Anti-Histone H2B (glcnac S112) antibody (ab130951)
Key features and details
- Rabbit polyclonal to Histone H2B (glcnac S112)
- Suitable for: IHC-P, WB, ICC/IF, PepArr
- Reacts with: Cow, Human
- Isotype: IgG
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.40
Preservative: 0.02% Sodium azide
Constituent: PBS
Batches of this product that have a concentration
Concentration information loading...Purity
Immunogen affinity purifiedClonality
PolyclonalIsotype
IgGResearch areas
Associated products
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Compatible Secondaries
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Control Peptide
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Immunizing Peptide (Blocking)
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Isotype control
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Recombinant Protein
Applications
Our Abpromise guarantee covers the use of ab130951 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application Abreviews Notes IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. WB Use a concentration of 1 µg/ml. Detects a band of approximately 18 kDa (predicted molecular weight: 14 kDa).Can be blocked with Human Histone H2B (glcnac S112) peptide (ab166685). ICC/IF Use a concentration of 1 µg/ml. PepArr Use a concentration of 0.2 - 0.02 µg/ml. Target
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Relevance
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. Subunit structure 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. Post-translational modification Monoubiquitination at Lys-35 (H2BK34Ub) by the MSL1/MSL2 dimer is required for histone H3 'Lys-4' (H3K4me) and 'Lys-79' (H3K79me) methylation and transcription activation at specific gene loci, such as HOXA9 and MEIS1 loci. Similarly, monoubiquitination at Lys-121 (H2BK120Ub) 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. H2BK120Ub also acts as a regulator of mRNA splicing: deubiquitination by USP49 is required for efficient cotranscriptional splicing of a large set of exons. Phosphorylation at Ser-37 (H2BS36ph) by AMPK in response to stress promotes transcription. Phosphorylated on Ser-15 (H2BS14ph) 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. GlcNAcylation at Ser-113 promotes monoubiquitination of Lys-121. It fluctuates in response to extracellular glucose, and associates with transcribed genes. 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. -
Cellular localization
Nuclear -
Database links
- Entrez Gene: 337874 Human
- Entrez Gene: 54145 Human
- Entrez Gene: 8349 Human
- Entrez Gene: 104021 Mouse
- SwissProt: P62808 Cow
- SwissProt: O60814 Human
- SwissProt: P06899 Human
- SwissProt: P23527 Human
see all -
Alternative names
- GL105 antibody
- H2B GL105 antibody
- H2B histone family member O antibody
see all
Images
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Peptide Array - Anti-Histone H2B (glcnac S112) antibody (ab130951)
All batches of ab130951 are tested in Peptide Array against peptides to different Histone H2B modifications. Six dilutions of each peptide are printed on to the Peptide Array in triplicate and results are averaged before being plotted on to a graph. Results show strong binding to Histone H2B - GlcNAc S112 peptide (ab166685), indicating that this antibody recognises the Histone H2B - GlcNAc S112 modification. This product shows
ab166685 - Histone H2B - GlcNAc S112
ab166686 - Histone H2B - unmodified
ab166688 - Histone H2B - GlcNAc S36
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Western blot - Anti-Histone H2B (glcnac S112) antibody (ab130951)Anti-Histone H2B (glcnac S112) antibody (ab130951) at 1 µg/ml + Calf Thymus Histone Preparation Lysate at 0.25 µg
Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution
Developed using the ECL technique.
Performed under reducing conditions.
Predicted band size: 14 kDa
Observed band size: 18 kDa why is the actual band size different from the predicted?
Exposure time: 4 minutes
This blot was produced using a 10% 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 5% Bovine Serum Albumin before being incubated with ab130951 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution. -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2B (glcnac S112) antibody (ab130951)IHC image of ab130951 staining Histone H2B (glcnac S112) in human colon formalin fixed paraffin embedded tissue sections, performed on a Leica Bond. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab130951, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. No primary antibody was used in the negative control (shown on the inset).
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.
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Immunocytochemistry/ Immunofluorescence - Anti-Histone H2B (glcnac S112) antibody (ab130951)ICC/IF image of ab130951 stained MCF-7 cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab130951 at 1µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- rabbit (ab96899) IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
Protocols
Datasheets and documents
References (3)
ab130951 has been referenced in 3 publications.
- Hirosawa M et al. Novel O-GlcNAcylation on Ser(40) of canonical H2A isoforms specific to viviparity. Sci Rep 6:31785 (2016). PubMed: 27615797
- Gagnon J et al. Undetectable histone O-GlcNAcylation in mammalian cells. Epigenetics 10:677-91 (2015). PubMed: 26075789
- Rønningen T et al. Pre-patterning of differentiation-driven nuclear lamin A/C-associated chromatin domains by GlcNAcylated histone H2B. Genome Res N/A:N/A (2015). ChIP ; Human . PubMed: 26359231
Images
-
Peptide Array - Anti-Histone H2B (glcnac S112) antibody (ab130951)
All batches of ab130951 are tested in Peptide Array against peptides to different Histone H2B modifications. Six dilutions of each peptide are printed on to the Peptide Array in triplicate and results are averaged before being plotted on to a graph. Results show strong binding to Histone H2B - GlcNAc S112 peptide (ab166685), indicating that this antibody recognises the Histone H2B - GlcNAc S112 modification. This product shows
ab166685 - Histone H2B - GlcNAc S112
ab166686 - Histone H2B - unmodified
ab166688 - Histone H2B - GlcNAc S36
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Western blot - Anti-Histone H2B (glcnac S112) antibody (ab130951)Anti-Histone H2B (glcnac S112) antibody (ab130951) at 1 µg/ml + Calf Thymus Histone Preparation Lysate at 0.25 µg
Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution
Developed using the ECL technique.
Performed under reducing conditions.
Predicted band size: 14 kDa
Observed band size: 18 kDa why is the actual band size different from the predicted?
Exposure time: 4 minutes
This blot was produced using a 10% 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 5% Bovine Serum Albumin before being incubated with ab130951 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution. -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H2B (glcnac S112) antibody (ab130951)
IHC image of ab130951 staining Histone H2B (glcnac S112) in human colon formalin fixed paraffin embedded tissue sections, performed on a Leica Bond. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab130951, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. No primary antibody was used in the negative control (shown on the inset).
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.
-
Immunocytochemistry/ Immunofluorescence - Anti-Histone H2B (glcnac S112) antibody (ab130951)ICC/IF image of ab130951 stained MCF-7 cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab130951 at 1µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- rabbit (ab96899) IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
