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Histone H4 (acetyl K16) Recombinant Rabbit Monoclonal Antibody [JB21-44] - ChIP Grade

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Specification

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Catalog# ET7107-89

Histone H4 (acetyl K16) Recombinant Rabbit Monoclonal Antibody [JB21-44] - ChIP Grade

Application

  • WB

  • IHC-P

  • FC

  • IF-Cell

  • IF-Tissue

  • ChIP

Reactivity

  • Human

  • Mouse

  • Rat

BSA and Azide free
Conjugation

  • unconjugated

This product has been cited in peer reviewed publications, see list HERE

Overview

Product Name

Histone H4 (acetyl K16) Recombinant Rabbit Monoclonal Antibody [JB21-44] - ChIP Grade

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Recombinant protein within Human Histone H4 (acetyl K16) aa 1-50 / 103.

Species Reactivity

Human, Mouse, Rat

Validated Applications

WB, IHC-P, FC, IF-Cell, IF-Tissue, ChIP

Target Molecular Weight

Predicted band size: 11 kDa

Positive Control

HeLa cell lysate, HeLa treated with 1μM TSA for 18 hours cell lysate, C6 cell lysate, C6 treated with 1μM TSA for 18 hours cell lysate, NIH/3T3 cell lysate, NIH/3T3 treated with 400nM TSA for 18 hours cell lysate, HeLa, human colon tissue, mouse colon tissue.

Conjugation

unconjugated

Clone Number

JB21-44

RRID

AB_3070794

Product Features

Form

Liquid

Concentration

1 mg/mL.(The concentration of this product may be batch-dependent)

Storage Instructions

Shipped at 4℃. Store at +4℃ short term (1-2 weeks). Store at -20℃ long term.

Storage Buffer

1*PBS (pH7.4), 0.1% BSA, 40% Glycerol, 0.2% Proclean 950.

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

  • WB

  • 1:1,000

  • IF-Cell

  • 1:50-1:200

  • IF-Tissue

  • 1:50-1:200

  • IHC-P

  • 1:200-1:1,000

  • FC

  • 1:1,000

  • ChIP

  • Use 0.5~2 μg for 25 μg of chromatin.

Target

Function

Eukaryotic histones are basic and water soluble nuclear proteins that form hetero-octameric nucleosome particles by wrapping 146 base pairs of DNA in a left-handed super-helical turn sequentially to form chromosomal fiber. Two molecules of each of the four core histones (H2A, H2B, H3, and H4) form the octamer; formed of two H2A-H2B dimers and two H3-H4 dimers, forming two nearly symmetrical halves by tertiary structure. Over 80% of nucleosomes contain the linker Histone H1, derived from an intronless gene, that interacts with linker DNA between nucleosomes and mediates compaction into higher order chromatin. Histones are subject to posttranslational modification by enzymes primarily on their N-terminal tails, but also in their globular domains. Such modifications include methylation, citrullination, acetylation, phosphorylation, sumoylation, ubiquitination and ADP-ribosylation.

Background References

1. Kurata M et al. Characterization of t(3;6)(q27;p21) breakpoints in B-cell non-Hodgkin\'s lymphoma and construction of the histone H4/BCL6 fusion gene, leading to altered expression of Bcl-6. Cancer Res 62:6224-6230 (2002).

2. Pesavento J J et al. Certain and progressive methylation of histone H4 at lysine 20 during the cell cycle. Mol Cell Biol 28:468-486 (2008).

Sequence Similarity

Belongs to the histone H4 family.

Post-translational Modification

Acetylation at Lys-6 (H4K5ac), Lys-9 (H4K8ac), Lys-13 (H4K12ac) and Lys-17 (H4K16ac) occurs in coding regions of the genome but not in heterochromatin.; Citrullination at Arg-4 (H4R3ci) by PADI4 impairs methylation.; Monomethylation and asymmetric dimethylation at Arg-4 (H4R3me1 and H4R3me2a, respectively) by PRMT1 favors acetylation at Lys-9 (H4K8ac) and Lys-13 (H4K12ac). Demethylation is performed by JMJD6. Symmetric dimethylation on Arg-4 (H4R3me2s) by the PRDM1/PRMT5 complex may play a crucial role in the germ-cell lineage.; Monomethylated, dimethylated or trimethylated at Lys-21 (H4K20me1, H4K20me2, H4K20me3). Monomethylation is performed by SET8. Dimethylation and trimethylation is performed by KMT5B and KMT5C and induces gene silencing (By similarity).; Phosphorylated by PAK2 at Ser-48 (H4S47ph). This phosphorylation increases the association of H3.3-H4 with the histone chaperone HIRA, thus promoting nucleosome assembly of H3.3-H4 and inhibiting nucleosome assembly of H3.1-H4.; Ubiquitinated by the CUL4-DDB-RBX1 complex in response to ultraviolet irradiation. This may weaken the interaction between histones and DNA and facilitate DNA accessibility to repair proteins. Monoubiquitinated at Lys-92 of histone H4 (H4K91ub1) in response to DNA damage. The exact role of H4K91ub1 in DNA damage response is still unclear but it may function as a licensing signal for additional histone H4 post-translational modifications such as H4 Lys-21 methylation (H4K20me).; Sumoylated, which is associated with transcriptional repression.; Crotonylation (Kcr) is specifically present in male germ cells and marks testis-specific genes in post-meiotic cells, including X-linked genes that escape sex chromosome inactivation in haploid cells. Crotonylation marks active promoters and enhancers and confers resistance to transcriptional repressors. It is also associated with post-meiotically activated genes on autosomes.; Butyrylation of histones marks active promoters and competes with histone acetylation.; Glutarylation at Lys-92 (H4K91glu) destabilizes nucleosomes by promoting dissociation of the H2A-H2B dimers from nucleosomes.

Subcellular Location

Chromosome, Nucleosome core, Nucleus.

UNIPROT

SwissProt: P62805 Human

SwissProt: P62806 Mouse

SwissProt: P62804 Rat

Synonyms

Histone gene cluster 1, H4A antibody

Histone gene cluster 2, H4 antibody

dJ160A22.1 antibody

dJ160A22.2 antibody

dJ221C16.1 antibody

dJ221C16.9 antibody

FO108 antibody

H4 antibody

H4 histone family, member A antibody

H4 histone family, member B antibody

Expand

Images

  • <span style="font-weight: bold;">☑ Cell treatment (CT)</span><br /><br />Western blot analysis of Histone H4 (acetyl K16) on different lysates with Rabbit anti-Histone H4 (acetyl K16) antibody (<a href="/products/ET7107-89" style="font-weight: bold;text-decoration: underline;">ET7107-89</a>) at 1/1,000 dilution.<br /><br />Lane 1: HeLa cell lysate<br />Lane 2: HeLa treated with 1μM TSA for 18 hours cell lysate<br />Lane 3: C6 cell lysate<br />Lane 4: C6 treated with 1μM TSA for 18 hours cell lysate<br />Lane 5: NIH/3T3 cell lysate<br />Lane 6: NIH/3T3 treated with 400nM TSA for 18 hours cell lysate<br /><br />Lysates/proteins at 10 µg/Lane.<br /><br />Predicted band size: 11 kDa<br />Observed band size: 11 kDa<br /><br />Exposure time: 3 minutes; ECL: K1801;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET7107-89" style="font-weight: bold;text-decoration: underline;">ET7107-89</a>) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    ☑ Cell treatment (CT)

    Western blot analysis of Histone H4 (acetyl K16) on different lysates with Rabbit anti-Histone H4 (acetyl K16) antibody (ET7107-89) at 1/1,000 dilution.

    Lane 1: HeLa cell lysate
    Lane 2: HeLa treated with 1μM TSA for 18 hours cell lysate
    Lane 3: C6 cell lysate
    Lane 4: C6 treated with 1μM TSA for 18 hours cell lysate
    Lane 5: NIH/3T3 cell lysate
    Lane 6: NIH/3T3 treated with 400nM TSA for 18 hours cell lysate

    Lysates/proteins at 10 µg/Lane.

    Predicted band size: 11 kDa
    Observed band size: 11 kDa

    Exposure time: 3 minutes; ECL: K1801;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET7107-89) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • Immunocytochemistry analysis of HeLa cells labeling Histone H4 (acetyl K16) with Rabbit anti-Histone H4 (acetyl K16) antibody (<a href="/products/ET7107-89" style="font-weight: bold;text-decoration: underline;">ET7107-89</a>) at 1/100 dilution.<br /><br />Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Histone H4 (acetyl K16) antibody (<a href="/products/ET7107-89" style="font-weight: bold;text-decoration: underline;">ET7107-89</a>) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.<br /><br />Beta tubulin (<a href="/products/HA601187" style="font-weight: bold;text-decoration: underline;">HA601187</a>, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor&trade; 594, <a href="/products/HA1126" style="font-weight: bold;text-decoration: underline;">HA1126</a>) was used as the secondary antibody at 1/1,000 dilution.

    Immunocytochemistry analysis of HeLa cells labeling Histone H4 (acetyl K16) with Rabbit anti-Histone H4 (acetyl K16) antibody (ET7107-89) at 1/100 dilution.

    Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Histone H4 (acetyl K16) antibody (ET7107-89) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

    Beta tubulin (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

  • Immunohistochemical analysis of paraffin-embedded human colon tissue with Rabbit anti-Histone H4 (acetyl K16) antibody (<a href="/products/ET7107-89" style="font-weight: bold;text-decoration: underline;">ET7107-89</a>) at 1/200 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET7107-89" style="font-weight: bold;text-decoration: underline;">ET7107-89</a>) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded human colon tissue with Rabbit anti-Histone H4 (acetyl K16) antibody (ET7107-89) at 1/200 dilution.

    The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-89) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunohistochemical analysis of paraffin-embedded mouse colon tissue with Rabbit anti-Histone H4 (acetyl K16) antibody (<a href="/products/ET7107-89" style="font-weight: bold;text-decoration: underline;">ET7107-89</a>) at 1/1,000 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET7107-89" style="font-weight: bold;text-decoration: underline;">ET7107-89</a>) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded mouse colon tissue with Rabbit anti-Histone H4 (acetyl K16) antibody (ET7107-89) at 1/1,000 dilution.

    The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-89) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Flow cytometric analysis of HeLa cells labeling Histone H4 (acetyl K16).<br /><br />Cells were fixed and permeabilized. Then stained with the primary antibody (<a href="/products/ET7107-89" style="font-weight: bold;text-decoration: underline;">ET7107-89</a>, 1ug/ml) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor&trade; 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (<a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

    Flow cytometric analysis of HeLa cells labeling Histone H4 (acetyl K16).

    Cells were fixed and permeabilized. Then stained with the primary antibody (ET7107-89, 1ug/ml) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

  • Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells with Histone H4 (acetyl K16) (<a href="/products/ET7107-89" style="font-weight: bold;text-decoration: underline;">ET7107-89</a>) or Normal Rabbit IgG according to the ChIP protocol. The enriched DNA was quantified by real-time PCR using indicated primers. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.

    Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells with Histone H4 (acetyl K16) (ET7107-89) or Normal Rabbit IgG according to the ChIP protocol. The enriched DNA was quantified by real-time PCR using indicated primers. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.

Citation

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