Synthetic peptide within Human Histone H3 aa 1-100 (N terminal) (citrulline R2) conjugated to Keyhole Limpet Haemocyanin (KLH). The exact sequence is proprietary. Also SwissProt: P84243, Q71DI3, Q16695, Q6NXT2. Database link: P68431
This antibody gave a positive signal in HL60 Treated with DMSO and Ca Ionophore whole cell lysate.
ICC/IF - HeLa cells
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 7 µg/ml.
For better results, cells should be fixed with methanol
Use a concentration of 1 µg/ml. Detects a band of approximately 18 kDa (predicted molecular weight: 15 kDa).
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- modifications of histones, also called histone code, and nucleosome remodeling.
P68431 and Q71DI3 are expressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation.
Q16695 is expressed in tesicular cells.
Q6NXT2 is specifically expressed in the seminiferous tubules of testis and this Hominid-specific H3.5/H3F3C preferentially colocalizes with euchromatin, and it is associated with actively transcribed genes.
P84243 is a variant histone H3 which replaces conventional H3 in a wide range of nucleosomes in active genes. Constitutes the predominant form of histone H3 in non-dividing cells and is incorporated into chromatin independently of DNA synthesis. Deposited at sites of nucleosomal displacement throughout transcribed genes, suggesting that it represents an epigenetic imprint of transcriptionally active chromatin.
Citrullination at Arg-9 (H3R8ci) and/or Arg-18 (H3R17ci) by PADI4 impairs methylation and represses transcription.
ab174992 staining Histone H3 (citrulline R2) in HeLa cells. The 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 ab174992 at 5µg/ml and ab7291 (anti beta Tubulin) at 1µg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat anti-rabbit AlexaFluor®488 secondary antibody (ab150081) at 2 μg/ml (shown in green) and a goat anti-mouse AlexaFluor®594 (ab150120) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labelled in blue with DAPI.
Negative controls: 1– Rabbit primary antibody 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.
Western blot - Anti-Histone H3 (citrulline R2) antibody (ab174992)
All lanes : Anti-Histone H3 (citrulline R2) antibody (ab174992) at 1 µg/ml
Lane 1 : HL60 (Human promyelocytic leukemia cell line) Whole Cell Lysate Lane 2 : HL60 (Human Caucasian promyelocytic leukaemia) DMSO and Calcium Lonophore treated Whole Cell Lysate
Lysates/proteins at 10 µg per lane.
Secondary All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 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 2% Bovine Serum Albumin before being incubated with ab174992 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution ab133406