trichostatin a  (Abcam)

 
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 81
    Name:
    Trichostatin A DMSO solution
    Description:

    Catalog Number:
    ab146598
    Price:
    None
    Buy from Supplier


    Structured Review

    Abcam trichostatin a
    Chromatin accessibility regulated STAT3 dimer-to-tetramer transition (cpCOMB analysis). ( a ) Chromatin organization in HeLa cells stained with Hoechst 33342 after no treatment, 18 h after addition of 400 nM of <t>trichostatin</t> A (loosened chromatin) and 30 min after addition of 5 μg ml −1 of actinomycin D (compacted chromatin). ( b ) Merged STAT3-GFP and STAT3-mCherry intensity images of control, trichostatin A- and actinomycin D-treated live HeLa cells. Scale bar, 5 μm. ( c ) STAT3-GFP and STAT3-mCherry fluorescence intensity within the plane in which the line scan was acquired. The nuclear boundary is indicated by the Hoechst 33342 stain. ( d ) Pseudocoloured brightness maps for STAT3-GFP in control, trichostatin A- and actinomycin D-treated HeLa cells. Dark green pixels represent monomers, light green pixels dimers and red pixels tetramer. ( e ) cpCOMB carpets in control, trichostatin A- and actinomycin D-treated live HeLa cells. White dashed circles highlight tetramer formation. CTYO, cytoplasm; Max, maximum; Min, minimum; NUC, nucleus.

    https://www.bioz.com/result/trichostatin a/product/Abcam
    Average 81 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    trichostatin a - by Bioz Stars, 2020-01
    81/100 stars

    Images

    1) Product Images from "Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness"

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness

    Journal: Nature Communications

    doi: 10.1038/ncomms11047

    Chromatin accessibility regulated STAT3 dimer-to-tetramer transition (cpCOMB analysis). ( a ) Chromatin organization in HeLa cells stained with Hoechst 33342 after no treatment, 18 h after addition of 400 nM of trichostatin A (loosened chromatin) and 30 min after addition of 5 μg ml −1 of actinomycin D (compacted chromatin). ( b ) Merged STAT3-GFP and STAT3-mCherry intensity images of control, trichostatin A- and actinomycin D-treated live HeLa cells. Scale bar, 5 μm. ( c ) STAT3-GFP and STAT3-mCherry fluorescence intensity within the plane in which the line scan was acquired. The nuclear boundary is indicated by the Hoechst 33342 stain. ( d ) Pseudocoloured brightness maps for STAT3-GFP in control, trichostatin A- and actinomycin D-treated HeLa cells. Dark green pixels represent monomers, light green pixels dimers and red pixels tetramer. ( e ) cpCOMB carpets in control, trichostatin A- and actinomycin D-treated live HeLa cells. White dashed circles highlight tetramer formation. CTYO, cytoplasm; Max, maximum; Min, minimum; NUC, nucleus.
    Figure Legend Snippet: Chromatin accessibility regulated STAT3 dimer-to-tetramer transition (cpCOMB analysis). ( a ) Chromatin organization in HeLa cells stained with Hoechst 33342 after no treatment, 18 h after addition of 400 nM of trichostatin A (loosened chromatin) and 30 min after addition of 5 μg ml −1 of actinomycin D (compacted chromatin). ( b ) Merged STAT3-GFP and STAT3-mCherry intensity images of control, trichostatin A- and actinomycin D-treated live HeLa cells. Scale bar, 5 μm. ( c ) STAT3-GFP and STAT3-mCherry fluorescence intensity within the plane in which the line scan was acquired. The nuclear boundary is indicated by the Hoechst 33342 stain. ( d ) Pseudocoloured brightness maps for STAT3-GFP in control, trichostatin A- and actinomycin D-treated HeLa cells. Dark green pixels represent monomers, light green pixels dimers and red pixels tetramer. ( e ) cpCOMB carpets in control, trichostatin A- and actinomycin D-treated live HeLa cells. White dashed circles highlight tetramer formation. CTYO, cytoplasm; Max, maximum; Min, minimum; NUC, nucleus.

    Techniques Used: Staining, Fluorescence

    2) Product Images from "Involvement of phosphorylated Apis mellifera CREB in gating a honeybee's behavioral response to an external stimulus"

    Article Title: Involvement of phosphorylated Apis mellifera CREB in gating a honeybee's behavioral response to an external stimulus

    Journal: Learning & Memory

    doi: 10.1101/lm.040964.115

    Inhibition of histone deacetylases enhances CS responsiveness. ( A ) Schematic representation of the behavioral experiments. Bees were injected with the histone deacetylase inhibitor Trichostatin A (TSA), the histone acetyl transferase inhibitor, Garcinol or the respective vehicles 2, 5, or 24 h prior to unpaired training ( B ) Percentage of bees responding to the CS during unpaired training after injection of Trichostatin A ( B ) or Garcinol ( C ). (*) Significant differences: P
    Figure Legend Snippet: Inhibition of histone deacetylases enhances CS responsiveness. ( A ) Schematic representation of the behavioral experiments. Bees were injected with the histone deacetylase inhibitor Trichostatin A (TSA), the histone acetyl transferase inhibitor, Garcinol or the respective vehicles 2, 5, or 24 h prior to unpaired training ( B ) Percentage of bees responding to the CS during unpaired training after injection of Trichostatin A ( B ) or Garcinol ( C ). (*) Significant differences: P

    Techniques Used: Inhibition, Injection, Histone Deacetylase Assay

    Related Articles

    Transfection:

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness
    Article Snippet: .. Loosening of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with trichostatin A (400 nM, Abcam) for 18 h. Compacting of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with actinomycin D (5 μg ml−1 , a concentration known to stop class III transcription, Sigma Aldrich) for 30 min. .. The compaction status of chromatin was assessed by staining the DNA of HeLa cells with Hoechst 33342 (1 μg ml−1 , Sigma Aldrich) 15 min before imaging.

    Selection:

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness
    Article Snippet: Loosening of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with trichostatin A (400 nM, Abcam) for 18 h. Compacting of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with actinomycin D (5 μg ml−1 , a concentration known to stop class III transcription, Sigma Aldrich) for 30 min. .. Under this selection criteria, variations in STAT3-mCherry expression levels had negligible impact on STAT3 oligomerization ( ).

    Concentration Assay:

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness
    Article Snippet: .. Loosening of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with trichostatin A (400 nM, Abcam) for 18 h. Compacting of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with actinomycin D (5 μg ml−1 , a concentration known to stop class III transcription, Sigma Aldrich) for 30 min. .. The compaction status of chromatin was assessed by staining the DNA of HeLa cells with Hoechst 33342 (1 μg ml−1 , Sigma Aldrich) 15 min before imaging.

    Mutagenesis:

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness
    Article Snippet: Stimulation of wild-type and mutant STAT3 activity was carried out by treating the HeLa cells with oncostatin M (10 nM, Sigma Aldrich) for 15 min. STAT3 activation was assessed by the degree of nuclear accumulation. .. Loosening of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with trichostatin A (400 nM, Abcam) for 18 h. Compacting of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with actinomycin D (5 μg ml−1 , a concentration known to stop class III transcription, Sigma Aldrich) for 30 min.

    Construct:

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness
    Article Snippet: Loosening of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with trichostatin A (400 nM, Abcam) for 18 h. Compacting of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with actinomycin D (5 μg ml−1 , a concentration known to stop class III transcription, Sigma Aldrich) for 30 min. .. In all line- and frame-scan experiments, cells exhibiting a low STAT3 fluorescent construct expression level were selected, as both the brightness and pCOMB methods are fluctuation-based analyses.

    Purification:

    Article Title: Selective in vivo metabolic cell-labeling-mediated cancer targeting
    Article Snippet: Trichostatin A was purchased from Abcam (Cambridge, UK). .. Anhydrous dichloromethane (DCM), hexane, ethyl acetate, diethyl ether, tetrahydrofuran (THF) and dimethylformamide (DMF) were purified by passing them through alumina columns and kept anhydrous by storing them in the presence of molecular sieves.

    Activation Assay:

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness
    Article Snippet: Stimulation of wild-type and mutant STAT3 activity was carried out by treating the HeLa cells with oncostatin M (10 nM, Sigma Aldrich) for 15 min. STAT3 activation was assessed by the degree of nuclear accumulation. .. Loosening of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with trichostatin A (400 nM, Abcam) for 18 h. Compacting of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with actinomycin D (5 μg ml−1 , a concentration known to stop class III transcription, Sigma Aldrich) for 30 min.

    HAT Assay:

    Article Title: Involvement of phosphorylated Apis mellifera CREB in gating a honeybee's behavioral response to an external stimulus
    Article Snippet: To explore the effect of transcription-, HAT-, and HDAC-inhibitors on the behavior of bees during and after training, drugs or their solvents were systemically injection into the flight muscle ( ). .. To examine the role of histone acetylation, bees were injected 2, 5, or 24 h prior to the unpaired training with 6 mM Garcinol (Abcam; dissolved in 100% DMSO), 1.65 mM Trichostatin A (TSA, Abcam; dissolved in 20% DMSO in PBS) or the appropriate solvent as control.

    Activity Assay:

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness
    Article Snippet: Stimulation of wild-type and mutant STAT3 activity was carried out by treating the HeLa cells with oncostatin M (10 nM, Sigma Aldrich) for 15 min. STAT3 activation was assessed by the degree of nuclear accumulation. .. Loosening of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with trichostatin A (400 nM, Abcam) for 18 h. Compacting of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with actinomycin D (5 μg ml−1 , a concentration known to stop class III transcription, Sigma Aldrich) for 30 min.

    Imaging:

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness
    Article Snippet: Loosening of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with trichostatin A (400 nM, Abcam) for 18 h. Compacting of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with actinomycin D (5 μg ml−1 , a concentration known to stop class III transcription, Sigma Aldrich) for 30 min. .. The compaction status of chromatin was assessed by staining the DNA of HeLa cells with Hoechst 33342 (1 μg ml−1 , Sigma Aldrich) 15 min before imaging.

    Expressing:

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness
    Article Snippet: Loosening of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with trichostatin A (400 nM, Abcam) for 18 h. Compacting of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with actinomycin D (5 μg ml−1 , a concentration known to stop class III transcription, Sigma Aldrich) for 30 min. .. In all line- and frame-scan experiments, cells exhibiting a low STAT3 fluorescent construct expression level were selected, as both the brightness and pCOMB methods are fluctuation-based analyses.

    Modification:

    Article Title: Selective in vivo metabolic cell-labeling-mediated cancer targeting
    Article Snippet: Trichostatin A was purchased from Abcam (Cambridge, UK). .. Phosphate-buffered saline (PBS) and Dulbecco’s Modified Eagle Medium (DMEM) were obtained from Invitrogen (Carlsbad, CA, USA).

    Staining:

    Article Title: Quantifying the dynamics of the oligomeric transcription factor STAT3 by pair correlation of molecular brightness
    Article Snippet: Loosening of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with trichostatin A (400 nM, Abcam) for 18 h. Compacting of chromatin was carried out by treating the HeLa cells transiently transfected with STAT3-GFP and STAT3-mCherry, with actinomycin D (5 μg ml−1 , a concentration known to stop class III transcription, Sigma Aldrich) for 30 min. .. The compaction status of chromatin was assessed by staining the DNA of HeLa cells with Hoechst 33342 (1 μg ml−1 , Sigma Aldrich) 15 min before imaging.

    Injection:

    Article Title: Involvement of phosphorylated Apis mellifera CREB in gating a honeybee's behavioral response to an external stimulus
    Article Snippet: .. To examine the role of histone acetylation, bees were injected 2, 5, or 24 h prior to the unpaired training with 6 mM Garcinol (Abcam; dissolved in 100% DMSO), 1.65 mM Trichostatin A (TSA, Abcam; dissolved in 20% DMSO in PBS) or the appropriate solvent as control. ..

    Activated Clotting Time Assay:

    Article Title: Involvement of phosphorylated Apis mellifera CREB in gating a honeybee's behavioral response to an external stimulus
    Article Snippet: For the investigation of the transcriptional dependence of memories, bees were injected 90 min prior to the paired or unpaired training with 1.5 mM actinomycin D (Act D, Sigma-Aldrich) or the solvent PBS (137 mM NaCl; 2,7 mM KCl; 10,1 mM; Na2 HPO4 ; 1,8 mM KH2 PO4 , pH 7.2). .. To examine the role of histone acetylation, bees were injected 2, 5, or 24 h prior to the unpaired training with 6 mM Garcinol (Abcam; dissolved in 100% DMSO), 1.65 mM Trichostatin A (TSA, Abcam; dissolved in 20% DMSO in PBS) or the appropriate solvent as control.

    Molecular Weight:

    Article Title: Selective in vivo metabolic cell-labeling-mediated cancer targeting
    Article Snippet: HOOC–PEG–NH2 (1 k molecular weight) was purchased from Nanocs Inc. (New York, NY, USA). .. Trichostatin A was purchased from Abcam (Cambridge, UK).

    Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 90
    Abcam donkey anti mouse alexa fluor 488
    Donkey Anti Mouse Alexa Fluor 488, supplied by Abcam, used in various techniques. Bioz Stars score: 90/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/donkey anti mouse alexa fluor 488/product/Abcam
    Average 90 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    donkey anti mouse alexa fluor 488 - by Bioz Stars, 2020-01
    90/100 stars
      Buy from Supplier

    86
    Abcam mouse lamin a c
    Immunogold-labeling of A- and B-type lamins Nuclei on EM-grids were treated with α-lamin A/C or α-lamin B1 antibodies and labeled with 6 nm gold conjugated protein A prior vitrification and cryo-ET analysis. Control samples were treated with protein A conjugate only. (a) Projection views of nuclei (contoured by the light-grey line) display the distribution of gold conjugate. Scale bar, 200 nm. (b) Zoomed-in images show 9 nm thick xy-slices through reconstructed volumes of the respective immunogold-labeled nuclei. A- and B-type filaments are labeled with gold conjugate as indicated (red circles). Scale bar, 100 nm. (c) Gallery view of immunogold-labeled, segmented and skeletonized filaments from 40 nm thick sub-volumes. Green dots indicate immunogold-labeled lamin A and red dots lamin B1 in nine sub-volumes each, locating both within sparsely and densely packed regions. Scale bar, 200 nm. (d) Box plot shows the immunogold-labeling density of lamin A/C and lamin B1 per μm 2  (+/− SD) from volumes shown in (c) (white line represents the median and black dot the average number of gold particles).
    Mouse Lamin A C, supplied by Abcam, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse lamin a c/product/Abcam
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse lamin a c - by Bioz Stars, 2020-01
    86/100 stars
      Buy from Supplier

    85
    Abcam anti tryptase antibody
    Immunogold-labeling of A- and B-type lamins Nuclei on EM-grids were treated with α-lamin A/C or α-lamin B1 antibodies and labeled with 6 nm gold conjugated protein A prior vitrification and cryo-ET analysis. Control samples were treated with protein A conjugate only. (a) Projection views of nuclei (contoured by the light-grey line) display the distribution of gold conjugate. Scale bar, 200 nm. (b) Zoomed-in images show 9 nm thick xy-slices through reconstructed volumes of the respective immunogold-labeled nuclei. A- and B-type filaments are labeled with gold conjugate as indicated (red circles). Scale bar, 100 nm. (c) Gallery view of immunogold-labeled, segmented and skeletonized filaments from 40 nm thick sub-volumes. Green dots indicate immunogold-labeled lamin A and red dots lamin B1 in nine sub-volumes each, locating both within sparsely and densely packed regions. Scale bar, 200 nm. (d) Box plot shows the immunogold-labeling density of lamin A/C and lamin B1 per μm 2  (+/− SD) from volumes shown in (c) (white line represents the median and black dot the average number of gold particles).
    Anti Tryptase Antibody, supplied by Abcam, used in various techniques. Bioz Stars score: 85/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti tryptase antibody/product/Abcam
    Average 85 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    anti tryptase antibody - by Bioz Stars, 2020-01
    85/100 stars
      Buy from Supplier

    Image Search Results


    Immunogold-labeling of A- and B-type lamins Nuclei on EM-grids were treated with α-lamin A/C or α-lamin B1 antibodies and labeled with 6 nm gold conjugated protein A prior vitrification and cryo-ET analysis. Control samples were treated with protein A conjugate only. (a) Projection views of nuclei (contoured by the light-grey line) display the distribution of gold conjugate. Scale bar, 200 nm. (b) Zoomed-in images show 9 nm thick xy-slices through reconstructed volumes of the respective immunogold-labeled nuclei. A- and B-type filaments are labeled with gold conjugate as indicated (red circles). Scale bar, 100 nm. (c) Gallery view of immunogold-labeled, segmented and skeletonized filaments from 40 nm thick sub-volumes. Green dots indicate immunogold-labeled lamin A and red dots lamin B1 in nine sub-volumes each, locating both within sparsely and densely packed regions. Scale bar, 200 nm. (d) Box plot shows the immunogold-labeling density of lamin A/C and lamin B1 per μm 2  (+/− SD) from volumes shown in (c) (white line represents the median and black dot the average number of gold particles).

    Journal: Nature

    Article Title: The molecular architecture of lamins in somatic cells

    doi: 10.1038/nature21382

    Figure Lengend Snippet: Immunogold-labeling of A- and B-type lamins Nuclei on EM-grids were treated with α-lamin A/C or α-lamin B1 antibodies and labeled with 6 nm gold conjugated protein A prior vitrification and cryo-ET analysis. Control samples were treated with protein A conjugate only. (a) Projection views of nuclei (contoured by the light-grey line) display the distribution of gold conjugate. Scale bar, 200 nm. (b) Zoomed-in images show 9 nm thick xy-slices through reconstructed volumes of the respective immunogold-labeled nuclei. A- and B-type filaments are labeled with gold conjugate as indicated (red circles). Scale bar, 100 nm. (c) Gallery view of immunogold-labeled, segmented and skeletonized filaments from 40 nm thick sub-volumes. Green dots indicate immunogold-labeled lamin A and red dots lamin B1 in nine sub-volumes each, locating both within sparsely and densely packed regions. Scale bar, 200 nm. (d) Box plot shows the immunogold-labeling density of lamin A/C and lamin B1 per μm 2 (+/− SD) from volumes shown in (c) (white line represents the median and black dot the average number of gold particles).

    Article Snippet: For immunogold-labeling experiments, rabbit monoclonal antibody directed against the rod domain of mouse lamin A/C (peptide between aa 200–300) (EP4520, ab133256) was purchased from Abcam.

    Techniques: Labeling

    Sample preparation of vimentin deficient MEFs for Cryo-ET and 3D-SIM (a) 3D-SIM images of immunolabeled wt and Vim −/−  MEFs shows similar localization and expression of lamin A and lamin B1. Scale bar, 5 μm. (b) Western blot analyses of the indicated MEF cell lines show unchanged expression levels of each of the lamin isoforms and the retention of the lamins following the shRNAi-mediated knockdown of vimentin. (c) 3D-SIM images of pre-permeabilized, nuclease treated and immunolabeled MEFs shows similar localization and expression of lamin A and lamin B1 compared to untreated cells (a). Scale bar, 1 μm. (d) The localization of the lamina associated proteins (LAPs), emerin and Lap2β, exhibit high similarities between pre-permeabilized and nuclease treated (+) and untreated (−) cells, as indicated by 3D-SIM analysis. Scale bar, 1 μm. (e) Illustration of the cryo-ET sample preparation procedure.

    Journal: Nature

    Article Title: The molecular architecture of lamins in somatic cells

    doi: 10.1038/nature21382

    Figure Lengend Snippet: Sample preparation of vimentin deficient MEFs for Cryo-ET and 3D-SIM (a) 3D-SIM images of immunolabeled wt and Vim −/− MEFs shows similar localization and expression of lamin A and lamin B1. Scale bar, 5 μm. (b) Western blot analyses of the indicated MEF cell lines show unchanged expression levels of each of the lamin isoforms and the retention of the lamins following the shRNAi-mediated knockdown of vimentin. (c) 3D-SIM images of pre-permeabilized, nuclease treated and immunolabeled MEFs shows similar localization and expression of lamin A and lamin B1 compared to untreated cells (a). Scale bar, 1 μm. (d) The localization of the lamina associated proteins (LAPs), emerin and Lap2β, exhibit high similarities between pre-permeabilized and nuclease treated (+) and untreated (−) cells, as indicated by 3D-SIM analysis. Scale bar, 1 μm. (e) Illustration of the cryo-ET sample preparation procedure.

    Article Snippet: For immunogold-labeling experiments, rabbit monoclonal antibody directed against the rod domain of mouse lamin A/C (peptide between aa 200–300) (EP4520, ab133256) was purchased from Abcam.

    Techniques: Sample Prep, Immunolabeling, Expressing, Western Blot

    Structural analysis of  in vitro  assembled A- and B-type lamin paracrystals The final step in the  in vitro  assembly of assembled A- and B-type lamins results in the formation of paracrystals that display identical organization, corroborating that A- and B-type lamins assemble into similar structures (a) TEM analysis and comparison of negatively stained human lamin C  2 , B1 and B2 paracrystals show an identical striped pattern with 20 nm repeats. Scale bar, 20 nm. (b) Cryo-ET analysis of  in vitro  assembled lamin A shows the same 20 nm repeating pattern compared to the lamin isoforms shown in (a). Scale bar, 20 nm. (c) The model shows the 2D arrangement of lamin protofilaments within a paracrystal. The rod-like structure is shown in grey and the globular tail domains in red. (d) Averaged structure of cryo-tomograms of  in vitro  assembled lamin A as shown in (b) displays the striped pattern comprising the purported Ig-fold domains at distances of 20 nm, comparable to the spacing of the repeating pairs of globular domains in some structural classes from our  in situ  structural analysis (). The distance of the putative Ig-fold domains within the stripes of the paracrystals is 6 nm. Scale bar, 20 nm. Fig. 3a and b

    Journal: Nature

    Article Title: The molecular architecture of lamins in somatic cells

    doi: 10.1038/nature21382

    Figure Lengend Snippet: Structural analysis of in vitro assembled A- and B-type lamin paracrystals The final step in the in vitro assembly of assembled A- and B-type lamins results in the formation of paracrystals that display identical organization, corroborating that A- and B-type lamins assemble into similar structures (a) TEM analysis and comparison of negatively stained human lamin C 2 , B1 and B2 paracrystals show an identical striped pattern with 20 nm repeats. Scale bar, 20 nm. (b) Cryo-ET analysis of in vitro assembled lamin A shows the same 20 nm repeating pattern compared to the lamin isoforms shown in (a). Scale bar, 20 nm. (c) The model shows the 2D arrangement of lamin protofilaments within a paracrystal. The rod-like structure is shown in grey and the globular tail domains in red. (d) Averaged structure of cryo-tomograms of in vitro assembled lamin A as shown in (b) displays the striped pattern comprising the purported Ig-fold domains at distances of 20 nm, comparable to the spacing of the repeating pairs of globular domains in some structural classes from our in situ structural analysis (). The distance of the putative Ig-fold domains within the stripes of the paracrystals is 6 nm. Scale bar, 20 nm. Fig. 3a and b

    Article Snippet: For immunogold-labeling experiments, rabbit monoclonal antibody directed against the rod domain of mouse lamin A/C (peptide between aa 200–300) (EP4520, ab133256) was purchased from Abcam.

    Techniques: In Vitro, Transmission Electron Microscopy, Staining, In Situ

    Co-Immunogold-labeling of A- and B-type lamins (a) Nuclei on EM-grids were treated with α-lamin A/C and α-lamin B1 and labeled with 6 nm and 10 nm gold conjugates, respectively, prior to vitrification and cryo-ET analysis. Control samples (middle and right) were either treated with α-lamin A/C prior to treatment with 6 nm gold conjugate, post-fixation and subsequent treatment with 10 nm gold conjugate (middle panel), or only with 6 nm and 10 nm gold conjugate (right panel), omitting incubation with antibodies. Large projection views of parts of nuclei (contoured by the light-grey line) display the distribution of gold conjugates under the indicated conditions. Scale bar, 200 nm. (b) Box plots show the labeling density of 6 nm (lamin A/C) and 10 nm (lamin B1) gold particles per μm 2  (+/− SD) under the indicated conditions (white line represents the median and black dot the average amount of gold particles). The number of gold particles was extracted from 47 sub-volumes (left panel), 9 sub-volumes (middle panel) or 11 sub-volumes (right panel). (c) To validate the reliable assignment of small and large gold colloids, 6 nm and 10 nm gold conjugated protein A label were manually picked from samples containing either 6 nm or 10 nm colloids (individual gold colloids), a mixture of both (mixed gold colloids) or from the co-immunogold labeling experiment (lamin A/C and B1 co-labeling experiments) and averaged (n = 200 per condition). The two images at the bottom show a collage of nine individual 6 nm (left) and 10 nm (right) gold conjugated protein A label, respectively, which were picked from the co-immunogold labeling experiments. The line plot shows the normalized intensity profile of the average diameter of the gold conjugated protein A labels that were picked from the indicated samples. The solid lines show the intensity profile of the averaged gold label from the samples containing only one type of colloid. The dashed lines show the intensity profile of the averaged gold labels picked from a mixture. The lines with diamonds show the profile of the averaged gold labels from the co-immunogold labeling experiment. The line profiles of the 6 nm and 10 nm averaged gold conjugated protein A labels are shown in green and red, respectively. The line plot shows almost identical diameters of the 6 nm or 10 nm gold labels for each condition.

    Journal: Nature

    Article Title: The molecular architecture of lamins in somatic cells

    doi: 10.1038/nature21382

    Figure Lengend Snippet: Co-Immunogold-labeling of A- and B-type lamins (a) Nuclei on EM-grids were treated with α-lamin A/C and α-lamin B1 and labeled with 6 nm and 10 nm gold conjugates, respectively, prior to vitrification and cryo-ET analysis. Control samples (middle and right) were either treated with α-lamin A/C prior to treatment with 6 nm gold conjugate, post-fixation and subsequent treatment with 10 nm gold conjugate (middle panel), or only with 6 nm and 10 nm gold conjugate (right panel), omitting incubation with antibodies. Large projection views of parts of nuclei (contoured by the light-grey line) display the distribution of gold conjugates under the indicated conditions. Scale bar, 200 nm. (b) Box plots show the labeling density of 6 nm (lamin A/C) and 10 nm (lamin B1) gold particles per μm 2 (+/− SD) under the indicated conditions (white line represents the median and black dot the average amount of gold particles). The number of gold particles was extracted from 47 sub-volumes (left panel), 9 sub-volumes (middle panel) or 11 sub-volumes (right panel). (c) To validate the reliable assignment of small and large gold colloids, 6 nm and 10 nm gold conjugated protein A label were manually picked from samples containing either 6 nm or 10 nm colloids (individual gold colloids), a mixture of both (mixed gold colloids) or from the co-immunogold labeling experiment (lamin A/C and B1 co-labeling experiments) and averaged (n = 200 per condition). The two images at the bottom show a collage of nine individual 6 nm (left) and 10 nm (right) gold conjugated protein A label, respectively, which were picked from the co-immunogold labeling experiments. The line plot shows the normalized intensity profile of the average diameter of the gold conjugated protein A labels that were picked from the indicated samples. The solid lines show the intensity profile of the averaged gold label from the samples containing only one type of colloid. The dashed lines show the intensity profile of the averaged gold labels picked from a mixture. The lines with diamonds show the profile of the averaged gold labels from the co-immunogold labeling experiment. The line profiles of the 6 nm and 10 nm averaged gold conjugated protein A labels are shown in green and red, respectively. The line plot shows almost identical diameters of the 6 nm or 10 nm gold labels for each condition.

    Article Snippet: For immunogold-labeling experiments, rabbit monoclonal antibody directed against the rod domain of mouse lamin A/C (peptide between aa 200–300) (EP4520, ab133256) was purchased from Abcam.

    Techniques: Labeling, Incubation