Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade

12 product images

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  ChIP - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Chromatin was prepared from HeLa cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25μg of chromatin, 2μg of ab45166 (red), and 20μl of Protein A/G sepharose beads. No antibody was added to the beads control (grey). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach). Primers and probes are located in the first kb of the transcribed region.

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  Immunoprecipitation - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  ab45166 (purified) at 1/20 immunoprecipitating Histone H4 (acetyl K8) in HeLa treated with Trichostatin A whole cell lysate.
  

  Lane 1 (input): HeLa treated with Trichostatin A whole cell lysate (10μg)

  Lane 2 (+): ab45166 + HeLa treated with Trichostatin A whole cell lysate.

  Lane 3 (-): Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab45166 in HeLa treated with Trichostatin A whole cell lysate.

  For western blotting, VeriBlot for IP Detection Reagent (HRP) ab131366 VeriBlot for IP Detection Reagent (HRP) was used for detection (1/10000).

  Blocking buffer and concentration: 5% NFDM/TBST.

  Diluting buffer and concentration: 5% NFDM /TBST.

  All lanes: Immunoprecipitation - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166) at 1/20 dilution

  Lane 1: HeLa (human cervix adenocarcinoma) treated with Trichostatin A whole cell lysate at 10 µg

  Lanes 2 - 3: HeLa (human cervix adenocarcinoma) treated with Trichostatin A whole cell lysate

  Secondary

  All lanes: Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) at 1/10000 dilution

  Predicted band size: 11 kDa

  Observed band size: 11 kDa

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  ChIP-sequencing - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Chromatin was prepared from HeLa cells. Cells were fixed with 1% formaldehyde for 10 minutes. ChIP was performed with 10^7 HeLa cells and 4 μg of ab45166 [EP1002Y]. ChIP DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 30 million reads.
  
  Additional screenshots of mapped reads can be downloaded here.

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  Flow Cytometry (Intracellular) - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) treated (Red)/untreated (Green) with 500ng/ml Trichostatin A for 4 hours with purified ab45166 at 1/20 dilution. The secondary antibody was Goat anti rabbit IgG (Alexa Fluorr^® 488) at 1/2000 dilution. A Rabbit monoclonal IgG (Black) was used as the isotype control and cells without incubation with primary antibody and secondary antibody (Blue) were used as unlabeled control.
  

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  Western blot - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Blocking and diluting buffer 5% NFDM/TBST

  All lanes: Western blot - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166) at 1/5000 dilution

  Lane 1: Untreated HeLa (human cervix adenocarcinoma) whole cell lysate at 10 µg

  Lane 2: HeLa (human cervix adenocarcinoma) treated with Trichostatin A whole cell lysate at 10 µg

  Secondary

  All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

  Predicted band size: 11 kDa

  Observed band size: 11 kDa

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  Immunocytochemistry/ Immunofluorescence - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Immunocytochemistry/immunofluorescence staining of 4% paraformaldehyde fixed; 0.1% triton X 100 permeabilized C6 (rat glioma) cells (non-treated-top panels) and (C6 + TSA(500ng/ml, 4hr)-middle panels) with purified ab45166 at dilution of 1/150. The secondary antibody used was Alexa Fluor® 488; goat anti-rabbit IgG (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) at a dilution of 1/1000. Nucleus was counter-stained with DAPI (blue). Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291, a mouse anti-tubulin antibody (1/1000) was used to stain tubulin along with Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120 (AlexaFluor®594 goat anti-mouse secondary, 1/1000) shown in the top right and middle right hand panels. The negative controls are shown in the bottom two panels- for negative control 1 rabbit primary antibody and anti-mouse secondary antibody (Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120) was used. For negative control 2 mouse primary antibody (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) and anti-rabbit secondary antibody (Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077) was used.

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Immunohistochemical staining of paraffin-embedded rat kidney sections labelling Histone H4 (acetyl K8) with purified ab45166 at dilution of 1:2500. The secondary antibody used was Goat Anti-Rabbit IgG H&L (HRP) ab97051; a goat anti-rabbit IgG H&L (HRP) at dilution of 1/500. The sample was counter-stained with hematoxylin. Antigen retrieval was performed using EDTA Buffer; pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.

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  Western blot - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Blocking and diluting buffer 5% NFDM/TBST

  All lanes: Western blot - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166) at 1/5000 dilution

  Lane 1: Untreated C6 (rat glioma) whole cell lysate at 10 µg

  Lane 2: C6 (rat glioma) treated with Trichostatin A whole cell lysate at 10 µg

  Secondary

  All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

  Predicted band size: 11 kDa

  Observed band size: 11 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Immunohistochemical staining of paraffin-embedded mouse kidney sections labelling Histone H4 (acetyl K8) with purified ab45166 at dilution of 1:2500. The secondary antibody used was Goat Anti-Rabbit IgG H&L (HRP) ab97051; a goat anti-rabbit IgG H&L (HRP) at dilution of 1/500. The sample was counter-stained with hematoxylin. Antigen retrieval was performed using EDTA Buffer; pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.

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  Western blot - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Blocking and diluting buffer 5% NFDM/TBST

  All lanes: Western blot - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166) at 1/5000 dilution

  Lane 1: Untreated NIH/3T3 (mouse embryo) whole cell lysate at 10 µg

  Lane 2: NIH/3T3 (mouse embryo) treated with Trichostatin A whole cell lysate at 10 µg

  Secondary

  All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution

  Predicted band size: 11 kDa

  Observed band size: 11 kDa

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Immunohistochemical staining of paraffin-embedded human colon sections labelling Histone H4 (acetyl K8) with purified ab45166 at dilution of 1:2500. The secondary antibody used was Goat Anti-Rabbit IgG H&L (HRP) ab97051; a goat anti-rabbit IgG H&L (HRP) at dilution of 1/500. The sample was counter-stained with hematoxylin. Antigen retrieval was performed using EDTA Buffer; pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.

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  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H4 (acetyl K8) antibody [EP1002Y] - ChIP Grade (ab45166)

  Immunohistochemical analysis of formalin fixed paraffin embedded human colon tissue sections labelling Histone H4 (acetyl K8) with unpurified ab45166 at dilution of 1/200.
  

  For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.