Synthetic peptide within Human Histone H2B aa 1-100 (N terminal) (acetyl K5) conjugated to Keyhole Limpet Haemocyanin (KLH). The exact sequence is proprietary. Database link: Q16778
This antibody gave a positive signal in HeLa Histone Nuclear prep and HeLa Butyrate treated whole cell lysate as well as NIH3T3 whole cell lysate.
ICC/IF - HeLa cells (treated with Sodium butyrate and untreated).
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
Batches of this product that have a concentration < 1mg/ml may have BSA added as a stabilising agent. If you would like information about the formulation of a specific lot, please contact our scientific support team who will be happy to help.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml. Detects a band of approximately 17 kDa (predicted molecular weight: 14 kDa).
Use 2 µg for 25 µg of chromatin.
Use a concentration of 0.5 µg/ml.
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.
Belongs to the histone H2B family.
Monoubiquitination of Lys-121 by the RNF20/40 complex gives a specific tag for epigenetic transcriptional activation and is also prerequisite for histone H3 'Lys-4' and 'Lys-79' methylation. It also functions cooperatively with the FACT dimer to stimulate elongation by RNA polymerase II. Phosphorylated on Ser-15 by STK4/MST1 during apoptosis; which facilitates apoptotic chromatin condensation. Also phosphorylated on Ser-15 in response to DNA double strand breaks (DSBs), and in correlation with somatic hypermutation and immunoglobulin class-switch recombination.
ab175046 staining Histone H2B (acetyl K5) in HeLa cells. The cells were incubated with 10mM Sodium butyrate (ab120948) for 6 hours (Treated) or solvent-only for control purposes (Non-treated). Cells were fixed with 100% methanol (5min) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated with ab175046 at 0.5µg/ml and ab7291 at 1µg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with an AlexaFluor®488 Goat anti-Rabbit secondary (ab150077) at 2 μg/ml (shown in green) and AlexaFluor®594 Goat anti-Mouse secondary (ab150120) at 2 μg/ml (shown in pseudo colour red). Nuclear DNA was labelled in blue with DAPI.
Negative controls: 1– Rabbit primary and anti-mouse secondary antibody; 2 – Mouse primary antibody and anti-rabbit secondary antibody. Controls 1 and 2 indicate that there is no unspecific reaction between primary and secondary antibodies used.
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 ab175046 (blue), and 20µl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach for active and inactive loci, Sybr green approach for heterochromatic loci). Primers and probes are located in the first kb of the transcribed region.
Western blot - Anti-Histone H2B (acetyl K5) antibody - ChIP Grade (ab175046)
All lanes : Anti-Histone H2B (acetyl K5) antibody - ChIP Grade (ab175046) at 1 µg/ml
Lane 1 : ATCC HeLa Butyrate (0µM, 6hr) Whole Cell Lysate (ab189243) Lane 2 : ATCC HeLa Butyrate (10µM, 6hr) Whole Cell Lysate (ab189242) Lane 3 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate
Lysates/proteins at 2.5 µg per lane.
Secondary All lanes : Goat Anti-Rabbit IgG H&L (Alexa Fluor® 790) (ab175781) at 1/10000 dilution
This blot was produced using a 4-12% Bis-tris gel under the MES buffer system. The gel was run at 200V for 35 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using Licor blocking buffer before being incubated with ab175046 overnight at 4°C. Antibody binding was detected using ab175781 at a 1:10,000 dilution for 1hr at room temperature and then imaged using the Licor Odyssey CLx.