula, CA), rabbit anti-Cx47 [1:1000 dilution ( )], mouse anti-CC1 [APC (Ab-7), 1:20 dilution; Oncogene Research Products, Boston, MA), and mouse anti-parvalbumin (P3088, 1:1000 dilution; Sigma, St. Louis, MO). .. The monoclonal mouse anti-β-gal Ab (clone 40-1a; 1:10 dilution) was developed by Joshua Sanes (Washington University, St. Louis, MO) and was obtained from the Developmental Studies Hybridoma Bank developed under the auspices of the National Institute of Child Health and Human Development and maintained by the University of Iowa (Department of Biological Sciences, Iowa City, IA).

    Formalin-fixed Paraffin-Embedded:

    Article Title: Polyploidization Without Mitosis Improves In Vivo Liver Transduction With Lentiviral Vectors
    Article Snippet: .. The presence of β-galactosidase- and GFP-positive hepatocytes was assessed by immunohistochemistry on formalin-fixed/paraffin-embedded sections, using anti-GFP (Clontech, Saint-Germain-en-Laye, France) or anti-β-Gal antibodies (Chemicon International, Temecula, CA). .. Endogenous peroxidase activity was blocked by incubation for 10 min in a 3% H2 O2 solution in methyl alcohol.

    Staining:

    Article Title: The receptor tyrosine kinase Pvr promotes tissue closure by coordinating corpse removal and epidermal zippering
    Article Snippet: .. Staining reagents were: rat anti-Pvr 1:500 , rat anti-Pvr 1:500 , mouse anti-Fas3 1:40 (7G10, DSHB), presorbed rabbit anti-β-gal 1:1500 (Cappel, 55976), mouse anti-phosphotyrosine 1:1000 (4G10, EMD Millipore, 05-321), Texas Red-phalloidin 1:500 (Life Technologies), and Vectashield with DAPI mounting media (Vector Labs). ..

    Article Title: Feline Immunodeficiency Virus as a Gene Transfer Vector in the Rat Nucleus Tractus Solitarii
    Article Snippet: .. For double-label immunofluorescent staining for β Gal and NeuN, brain stem sections were washed with PBS and then blocked with 10% donkey normal serum (Jackson ImmunoResearch Labs, USA) in PBS at 25°C for 1 h. Sections were then incubated in a mixture of rabbit anti- β Gal antibody (1:200, Chemicon, USA) and mouse anti-NeuN (1:50, Chemicon) in 10% donkey normal serum for 24 h in a humid chamber at 25°C. ..

    Article Title: Pkd1 regulates immortalized proliferation of renal tubular epithelial cells through p53 induction and JNK activation
    Article Snippet: .. For immunofluorescence, frozen section were stained with YCC2 (anti–polycystin-2; a kind gift from Y. Cai, Yale University, New Haven, Connecticut, USA), anti–β-gal (Chemicon International Inc.), anti–Na-K ATPase (Upstate), anti–acetylated tubulin, anti-DBA, anti–lectin Lotus tetragonolobus (Sigma-Aldrich), anti–p-ERK, anti–p-Akt, or anti–p-JNK (Cell signaling Technology Inc.). .. Photomicrographs were obtained using a microscope (Carl Zeiss International). β -gal staining of kidneys .

    Immunofluorescence:

    Article Title: Pkd1 regulates immortalized proliferation of renal tubular epithelial cells through p53 induction and JNK activation
    Article Snippet: .. For immunofluorescence, frozen section were stained with YCC2 (anti–polycystin-2; a kind gift from Y. Cai, Yale University, New Haven, Connecticut, USA), anti–β-gal (Chemicon International Inc.), anti–Na-K ATPase (Upstate), anti–acetylated tubulin, anti-DBA, anti–lectin Lotus tetragonolobus (Sigma-Aldrich), anti–p-ERK, anti–p-Akt, or anti–p-JNK (Cell signaling Technology Inc.). .. Photomicrographs were obtained using a microscope (Carl Zeiss International). β -gal staining of kidneys .

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  • 95
    Millipore rabbit anti β galactosidase
    AvidinOX-anchored bTrast and bPert induce apoptosis and senescence of SKBR3 cells ( A ) Cells, with or without AvidinOX (AvOX), were stained with BrdU after 72-hour exposure to bTrast (1 μg/mL) or bPert (0.5 μg/mL) and then analyzed by flow cytometry. The Sub-G1 fraction of total cell population in grey. Representative results of one out of three independent samples. ( B ) HCS fluorescence imaging of cells cultivated 48 hours with bTrast (1 μg/mL) or bPert (0.5 μg/mL) then washed, fixed and stained with rabbit anti-cleaved caspase 3, followed by phycoherythrin-conjugated goat anti-Rabbit IgG (green). Draq5 dye staining of nucleus and cytoplasm (grey). Each image is representative of at least 5 fields of duplicate wells. Magnification 60x. Data are from one representative experiment out of two. ( C ) Caspase 3/7 activity in cells treated 72 hours with Pert or bPert at indicated concentrations, measured by Caspase-Glo 3/7 Assay. Data are expressed as fold change of activity compared to control cells and are the average of four replicates (± SE). Staurosporine (0.1 μg/mL) was included as positive control. Mann-Whitney's test: *** p ≤ 0.001, ** p ≤ 0.01 and * p ≤ 0.05 vs AvOX. ( D ) Cells were cultivated 6 days with antibodies as in B, and additional 3 days without antibodies. Cells in left panel were stained with rabbit <t>anti-β-Galactosidase</t> antibody followed by FITC-conjugated goat anti-rabbit IgG (violet). Cells in right panel were treated 1 hour with bafilomycin A1 and additional 2 hours with C 12 FDG substrate (green). Fluorescence imaging data analysis and staining of nucleus and cytoplasm as described above. Each image is representative of at least 5 fields of duplicate wells. Magnification 60×.
    Rabbit Anti β Galactosidase, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore rabbit anti β galactosidase antibody
    A) Immunofluorescent analyses of liver sections of Nrf2 -reporter mice orally treated with either DMSO, 30 μmol/kg CDDO-Im or 7 mg/kg WA for 3 hours by <t>anti-β-galactosidase</t> antibody. Nrf2-LacZ fusion protein expression and nuclei were stained by Alexa Fluor 546 and DAPI, respectively. Scale bars=10 μm. Transcript induction of B) Nqo1 C) Ho-1 in WT and Nrf2- disrupted MEF treated with graded doses of WA (0–1 μM) for 20 hours. D) Immunoblot of Nrf2 protein in the nuclear fraction of WT MEF treated with pharmacologic agents; DMSO, 25 nM CDDO-Im after 1 hour, 1 μM WA after 1 hour, 25 nM CDDO-Im after 3 hours, 1 μM WA after 3 hours. Lamin B was used as the loading control. E) Quantification of densitometry of 3 replicate western blots representing C. F) Nqo1 transcript induction by 1 μM WA, 25 nM CDDO-Im or 10 μM sulforaphane in WT MEF 20 hours post treatment. G) Relative luciferase activity in MCF7 ARE-luciferase reporter cells treated with 1 μM WA, 25 nM CDDO-Im or 10 μM sulforaphane for 20 hours. Gapdh was used as the normalization control for RT-PCR. All values are mean ± SEM (n=3). *p
    Rabbit Anti β Galactosidase Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Millipore anti cry1
    TCP2 is a light-regulated protein. (A) Immunoblots showing the effect of light on the expression of Myc-TCP2 protein in transgenic lines expressing 35S:: Myc-TCP2 . Plants were grown in long days (LDs; 16h/8h) for 3 weeks, moved to darkness for 16h, then exposed to 20 μmol m −2 s −1 far-red light, 18 μmol m −2 s −1 red light, or 20 μmol m −2 s −1 blue light for the indicated time. Samples were fractionated by SDS–PAGE, blotted to a nitrocellulose membrane, and probed with the anti-Myc antibody; HSP90 protein or the amounts of Rubisco (Ponceau) indicate relative loading of samples. (B) TCP2 protein accumulation increased as the blue light fluence rate increased. Plants grown in LDs (16h/8h) for 3 weeks were transferred to darkness for 16h, and then exposed to blue light of the indicated fluence rate (B 3, 3 μmol m −2 s −1 , B 10, 10 μmol m −2 s −1 , B 60, 60 μmol m −2 s −1 ) and time. Samples were analyzed by immunoblotting. (C) The immunoblots shown in (A and B) was quantified (Image J), calculated by the formula [(MYC-TCP2) t /Ponceau) t ]/ [(Myc-TCP2) 0 /Ponceau 0 ] or [(MYC-TCP2) t /(HSP90) t ]/[(Myc-TCP2) 0 /HSP90 0 ]; (Myc-TCP2) 0 and (MYC-TCP2) t are the signal at time zero and time indicated of Myc-TCP2, respectively; Ponceau 0 and Ponceau t are thesignal at time zero and time indicated of Rubisco protein, respectively; HSP90 0 and HSP90 t are the signal at time zero and time indicated of HSP90, respectively. (D) TCP2 protein accumulated in blue light. Three-week-old TCP2 /WT and TCP2 / <t>cry1</t> transgenic plants grown in a LD (16h/8h) photoperiod were moved to red light (20 μmol m −2 s −1 ) and far-red light (20 μmol m −2 s −1 ) for 16h, and then transferred to blue light (35 μmol m −2 s −1 ) for the indicated time before sample collection. (E) Analysis of TCP2 protein expression level change when transferred from blue light to darkness, far-red light, and red light. Three-week-old LD- (16h/8h) grown plants were transferred to blue light (35 μmol m −2 s −1 ) for 16h, and then transferred to darkness, far-red light (20 μmol m −2 s −1 ), and red light (20 μmol m −2 s −1 ) for the indicated time before sample collection. (F) Analysis of TCP2 protein degradation mechanism. Plants were grown in white light in LDs (16h/8h) for 3 weeks, moved to blue light (35 μmol m −2 s −1 ) for 16h, and leaves were excised and incubated in the presence of MG132 solution (50 μmol l –1 ) or in its absence (0.1% DMSO) for 4h, after which they were transferred to darkness for the indicated time before sample collection. (G) The immunoblots shown in (D–F) were quantified (Image J). The formula used for calculation is the same as that in (C). (This figure is available in colour at JXB online.)
    Anti Cry1, supplied by Millipore, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    AvidinOX-anchored bTrast and bPert induce apoptosis and senescence of SKBR3 cells ( A ) Cells, with or without AvidinOX (AvOX), were stained with BrdU after 72-hour exposure to bTrast (1 μg/mL) or bPert (0.5 μg/mL) and then analyzed by flow cytometry. The Sub-G1 fraction of total cell population in grey. Representative results of one out of three independent samples. ( B ) HCS fluorescence imaging of cells cultivated 48 hours with bTrast (1 μg/mL) or bPert (0.5 μg/mL) then washed, fixed and stained with rabbit anti-cleaved caspase 3, followed by phycoherythrin-conjugated goat anti-Rabbit IgG (green). Draq5 dye staining of nucleus and cytoplasm (grey). Each image is representative of at least 5 fields of duplicate wells. Magnification 60x. Data are from one representative experiment out of two. ( C ) Caspase 3/7 activity in cells treated 72 hours with Pert or bPert at indicated concentrations, measured by Caspase-Glo 3/7 Assay. Data are expressed as fold change of activity compared to control cells and are the average of four replicates (± SE). Staurosporine (0.1 μg/mL) was included as positive control. Mann-Whitney's test: *** p ≤ 0.001, ** p ≤ 0.01 and * p ≤ 0.05 vs AvOX. ( D ) Cells were cultivated 6 days with antibodies as in B, and additional 3 days without antibodies. Cells in left panel were stained with rabbit anti-β-Galactosidase antibody followed by FITC-conjugated goat anti-rabbit IgG (violet). Cells in right panel were treated 1 hour with bafilomycin A1 and additional 2 hours with C 12 FDG substrate (green). Fluorescence imaging data analysis and staining of nucleus and cytoplasm as described above. Each image is representative of at least 5 fields of duplicate wells. Magnification 60×.

    Journal: Oncotarget

    Article Title: AvidinOX-anchored biotinylated trastuzumab and pertuzumab induce down-modulation of ErbB2 and tumor cell death at concentrations order of magnitude lower than not-anchored antibodies

    doi: 10.18632/oncotarget.15145

    Figure Lengend Snippet: AvidinOX-anchored bTrast and bPert induce apoptosis and senescence of SKBR3 cells ( A ) Cells, with or without AvidinOX (AvOX), were stained with BrdU after 72-hour exposure to bTrast (1 μg/mL) or bPert (0.5 μg/mL) and then analyzed by flow cytometry. The Sub-G1 fraction of total cell population in grey. Representative results of one out of three independent samples. ( B ) HCS fluorescence imaging of cells cultivated 48 hours with bTrast (1 μg/mL) or bPert (0.5 μg/mL) then washed, fixed and stained with rabbit anti-cleaved caspase 3, followed by phycoherythrin-conjugated goat anti-Rabbit IgG (green). Draq5 dye staining of nucleus and cytoplasm (grey). Each image is representative of at least 5 fields of duplicate wells. Magnification 60x. Data are from one representative experiment out of two. ( C ) Caspase 3/7 activity in cells treated 72 hours with Pert or bPert at indicated concentrations, measured by Caspase-Glo 3/7 Assay. Data are expressed as fold change of activity compared to control cells and are the average of four replicates (± SE). Staurosporine (0.1 μg/mL) was included as positive control. Mann-Whitney's test: *** p ≤ 0.001, ** p ≤ 0.01 and * p ≤ 0.05 vs AvOX. ( D ) Cells were cultivated 6 days with antibodies as in B, and additional 3 days without antibodies. Cells in left panel were stained with rabbit anti-β-Galactosidase antibody followed by FITC-conjugated goat anti-rabbit IgG (violet). Cells in right panel were treated 1 hour with bafilomycin A1 and additional 2 hours with C 12 FDG substrate (green). Fluorescence imaging data analysis and staining of nucleus and cytoplasm as described above. Each image is representative of at least 5 fields of duplicate wells. Magnification 60×.

    Article Snippet: Expression of protein targets after cell fixation, permeabilization and blocking as described above, was evaluated by adding the following specific primary antibodies: rabbit anti-cleaved caspase-3 (Cell Signaling), rabbit anti-β-Galactosidase (Millipore), rabbit anti-Bcl-XL (Cell Signaling), rabbit anti-p-Bcl-XL (Invitrogen), rabbit anti-Bcl-2, (abcam), mouse anti-cIAP2 (R & D Systems), rabbit anti-phospho-PERK (Santa Cruz Biot.), rabbit anti-ATF4 (proteintech), rabbit anti-ATF-6α (Santa Cruz Biot).

    Techniques: Staining, Flow Cytometry, Cytometry, Fluorescence, Imaging, Activity Assay, Caspase-Glo Assay, Positive Control, MANN-WHITNEY

    A) Immunofluorescent analyses of liver sections of Nrf2 -reporter mice orally treated with either DMSO, 30 μmol/kg CDDO-Im or 7 mg/kg WA for 3 hours by anti-β-galactosidase antibody. Nrf2-LacZ fusion protein expression and nuclei were stained by Alexa Fluor 546 and DAPI, respectively. Scale bars=10 μm. Transcript induction of B) Nqo1 C) Ho-1 in WT and Nrf2- disrupted MEF treated with graded doses of WA (0–1 μM) for 20 hours. D) Immunoblot of Nrf2 protein in the nuclear fraction of WT MEF treated with pharmacologic agents; DMSO, 25 nM CDDO-Im after 1 hour, 1 μM WA after 1 hour, 25 nM CDDO-Im after 3 hours, 1 μM WA after 3 hours. Lamin B was used as the loading control. E) Quantification of densitometry of 3 replicate western blots representing C. F) Nqo1 transcript induction by 1 μM WA, 25 nM CDDO-Im or 10 μM sulforaphane in WT MEF 20 hours post treatment. G) Relative luciferase activity in MCF7 ARE-luciferase reporter cells treated with 1 μM WA, 25 nM CDDO-Im or 10 μM sulforaphane for 20 hours. Gapdh was used as the normalization control for RT-PCR. All values are mean ± SEM (n=3). *p

    Journal: Free radical biology & medicine

    Article Title: Withaferin A induces Nrf2-dependent protection against liver injury: role of Keap1-independent mechanisms

    doi: 10.1016/j.freeradbiomed.2016.10.003

    Figure Lengend Snippet: A) Immunofluorescent analyses of liver sections of Nrf2 -reporter mice orally treated with either DMSO, 30 μmol/kg CDDO-Im or 7 mg/kg WA for 3 hours by anti-β-galactosidase antibody. Nrf2-LacZ fusion protein expression and nuclei were stained by Alexa Fluor 546 and DAPI, respectively. Scale bars=10 μm. Transcript induction of B) Nqo1 C) Ho-1 in WT and Nrf2- disrupted MEF treated with graded doses of WA (0–1 μM) for 20 hours. D) Immunoblot of Nrf2 protein in the nuclear fraction of WT MEF treated with pharmacologic agents; DMSO, 25 nM CDDO-Im after 1 hour, 1 μM WA after 1 hour, 25 nM CDDO-Im after 3 hours, 1 μM WA after 3 hours. Lamin B was used as the loading control. E) Quantification of densitometry of 3 replicate western blots representing C. F) Nqo1 transcript induction by 1 μM WA, 25 nM CDDO-Im or 10 μM sulforaphane in WT MEF 20 hours post treatment. G) Relative luciferase activity in MCF7 ARE-luciferase reporter cells treated with 1 μM WA, 25 nM CDDO-Im or 10 μM sulforaphane for 20 hours. Gapdh was used as the normalization control for RT-PCR. All values are mean ± SEM (n=3). *p

    Article Snippet: For fluorescence staining, liver sections were stained with rabbit anti-β-galactosidase antibody (Millipore, Billerica MA; AB986) at a 1:150 dilution.

    Techniques: Mouse Assay, Expressing, Staining, Western Blot, Luciferase, Activity Assay, Reverse Transcription Polymerase Chain Reaction

    TCP2 is a light-regulated protein. (A) Immunoblots showing the effect of light on the expression of Myc-TCP2 protein in transgenic lines expressing 35S:: Myc-TCP2 . Plants were grown in long days (LDs; 16h/8h) for 3 weeks, moved to darkness for 16h, then exposed to 20 μmol m −2 s −1 far-red light, 18 μmol m −2 s −1 red light, or 20 μmol m −2 s −1 blue light for the indicated time. Samples were fractionated by SDS–PAGE, blotted to a nitrocellulose membrane, and probed with the anti-Myc antibody; HSP90 protein or the amounts of Rubisco (Ponceau) indicate relative loading of samples. (B) TCP2 protein accumulation increased as the blue light fluence rate increased. Plants grown in LDs (16h/8h) for 3 weeks were transferred to darkness for 16h, and then exposed to blue light of the indicated fluence rate (B 3, 3 μmol m −2 s −1 , B 10, 10 μmol m −2 s −1 , B 60, 60 μmol m −2 s −1 ) and time. Samples were analyzed by immunoblotting. (C) The immunoblots shown in (A and B) was quantified (Image J), calculated by the formula [(MYC-TCP2) t /Ponceau) t ]/ [(Myc-TCP2) 0 /Ponceau 0 ] or [(MYC-TCP2) t /(HSP90) t ]/[(Myc-TCP2) 0 /HSP90 0 ]; (Myc-TCP2) 0 and (MYC-TCP2) t are the signal at time zero and time indicated of Myc-TCP2, respectively; Ponceau 0 and Ponceau t are thesignal at time zero and time indicated of Rubisco protein, respectively; HSP90 0 and HSP90 t are the signal at time zero and time indicated of HSP90, respectively. (D) TCP2 protein accumulated in blue light. Three-week-old TCP2 /WT and TCP2 / cry1 transgenic plants grown in a LD (16h/8h) photoperiod were moved to red light (20 μmol m −2 s −1 ) and far-red light (20 μmol m −2 s −1 ) for 16h, and then transferred to blue light (35 μmol m −2 s −1 ) for the indicated time before sample collection. (E) Analysis of TCP2 protein expression level change when transferred from blue light to darkness, far-red light, and red light. Three-week-old LD- (16h/8h) grown plants were transferred to blue light (35 μmol m −2 s −1 ) for 16h, and then transferred to darkness, far-red light (20 μmol m −2 s −1 ), and red light (20 μmol m −2 s −1 ) for the indicated time before sample collection. (F) Analysis of TCP2 protein degradation mechanism. Plants were grown in white light in LDs (16h/8h) for 3 weeks, moved to blue light (35 μmol m −2 s −1 ) for 16h, and leaves were excised and incubated in the presence of MG132 solution (50 μmol l –1 ) or in its absence (0.1% DMSO) for 4h, after which they were transferred to darkness for the indicated time before sample collection. (G) The immunoblots shown in (D–F) were quantified (Image J). The formula used for calculation is the same as that in (C). (This figure is available in colour at JXB online.)

    Journal: Journal of Experimental Botany

    Article Title: TCP2 positively regulates HY5/HYH and photomorphogenesis in Arabidopsis

    doi: 10.1093/jxb/erv495

    Figure Lengend Snippet: TCP2 is a light-regulated protein. (A) Immunoblots showing the effect of light on the expression of Myc-TCP2 protein in transgenic lines expressing 35S:: Myc-TCP2 . Plants were grown in long days (LDs; 16h/8h) for 3 weeks, moved to darkness for 16h, then exposed to 20 μmol m −2 s −1 far-red light, 18 μmol m −2 s −1 red light, or 20 μmol m −2 s −1 blue light for the indicated time. Samples were fractionated by SDS–PAGE, blotted to a nitrocellulose membrane, and probed with the anti-Myc antibody; HSP90 protein or the amounts of Rubisco (Ponceau) indicate relative loading of samples. (B) TCP2 protein accumulation increased as the blue light fluence rate increased. Plants grown in LDs (16h/8h) for 3 weeks were transferred to darkness for 16h, and then exposed to blue light of the indicated fluence rate (B 3, 3 μmol m −2 s −1 , B 10, 10 μmol m −2 s −1 , B 60, 60 μmol m −2 s −1 ) and time. Samples were analyzed by immunoblotting. (C) The immunoblots shown in (A and B) was quantified (Image J), calculated by the formula [(MYC-TCP2) t /Ponceau) t ]/ [(Myc-TCP2) 0 /Ponceau 0 ] or [(MYC-TCP2) t /(HSP90) t ]/[(Myc-TCP2) 0 /HSP90 0 ]; (Myc-TCP2) 0 and (MYC-TCP2) t are the signal at time zero and time indicated of Myc-TCP2, respectively; Ponceau 0 and Ponceau t are thesignal at time zero and time indicated of Rubisco protein, respectively; HSP90 0 and HSP90 t are the signal at time zero and time indicated of HSP90, respectively. (D) TCP2 protein accumulated in blue light. Three-week-old TCP2 /WT and TCP2 / cry1 transgenic plants grown in a LD (16h/8h) photoperiod were moved to red light (20 μmol m −2 s −1 ) and far-red light (20 μmol m −2 s −1 ) for 16h, and then transferred to blue light (35 μmol m −2 s −1 ) for the indicated time before sample collection. (E) Analysis of TCP2 protein expression level change when transferred from blue light to darkness, far-red light, and red light. Three-week-old LD- (16h/8h) grown plants were transferred to blue light (35 μmol m −2 s −1 ) for 16h, and then transferred to darkness, far-red light (20 μmol m −2 s −1 ), and red light (20 μmol m −2 s −1 ) for the indicated time before sample collection. (F) Analysis of TCP2 protein degradation mechanism. Plants were grown in white light in LDs (16h/8h) for 3 weeks, moved to blue light (35 μmol m −2 s −1 ) for 16h, and leaves were excised and incubated in the presence of MG132 solution (50 μmol l –1 ) or in its absence (0.1% DMSO) for 4h, after which they were transferred to darkness for the indicated time before sample collection. (G) The immunoblots shown in (D–F) were quantified (Image J). The formula used for calculation is the same as that in (C). (This figure is available in colour at JXB online.)

    Article Snippet: Total supernatants were fractioned by 10% SDS–PAGE, and the membranes were probed by anti-CRY1, and then stripped, and re-probed by anti-Myc antibody (Millipore; Cat #05-724).

    Techniques: Western Blot, Expressing, Transgenic Assay, SDS Page, Incubation

    Analysis of the blue light response of the interaction between CRY1 and TCP2 in yeast cells and plant cells. (A) β-Galactosidase (β-gal) assay showing the interaction of CRY1 with TCP2 in the yeast cells treated with blue light (B 40, 40 μmol m −2 s −1 ), red light (R 18, 18 μmol m −2 s −1 ), or darkness (D) for 4h. (B) β-Gal assay showing the interaction of CRY1 with TCP2 in the yeast cells in response to blue light [10 μmol m −2 s −1 (B 10), 25 μmol m −2 s −1 (B 25), and 40 μmol m −2 s −1 (B 40)], red light (R 18, 18 μmol m −2 s −1 ), or darkness (D) for the indicated times. (C) Slopes of linear regression curves of different light fluence rates as shown in (B). The error bar shows the SD for triplicae samples at the same fluence rate. (D) BiFC assay showing the CRY1–TCP2 interaction in Nicotiana benthamiana . Plants were grown in long days (LDs; 16h/8h) for 4 weeks and the young leaves were infiltrated with an Agrobacterium mix carrying the plasmids nYFP- TCP2 and cCFP- CRY1 independently, incubated in the dark for 12h, and then transferred to white light for 48h, before fluorescence microscope assay, transferred to blue light (50 μmol m −2 s −1 ) and to darkness for 4h. DAPI, nuclear fluorescence; YFP, yellow fluorescent protein fluorescence; Merge, merge of DAPI and YFP; scale bar=2 μm. (E) The percentage of N. benthamiana leaf cells that showed the BiFC fluorescence signals in (D) were counted. For each sample at least 200 cells were counted; P =0.003. (F) A co-immunoprecipitation assay showing the CRY1–TCP2 interaction in blue light in Arabidopsis with the indicated treatment times. The transgenic plants expressing 35S ::Myc-TCP2 /WT were grown in LDs (16h/ 8h) for 3 weeks. Plants were transferred to darkness for 16h, and the leaves were excised, incubated in MG132 (50 μmol l –1 ) under darkness for 4h, and then exposed to blue light (B, 35 μmol m −2 s −1 ) for the time indicated (B 0, 0min; B 30, 30min; B 240, 240min). Total protein (Input) or immunoprecipitation (IP) products using agarose conjugated with anti-Myc antibody (α-Myc) were probed by anti-Myc antibody, stripped, and re-probed by the anti-CRY1 antibody (α-CRY1). (G) Schematic representation depicting the domains of CRY1 and TCP2 that are required for CRY1–TCP2 interaction (shaded area). (This figure is available in colour at JXB online.)

    Journal: Journal of Experimental Botany

    Article Title: TCP2 positively regulates HY5/HYH and photomorphogenesis in Arabidopsis

    doi: 10.1093/jxb/erv495

    Figure Lengend Snippet: Analysis of the blue light response of the interaction between CRY1 and TCP2 in yeast cells and plant cells. (A) β-Galactosidase (β-gal) assay showing the interaction of CRY1 with TCP2 in the yeast cells treated with blue light (B 40, 40 μmol m −2 s −1 ), red light (R 18, 18 μmol m −2 s −1 ), or darkness (D) for 4h. (B) β-Gal assay showing the interaction of CRY1 with TCP2 in the yeast cells in response to blue light [10 μmol m −2 s −1 (B 10), 25 μmol m −2 s −1 (B 25), and 40 μmol m −2 s −1 (B 40)], red light (R 18, 18 μmol m −2 s −1 ), or darkness (D) for the indicated times. (C) Slopes of linear regression curves of different light fluence rates as shown in (B). The error bar shows the SD for triplicae samples at the same fluence rate. (D) BiFC assay showing the CRY1–TCP2 interaction in Nicotiana benthamiana . Plants were grown in long days (LDs; 16h/8h) for 4 weeks and the young leaves were infiltrated with an Agrobacterium mix carrying the plasmids nYFP- TCP2 and cCFP- CRY1 independently, incubated in the dark for 12h, and then transferred to white light for 48h, before fluorescence microscope assay, transferred to blue light (50 μmol m −2 s −1 ) and to darkness for 4h. DAPI, nuclear fluorescence; YFP, yellow fluorescent protein fluorescence; Merge, merge of DAPI and YFP; scale bar=2 μm. (E) The percentage of N. benthamiana leaf cells that showed the BiFC fluorescence signals in (D) were counted. For each sample at least 200 cells were counted; P =0.003. (F) A co-immunoprecipitation assay showing the CRY1–TCP2 interaction in blue light in Arabidopsis with the indicated treatment times. The transgenic plants expressing 35S ::Myc-TCP2 /WT were grown in LDs (16h/ 8h) for 3 weeks. Plants were transferred to darkness for 16h, and the leaves were excised, incubated in MG132 (50 μmol l –1 ) under darkness for 4h, and then exposed to blue light (B, 35 μmol m −2 s −1 ) for the time indicated (B 0, 0min; B 30, 30min; B 240, 240min). Total protein (Input) or immunoprecipitation (IP) products using agarose conjugated with anti-Myc antibody (α-Myc) were probed by anti-Myc antibody, stripped, and re-probed by the anti-CRY1 antibody (α-CRY1). (G) Schematic representation depicting the domains of CRY1 and TCP2 that are required for CRY1–TCP2 interaction (shaded area). (This figure is available in colour at JXB online.)

    Article Snippet: Total supernatants were fractioned by 10% SDS–PAGE, and the membranes were probed by anti-CRY1, and then stripped, and re-probed by anti-Myc antibody (Millipore; Cat #05-724).

    Techniques: β-Gal Assay, Bimolecular Fluorescence Complementation Assay, Incubation, Fluorescence, Microscopy, Co-Immunoprecipitation Assay, Transgenic Assay, Expressing, Immunoprecipitation

    TCP2 regulates the expression of genes downstream of the CRY1 signaling pathway and binds to the chromatins of HY5 and HYH to promote photomorphogensis. (A) mRNA expression of genes downstream of CRY1 in different TCP2 genotypes as indicated. Seedlings were grown in continuous white light for 10 d, and moved to darkness or remained in continuous white light for 33h before tissues were collected. (B) Diagrams depicting the promoter (arrow) and genome (white box, intron; striped box, exon) of HY5 and HYH . The black circles and asterisk indicate the position of potential TCP-binding sites (the asterisk indicates the position of published TCP Class II gene-binding sites; circles indicate the positions of sequences which are partly similar to the published binding sites). Different regions of the HY5 and HYH genomic DNA examined by ChIP-qPCR are indicated with short underlines (underlining of the b and c regions of HY5, and the c and d regions of HYH mark the DNA regions including sequences indicated by an asterisk, and other underlines mark DNA regions including sequences indicated only by circles). (C) ChIP-qPCR analysis of the indicated chromatin regions of HY5 and HYH in darkness and blue light (35 μmol m −2 s −1 ) of samples collected from the TCP2 -overexpressing transgenic line and the cry1 mutant background. Seedlings were grown in continuous white light for 10 d and then transferred to blue light or darkness for 2 d before sample collection. ChIP samples were prepared by the anti-Myc antibody and subjected to qPCR analysis. Results of ChIP-qPCR were quantified by normalization of the IP signal with the corresponding input signal. The SDs are shown ( n =3). RBU (relative binding unit)=PCR signal of the IP reaction/PCR signal of the mock reaction without antibody. (This figure is available in colour at JXB online.)

    Journal: Journal of Experimental Botany

    Article Title: TCP2 positively regulates HY5/HYH and photomorphogenesis in Arabidopsis

    doi: 10.1093/jxb/erv495

    Figure Lengend Snippet: TCP2 regulates the expression of genes downstream of the CRY1 signaling pathway and binds to the chromatins of HY5 and HYH to promote photomorphogensis. (A) mRNA expression of genes downstream of CRY1 in different TCP2 genotypes as indicated. Seedlings were grown in continuous white light for 10 d, and moved to darkness or remained in continuous white light for 33h before tissues were collected. (B) Diagrams depicting the promoter (arrow) and genome (white box, intron; striped box, exon) of HY5 and HYH . The black circles and asterisk indicate the position of potential TCP-binding sites (the asterisk indicates the position of published TCP Class II gene-binding sites; circles indicate the positions of sequences which are partly similar to the published binding sites). Different regions of the HY5 and HYH genomic DNA examined by ChIP-qPCR are indicated with short underlines (underlining of the b and c regions of HY5, and the c and d regions of HYH mark the DNA regions including sequences indicated by an asterisk, and other underlines mark DNA regions including sequences indicated only by circles). (C) ChIP-qPCR analysis of the indicated chromatin regions of HY5 and HYH in darkness and blue light (35 μmol m −2 s −1 ) of samples collected from the TCP2 -overexpressing transgenic line and the cry1 mutant background. Seedlings were grown in continuous white light for 10 d and then transferred to blue light or darkness for 2 d before sample collection. ChIP samples were prepared by the anti-Myc antibody and subjected to qPCR analysis. Results of ChIP-qPCR were quantified by normalization of the IP signal with the corresponding input signal. The SDs are shown ( n =3). RBU (relative binding unit)=PCR signal of the IP reaction/PCR signal of the mock reaction without antibody. (This figure is available in colour at JXB online.)

    Article Snippet: Total supernatants were fractioned by 10% SDS–PAGE, and the membranes were probed by anti-CRY1, and then stripped, and re-probed by anti-Myc antibody (Millipore; Cat #05-724).

    Techniques: Expressing, Binding Assay, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Transgenic Assay, Mutagenesis, Polymerase Chain Reaction

    Light response phenotype analysis of transgenic lines with altered TCP2 expression in different backgrounds. (A) Images of the hypocotyl phenotypes of 5-day-old cry1 , WT, and Myc-TCP2 /WT transgenic seedlings grown under different light (D, darkness; FR 20, far-red 20 μmol m −2 s −1 ; R 18, red light 18 μmol m −2 s −1 ; B 2, blue light 2 μmol m −2 s −1 ). Western blots show the overexpression of TCP2 in the WT. Error bars represent ±SD of three biological replicates. (B) Hypocotyl lengths of the genotypes indicated in (A) grown in the different light conditions as in (A) were measured and are shown. The P -values of the hypocotyl length difference between WT and transgenic lines were 0.006, 0.027, and 0.014 for TCP2/ WT (#13, # 19,#22) in B 2 (blue light 2 μmol m −2 s −1 ), respectively. (C) Images of hypocotyl phenotypes of 5-day-old cry1 and Myc-TCP2 / cry1 transgenic seedlings grown under different light (D, darkness; FR 20, far-red light 20 μmol m −2 s −1 ; R 18, red light 18 μmol m −2 s −1 ; B 2, blue light 2 μmol m −2 s −1 ). Western blot shows the overexpression of TCP2 in cry1 . (D) Hypocotyl lengths of the genotypes indicated in (A) grown in the different light conditions as in (C) were measured and are shown. Error bars represent ±SD of three biological replicates. The P -values of the hypocotyl length difference between cry1 mutant and transgenic lines were

    Journal: Journal of Experimental Botany

    Article Title: TCP2 positively regulates HY5/HYH and photomorphogenesis in Arabidopsis

    doi: 10.1093/jxb/erv495

    Figure Lengend Snippet: Light response phenotype analysis of transgenic lines with altered TCP2 expression in different backgrounds. (A) Images of the hypocotyl phenotypes of 5-day-old cry1 , WT, and Myc-TCP2 /WT transgenic seedlings grown under different light (D, darkness; FR 20, far-red 20 μmol m −2 s −1 ; R 18, red light 18 μmol m −2 s −1 ; B 2, blue light 2 μmol m −2 s −1 ). Western blots show the overexpression of TCP2 in the WT. Error bars represent ±SD of three biological replicates. (B) Hypocotyl lengths of the genotypes indicated in (A) grown in the different light conditions as in (A) were measured and are shown. The P -values of the hypocotyl length difference between WT and transgenic lines were 0.006, 0.027, and 0.014 for TCP2/ WT (#13, # 19,#22) in B 2 (blue light 2 μmol m −2 s −1 ), respectively. (C) Images of hypocotyl phenotypes of 5-day-old cry1 and Myc-TCP2 / cry1 transgenic seedlings grown under different light (D, darkness; FR 20, far-red light 20 μmol m −2 s −1 ; R 18, red light 18 μmol m −2 s −1 ; B 2, blue light 2 μmol m −2 s −1 ). Western blot shows the overexpression of TCP2 in cry1 . (D) Hypocotyl lengths of the genotypes indicated in (A) grown in the different light conditions as in (C) were measured and are shown. Error bars represent ±SD of three biological replicates. The P -values of the hypocotyl length difference between cry1 mutant and transgenic lines were

    Article Snippet: Total supernatants were fractioned by 10% SDS–PAGE, and the membranes were probed by anti-CRY1, and then stripped, and re-probed by anti-Myc antibody (Millipore; Cat #05-724).

    Techniques: Transgenic Assay, Expressing, Western Blot, Over Expression, Mutagenesis