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H3k9ac

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H3K9ac is a histone modification antibody that specifically recognizes acetylation at lysine 9 of histone H3. This antibody can be used in various applications such as chromatin immunoprecipitation (ChIP), Western blotting, and immunofluorescence to detect and study the presence and distribution of this epigenetic mark.

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Lab products found in correlation

H3k27ac, by Abcam (4 mentions) H3k27me3, by Cell Signaling Technology (3 mentions) H3k27me3, by Merck Group (3 mentions) H3k9me3, by Abcam (3 mentions) H3k4me1, by Abcam (3 mentions) H3k14ac, by Active Motif (3 mentions) H3k36me3, by Abcam (3 mentions) H3k4me3, by Abcam (3 mentions) H3k9me3, by Active Motif (2 mentions) H3k27me3, by Active Motif (2 mentions) Suv39h1, by Merck Group (2 mentions) H3k4me3, by Merck Group (2 mentions) Chip isolation kit, by Merck Group (2 mentions) H3k4me, by Merck Group (2 mentions) Eset setdb1, by Merck Group (2 mentions) G9a ehmt2 c6h3, by Abcam (2 mentions) H3k27me2, by Abcam (2 mentions) H3k4me2, by Abcam (2 mentions) H3k9me2, by Merck Group (2 mentions) Total h3, by Cell Signaling Technology (2 mentions)

Spelling variants of this product

Anti-H3K9ac (12 citations) Histone H3K9ac (4 citations) H3K9ac antibody (4 citations) Acetylated histone H3K9 (H3K9Ac) antibody (2 citations) Anti-H3K9ac (61251) (2 citations) Anti-H3K9ac antibody (2 citations)

17 protocols using h3k9ac

1

Immunoblotting for Nuclear Protein Markers

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Reduced samples were loaded onto NuPAGE Bis-Tris (Thermo Fisher Scientific) gels in MOPS buffer and separated proteins were transferred onto a PVDF membrane for immunodetection with anti-PPARγ (C26H12, 1:2000: Cell Signaling Technologies, Danvers, MA, USA), HSP90α (GTX109753: Gene Tex, Irvine, CA, USA, 1:2000), HDAC1 (GTX100513: Gene Tex, 1:2000), GAPDH (GTX100118: Gene Tex, 1:1000), H3K27ac (39134: Active Motif, Carlsbad, CA, USA, 1:2000), H3K9ac (39918: Active Motif, 1:2000), H3k27me3 (C36B11: Cell Signaling Technologies, 1:2000), and Histone H3 (D2B12: Cell Signaling Technologies, 1:2000) antibodies as primary antibodies, and the HRP-conjugated goat anti-rabbit antibody (#7074: Cell Signaling Technologies, 1:2000) as the secondary antibody.
Yuan H., Suzuki S., Hirata-Tsuchiya S., Sato A., Nemoto E., Saito M., Shiba H, & Yamada S. (2021). PPARγ-Induced Global H3K27 Acetylation Maintains Osteo/Cementogenic Abilities of Periodontal Ligament Fibroblasts. International Journal of Molecular Sciences, 22(16), 8646.
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2

Chromatin Immunoprecipitation and Analysis

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Chromatin isolation and chromatin immunoprecipitation (ChIP) analyses were carried out as described [42 (link)]. Antibodies used for ChIP were directed against H2A.Zac (Diagenode, Liège, Belgium), H3K4me1 (Diagenode, Liège, Belgium), H3K4me3 (Diagenode, Liège, Belgium), H3K9ac (Active Motif, Carlsbad, CA, USA), H3K9me3 (Active Motif, Carlsbad, CA, USA), H3K9me3S10ph (Diagenode, Liège, Belgium) or H3K27me3 (Active Motif, Carlsbad, CA, USA). Relative enrichment of precipitated sequences in ChIP was assessed using qPCR.
Jenke A.C., Hensel K.O., Klein A., Willuhn L., Prax S., Weil P.P., Winkler T., Deba T., Orth V., Baiker A., Wirth S, & Postberg J. (2014). Restitution of gene expression and histone acetylation signatures altered by hepatitis B virus through antiviral microRNA-like molecules in nontransformed murine hepatocytes. Clinical Epigenetics, 6(1), 26.
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3

Chromatin Immunoprecipitation Assay Protocol

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The ChIP assays were performed using a ChIP isolation kit (Millipore, Billerica, MA). We treated 1 × 105 to 5 × 105 murine naïve T cells with 1% formaldehyde to cross-link histones to DNA. The fixed cells were sonicated to yield chromatin fragments of 200–500 base pairs (bp). The antibodies used in the ChIP assays included: H3Ac (Millipore), H4Ac (Upstate Biotechnology, Lake Placid, NY), H3K9Ac (Active Motif, Carlsbad, CA), H3K4Me3 (Millipore), H3K4Me2 (Abcam), H3K4Me (Millipore), H3K9Me2 (Millipore), H3K9Me3 (Abcam) H3K27me2 (Abcam) H3K27me3 (Millipore), p300 (Abcam), PCAF (Abcam), SUV39H1 (Millipore), CBP (Abcam), ESET/SetDB1(Millipore), Ezh2 (Cell Signaling Technology, Beverly, MA), G9a/EHMT2 (C6H3) (Abcam), HP1α (Millipore), HP1β (Active Motif), and HP1γ (Millipore). DNA was recovered by a Chelex 10% slurry and sample boiling method,10 (link) or using the IP-STAR (Diagenode, Denville, NJ) direct chip protocol followed by the IPPURE DNA purification. 1% Pre-enriched chromatin (input) served as the percentage input for sample quantification, confirmed by fold over IgG changes. The UV-exposed 4% agarose gels were imaged, and the digitized images were analyzed using the software VisionWorks (UVP). Protein signals were indexed by measuring their relative mean grey value per area.
Sarmento O.F., Svingen P.A., Xiong Y., Xavier R.J., McGovern D., Smyrk T.C., Papadakis K.A., Urrutia R.A, & Faubion W.A. (2014). A Novel Role for Kruppel-like Factor 14 (KLF14) in T-Regulatory Cell Differentiation. Cellular and Molecular Gastroenterology and Hepatology, 1(2), 188-202.e4.
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4

Chromatin Immunoprecipitation from Rat DRG Tissue

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We performed chromatin immunoprecipitation (ChIP) using the Magna ChIP G tissue kit (Millipore; catalog #17-20000), according to the manufacturer’s instructions. Briefly, fresh DRG tissues were rapidly removed from anesthetized rats and were stabilized for 3 min using stabilization buffer. Then, the DRGs were incubated in 2% formaldehyde for 20 min at ~26 °C. After being washed three times with PBS, the DRGs were incubated in lysis buffer for 15 min on ice. Finally, the DRG tissues were sonicated (30 s on and 30 s off, repeated 200 times) in ChIP dilution buffer using a water bath sonicator (Qsonica, Newtown, CT) at 4 °C. The sonicated DRG samples were centrifuged at 12,000 rpm for 10 min at 4 °C. Chromatin was pulled down using the following antibodies: IgG (as a negative control; catalog #ab124055, Abcam), total H3 (catalog #2650s, Cell Signaling Technology), H3K9me2 (catalog #ab1220, Abcam), H3K9ac (catalog #39917, Active Motif), H3K27me3 (catalog #9773s, Cell Signaling Technology), and H3K4me3 (catalog #9751s, Cell Signaling Technology). After chromatin precipitation, we performed quantitative PCR using the primers described in Supplementary Table 4. Data were analyzed and corrected by input and total H3 conditions 60 (link).
Laumet G., Garriga J., Chen S.R., Zhang Y., Li D.P., Smith T.M., Dong Y., Jelinek J., Cesaroni M., Issa J.P, & Pan H.L. (2015). G9a Is Essential for Epigenetic Silencing of K+ Channel Genes in Acute-to-Chronic Pain Transition. Nature neuroscience, 18(12), 1746-1755.
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5

Western Blot Analysis of Protein Expression

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Cells were lysed in modified RIPA buffer containing 150 mM NaCl, 1% NP-40, 50 mM Tris-Cl, pH 8.0, and 1% SDS, supplemented with protease inhibitors (Life Technologies, #78446) before use. Protein concentration was determined by BCA protein assay (Life Technologies, #23227), following which equal amount of proteins were loaded and separated in polyacrylamide gels. Proteins were then transferred to nitrocellulose membrane. Antibodies used in this study were as follows: p53 monoclonal antibody DO-1 (Calbiochem EMD); p53 polyclonal antibody FL393 (Santa Cruz Biotechnology Inc., sc-6243). Flag (Sigma, M2, F1804), HA (Rockland, 600-401-384), histone H3 (abcam, ab1791), H3K4me1 (abcam, ab8895), H3K4me2 (Active Motif, 39142), H3K4me3 (abcam, ab8580), H3K9ac (Active Motif, 39137), H3K14ac (Active Motif, 39616), H3K27ac (abcam, ab4729), H3K36me3 (abcam, ab9050), ETS2 (Santa Cruz Biotechnology Inc., sc-351), MLL1 (Bethyl Laboratories Inc., A300-086A), MOZ (Novus Biologicals, 21620002), mouse p53 antibody for ChIP experiments (Santa Cruz Biotechnology Inc., sc-1312 (M-19)), mouse p53 antibody for western blot analysis (Cell Signaling Technology, #2524), RNA polymerase II (abcam, ab817).
Zhu J., Sammons M.A., Donahue G., Dou Z., Vedadi M., Getlik M., Barsyte-Lovejoy D., Al-Awar R., Katona B.W., Shilatifard A., Huang J., Hua X., Arrowsmith C.H, & Berger S.L. (2015). Prevalent p53 mutants co-opt chromatin pathways to drive cancer growth. Nature, 525(7568), 206-211.
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6

Mitochondrial and Cell Cycle Markers

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The following antibodies were used: NDUFB8 (Abcam ab110242), SDHA (Cell Signaling Technology 11998), UQCR2 (Abcam ab14745), Tom20 (Santa Cruz Biotechnology sc-11415), p-Rb ser807/811 (Cell Signaling Technology 9308), Cyclin A (Santa Cruz Biotechnology sc-751), p21 (Santa Cruz Biotechnology sc-471 and HUGO 291 Abcam ab107099), IL8 (Abcam ab18672), p-JNK Thr183/Tyr188 (Cell Signaling Technology 9251), JNK (Cell Signaling Technology 9252 and Santa Cruz Biotechnology sc-7345), γH2AX ser139 (Millipore-Merck 05-636 and Cell Signaling Technology 9718), 53BP1 (Abcam 21083 and Cell Signaling Technology 4937), H4K16ac (Millipore-Merck 07-329), H4 (Active Motif 39163), H3K9ac (Active Motif 61252), H3 (Active Motif 39763), cleaved caspase 3 (Cell Signaling Technology 9661), p-38 MAPK Thr180/Tyr182 (Cell Signaling Technology 9211), p38 MAPK (Cell Signaling Technology 9212), p-S6 ser235/236 (Cell Signaling Technology 2211), pS6 (Cell Signaling Technology 2317), IRAK1 (Santa Cruz Biotechnology 5288), Actin (Sigma-Aldrich A1978), GAPDH (Cell Signaling Technology 5174), IL1α (R&D Systems AF-400), and 4-HNE (JaICA).
Vizioli M.G., Liu T., Miller K.N., Robertson N.A., Gilroy K., Lagnado A.B., Perez-Garcia A., Kiourtis C., Dasgupta N., Lei X., Kruger P.J., Nixon C., Clark W., Jurk D., Bird T.G., Passos J.F., Berger S.L., Dou Z, & Adams P.D. (2020). Mitochondria-to-nucleus retrograde signaling drives formation of cytoplasmic chromatin and inflammation in senescence. Genes & Development, 34(5-6), 428-445.
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7

Histone Acetylation and Methylation Analysis

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PU139 was dissolved in a solution of Tween-80/0.9% NaCl (1:10). Mice were injected i.p. with 100 μl PU139 (10 mg/ml, 25 μg/g body weight). Bone marrow cells were flushed in IMDM medium using a 26-G syringe on day 1 post PU139 injections. Red blood cell lysis was carried out with erythrocyte lysis buffer (Sigma) followed by centrifugation and resuspension of the pellet in IMDM. Cell counting was done using a Neubauer counting chamber. For western analysis, cells were resuspended in IMDM containing SDS loading dye and 10 mM sodium butyrate. Typically, 105 cells were lysed in 100 μl IMDM. Cells were sheared by passing the cells five or six times through a 26-G 1-ml syringe, immediately followed by boiling at 95 oC for 5 min. Lysates were stored at −20 oC until use. To check acetylation levels, lysates were probed with antibodies against H3K14-Ac (Active Motif), H4K16-Ac (Active Motif), H3K9-Ac (Active Motif) and H4K8-Ac (Cell Signaling, Danvers, MA, USA). In addition, western blot analysis was done for H3K27me3 (Millipore, Billerica, MA, USA) and H3K9me3 (Millipore). Blots were stripped and reprobed with antibodies against total H3 (Abcam, Cambridge, UK) or total H4 (Abcam) to ensure equal loading.
Gajer J.M., Furdas S.D., Gründer A., Gothwal M., Heinicke U., Keller K., Colland F., Fulda S., Pahl H.L., Fichtner I., Sippl W, & Jung M. (2015). Histone acetyltransferase inhibitors block neuroblastoma cell growth in vivo. Oncogenesis, 4(2), e137-.
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8

Chromatin Immunoprecipitation (ChIP) Assay

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ChIP assays were performed using a ChIP isolation kit (Millipore). 1 × 105 to 5 × 105 murine naïve T cells were treated with 1% formaldehyde to cross-link histones to DNA. Fixed cells were sonicated to yield chromatin fragments of 200–500 bp. Antibodies used in the ChIP assays included: H3Ac (Millipore), H4Ac (upstate), H3K9Ac (Active Motif), H3K4Me3 (Millipore), H3K4Me2 (Abcam), H3K4Me (Millipore), H3K9Me2 (Millipore), H3K9Me3 (Abcam) H3K27me2 (Abcam) H3K27me3 (Millipore), p300 (Abcam), PCAF (Abcam), SUV39H1(Millipore) , CBP (Abcam), ESET/SetDB1(Millipore), Ezh2 (Cell Signalling), G9a/EHMT2 (C6H3) (Abcam), HP1α (millipore), HP1β (Active Motif), and HP1γ (Millipore ). DNA was recovered by Chelex 10% slurry and sample boiling method (Nature protocols), or utilizing the IP-STAR (diagenode) direct chip protocol followed by the IPPURE DNA purification. 1% Pre-enriched chromatin (Input) served as percent Input for sample quantification, confirmed by fold over IgG changes. UV exposed 4% agarose gels were imagined and digitized images were analyzed using the software visionworks (UVP). Protein signals were indexed by measuring their relative mean grey value per area.
Sarmento O.F., Svingen P.A., Xiong Y., Xavier R.J., McGovern D., Smyrk T.C., Papadakis K.A., Urrutia R.A, & Faubion W.A. (2014). A Novel Role for Kruppel-like Factor 14 (KLF14) in T-Regulatory Cell Differentiation. Cellular and Molecular Gastroenterology and Hepatology, 1(2), 188-202.e4.
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9

Western Blot Analysis of Apoptosis and Histone Modification

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Tissue or cell lysates were prepared in a RIPA buffer supplemented with protease inhibitor cocktail and phosphatase inhibitor cocktail. Electrophoresis was performed by using BOLT 4%–12% gradient gel (Thermo Fisher Scientific). Western blots were developed and imaged using a ChemiDoc Touch Imaging System (Bio-Rad, Hercules, CA), band density was quantified by using Image Lab 5.2.1 software. The following antibodies were used: cleaved caspase 3 (9661; Cell Signaling), cleaved caspase 8 (8592; Cell Signaling, Danvers, MA), H3K9ac (39137; Active Motif, Carlsbad, CA), H3K27ac (ab4729; Abcam, Cambridge, MA), and anti–α-tubulin (T9026; Sigma-Aldrich, St. Louis, MO).
Mennillo E., Yang X., Paszek M., Auwerx J., Benner C, & Chen S. (2020). NCoR1 Protects Mice From Dextran Sodium Sulfate–Induced Colitis by Guarding Colonic Crypt Cells From Luminal Insult. Cellular and Molecular Gastroenterology and Hepatology, 10(1), 133-147.
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10

Chromatin Immunoprecipitation from Rat DRG Tissue

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We performed chromatin immunoprecipitation (ChIP) using the Magna ChIP G tissue kit (Millipore; catalog #17-20000), according to the manufacturer’s instructions. Briefly, fresh DRG tissues were rapidly removed from anesthetized rats and were stabilized for 3 min using stabilization buffer. Then, the DRGs were incubated in 2% formaldehyde for 20 min at ~26 °C. After being washed three times with PBS, the DRGs were incubated in lysis buffer for 15 min on ice. Finally, the DRG tissues were sonicated (30 s on and 30 s off, repeated 200 times) in ChIP dilution buffer using a water bath sonicator (Qsonica, Newtown, CT) at 4 °C. The sonicated DRG samples were centrifuged at 12,000 rpm for 10 min at 4 °C. Chromatin was pulled down using the following antibodies: IgG (as a negative control; catalog #ab124055, Abcam), total H3 (catalog #2650s, Cell Signaling Technology), H3K9me2 (catalog #ab1220, Abcam), H3K9ac (catalog #39917, Active Motif), H3K27me3 (catalog #9773s, Cell Signaling Technology), and H3K4me3 (catalog #9751s, Cell Signaling Technology). After chromatin precipitation, we performed quantitative PCR using the primers described in Supplementary Table 4. Data were analyzed and corrected by input and total H3 conditions 60 (link).
Laumet G., Garriga J., Chen S.R., Zhang Y., Li D.P., Smith T.M., Dong Y., Jelinek J., Cesaroni M., Issa J.P, & Pan H.L. (2015). G9a Is Essential for Epigenetic Silencing of K+ Channel Genes in Acute-to-Chronic Pain Transition. Nature neuroscience, 18(12), 1746-1755.
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