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Key Specifications Table
| Species Reactivity | Key Applications |
|---|---|
| B, H, M, Yeast (S. cerevisiae) | WB, ChIP |
| Description | |
|---|---|
| Catalogue Number | 17-10241 |
| Trade Name |
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| Description | ChIPAb+™ Acetyl-Histone H3 (Lys9/18) - ChIP Validated Antibody and Primer Set |
| Overview | All ChIPAb+ antibodies are individually validated for chromatin precipitation, every lot, every time. Each ChIPAb+ antibody set includes control primers (tested every lot by qPCR) to biologically validate your IP results in a locus-specific context. The qPCR protocol and primer sequences are provided, allowing researchers to validate ChIP protocols when using our antibody in their chromatin context. Each set also includes a negative control antibody to ensure specificity of the ChIP reaction. The ChIPAb+ Acetyl-Histone H3 (Lys9/18) set includes the Acetyl-Histone H3 (Lys9/18) antibody, a Normal rabbit serum, and control primers which amplify a 166 bp region of human GAPDH promoter. The Acetyl-Histone H3 (Lys9/18) and negative controls are supplied in a scalable "per ChIP" reaction size and can be used to functionally validate the precipitation of Acetyl-Histone H3 (Lys9/18)-associated chromatin. |
| Alternate Names |
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| Background Information | Histone H3 is one of the 5 main histone proteins involved in the structure of chromatin in eukaryotic cells. Featuring a main globular domain and a long N terminal tail H3 is involved with the structure of the nucleosomes of the beads on a string structure. Acetylation of histone H3 occurs at several different lysine positions in the histone tail and is performed by a family of enzymes known as Histone Acetyl Transferases (HATs). |
| Product Information | |
|---|---|
| Format | Serum |
| Control |
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| Presentation | Anti-Acetyl-Histone H3 (Lys9/18) (rabbit polyclonal), . One vial containing 25 µL of rabbit antiserum containing 0.05% sodium azide before the addition of glycerol to 30%. Store at -20°C. Normal Rabbit Serum, . One vial containing 25 μL of antiserum containing 0.05% sodium azide. Store at -20°C. Control Primers, human GAPDH promoter, . One vial containing 75 μL of 5 μM of each primer specific for human GAPDH. Store at -20°C. FOR: TAC TAG CGG TTT TAC GGG CG REV: TCG AAC AGG AGG AGC AGA GAG CGA |
| Quality Level | MQ100 |
| Applications | |
|---|---|
| Application | This ChIPAb+ Acetyl-Histone H3 (Lys9/18) -ChIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers. |
| Key Applications |
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| Application Notes | Chromatin Immunoprecipitation: Representative lot data. Sonicated chromatin prepared from HeLa cells (1 X 10E6 cell equivalents per IP) were subjected to chromatin immunoprecipitation using 1 µL of either Normal Rabbit Serum, or 1 µL of Anti-Acetyl-Histone H3 (Lys9/18), and the Magna ChIP™ A Kit (Cat. # 17-610). Successful immunoprecipitation of acetyl-Histone H3 (Lys9/18) associated DNA fragments was verified by qPCR using Control Primers specific for the human GAPDH promoter region as a positive locus and human β-globin primers as a negative locus. (Figure 2). Data is presented as percent input of each IP sample relative to input chromatin for each amplicon and ChIP sample as indicated. Please refer to the EZ-Magna ChIP™ A (Cat. # 17-408) or EZ-ChIP™ (Cat. # 17-371) protocol for experimental details Western Blot Analysis: Representative lot data. Acid extracts from untreated (lane 1), sodium butyrate treated (lane 2, Catalog # 17-305) HeLa cells and recombinant Histone H3, (Lane 3, Catalog # 14-411) were resolved by electrophoresis, transferred to nitrocellulose and probed with Anti-Acetyl-Histone H3 (Lys9/18) (1:5,000 dilution). Proteins were visualized using a goat anti-rabbit secondary antibody conjugated to HRP and a chemiluminescence detection system. Arrow indicates Acetyl-histone H3 (Lys9/18) (~17kDa). (Figure 3). Beadlyte® Histone-Peptide Specificity Assay: Representative lot data. 1:1,000-1:40,000 dilutions of anti-acetyl-Histone H3 (Lys9/18) were incubated with a cocktail of microspheres conjugated to histone H3 peptides with the following modifications: 1. acetyl-lysine 4 2. acetyl-lysine 14 3. acetyl-lysine 9 4. acetyl-lysine 18 5. No modifications, containing aa 1-20 Unbound antibody was removed by filtration. The peptide-antibody complexes were incubated with a biotin-conjugated anti-mouse secondary antibody followed by incubation with a phycoerythrin-streptavidin conjugate. Fluorescence was read on a Luminex® 100™ instrument. Median fluorescence intensity (MFI) is plotted. (Figure 4). |
| Biological Information | |
|---|---|
| Immunogen | KLH-conjugated synthetic peptide (..ARAcKSTGGKAPRAcKQL..) in which AcK corresponds to acetyl-lysine at residue 9 and 18 of human Histone H3. |
| Epitope | Acetylated Lys9/18 |
| Host | Rabbit |
| Specificity | Recognizes Histone H3 acetylated on lysines 9 and 18. |
| Species Reactivity |
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| Species Reactivity Note | Broad species cross-reactivity expected |
| Antibody Type | Polyclonal Antibody |
| Entrez Gene Number |
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| Entrez Gene Summary | Histones are basic nuclear proteins that are responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. Two molecules of each of the four core histones (H2A, H2B, H3, and H4) form an octamer, around which approximately 146 bp of DNA is wrapped in repeating units, called nucleosomes. The linker histone, H1, interacts with linker DNA between nucleosomes and functions in the compaction of chromatin into higher order structures. This gene contains introns and its mRNA is polyadenylated, unlike most histone genes. The protein encoded is a replication-independent member of the histone H3 family. |
| Gene Symbol |
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| Modifications |
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| Purification Method | Unpurified |
| UniProt Number |
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| UniProt Summary | FUNCTION: SwissProt: Q16695 # Core component of nucleosome. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling. SIZE: 136 amino acids; 15508 Da SUBUNIT: The nucleosome is a histone octamer containing two molecules each of H2A, H2B, H3 and H4 assembled in one H3-H4 heterotetramer and two H2A-H2B heterodimers. The octamer wraps approximately 147 bp of DNA. SUBCELLULAR LOCATION: Nucleus. PTM: Acetylation is generally linked to gene activation. Acetylation on Lys-10 impairs methylation at Arg-9. Acetylation on Lys-19 and Lys-24 favors methylation at Arg-18 (By similarity). & Citrullination at Arg-9 and/or Arg-18 by PADI4 impairs methylation and represses transcription (By similarity). & Asymmetric dimethylation at Arg-18 by CARM1 is linked to gene activation. Symmetric dimethylation at Arg-9 by PRMT5 is linked to gene repression (By similarity). & Methylation at Lys-5, Lys-37 and Lys-80 are linked to gene activation. Methylation at Lys-5 facilitates subsequent acetylation of H3 and H4. Methylation at Lys-80 is associated with DNA double-strand break (DSB) responses and is a specific target for TP53BP1. Methylation at Lys-10 and Lys-28 are linked to gene repression. Methylation at Lys-10 is a specific target for HP1 proteins (CBX1, CBX3 and CBX5) and prevents subsequent phosphorylation at Ser-11 and acetylation of H3 and H4. Methylation at Lys-5 and Lys-80 require preliminary monoubiquitination of H2B at Lys-120. Methylation at Lys-10 and Lys-28 are enriched in inactive X chromosome chromatin (By similarity). & Phosphorylated at Thr-4 by GSG2/haspin during prophase and dephosphorylated during anaphase. At centromeres, specifically phosphorylated at Thr-12 from prophase to early anaphase. Phosphorylated at Ser-11 during the whole mitosis. Phosphorylation at Ser-11, which is linked to gene activation, prevents methylation at Lys-10 but facilitates acetylation of H3 and H4. Phosphorylated at Ser-29 by MLTK isoform 1, RPS6KA5 or AURKB during mitosis or upon ultraviolet B irradiation (By similarity). & Phosphorylation at Ser-11 is crucial for chromosome condensation and cell-cycle progression during mitosis and meiosis. In addition phosphorylation at Ser-11 is important during interphase because it enables the transcription of genes following external stimulation, like stress or growth factors. Phosphorylation at Ser-11 is also an essential regulatory mechanism for neoplastic cell transformation. Phosphorylation at Ser-11 by AURKB/Aurora-B mediates the dissociation of HP1 proteins (CBX1, CBX3 and CBX5) from heterochromatin. & Ubiquitinated (By similarity). SIMILARITY: SwissProt: Q16695 ## Belongs to the histone H3 family. |
| Molecular Weight | 17 kDa |
| Product Usage Statements | |
|---|---|
| Quality Assurance | Chromatin Immunoprecipitation: Representative lot data. Sonicated chromatin prepared from HeLa cells (1 X 10E6 cell equivalents per IP) were subjected to chromatin immunoprecipitation using 1 µL of either Normal Rabbit Serum,or 1 µL of Anti-Acetyl-Histone H3 (Lys9/18) and the Magna ChIP™ A Kit (Cat. # 17-610). Successful immunoprecipitation of Acetyl-Histone H3 (Lys9/18) associated DNA fragments was verified by qPCR using Control Primers specific for the human GAPDH promoter region as a positive locus (Figure 1). Please refer to the EZ-Magna ChIP™ A (Cat. # 17-408) or EZ-ChIP™ (Cat. # 17-371) protocol for experimental details. |
| Usage Statement |
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| Storage and Shipping Information | |
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| Storage Conditions | Stable for 1 year at -20°C from date of receipt. Handling Recommendations: Upon first thaw, and prior to removing the cap, centrifuge the vial and gently mix the solution. Aliquot into microcentrifuge tubes and store at -20°C. Avoid repeated freeze/thaw cycles, which may damage IgG and affect product performance. Note: Variability in freezer temperatures below -20°C may cause glycerol containing solutions to become frozen during storage. |
| Packaging Information | |
|---|---|
| Material Size | 25 assays |
| Material Package | 25 assays per set. Recommended use: ~1 μL of antibody per chromatin immunoprecipitation (dependent upon biological context). |