Histone H4 Antibody (Methyl-Lys20) (OADC00271)

• Product Number: OADC00271
• Name: Histone H4 Antibody (Methyl-Lys20) (OADC00271)
• Isotype: IgG1
• Conjugation: Unconjugated
• NCBI Gene Id: 121504; 554313; 8294; 8359; 8360; 8361; 8362; 8363; 8364; 8365; 8366; 8367; 8368; 8370
• Host: Mouse
• Clonality: Monoclonal
• Concentration: 2.8 mg/mL
• Gene Full Name: histone cluster 4, H4
• Alias Symbols: H4/p, H4C1, H4C2, H4C3, H4C4, H4C5, H4C6, H4C8, H4C9, H4C11, H4C12, H4C13, H4C14, H4C15, HIST4H4
• Product Format: Liquid. Provided in PBS containing 0.05% azide and 0.05% ProClin 300.
• Description of Target: Histones are basic nuclear proteins that are responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. Nucleosomes consist of approximately 146 bp of DNA wrapped around a histone octamer composed of pairs of each of the four core histones (H2A, H2B, H3, and H4). The chromatin fiber is further compacted through the interaction of a linker histone, H1, with the DNA between the nucleosomes to form higher order chromatin structures. This gene is intronless and encodes a replication-dependent histone that is a member of the histone H4 family. Transcripts from this gene lack polyA tails; instead, they contain a palindromic termination element.
• Reconstitution and Storage: Store at -20C. Avoid repeated freeze/thaw cycles.
• Specificity: Monoclonal antibody raised in mouse against histone H4 containing the monomethylated lysine 20 (H4K20me1), using a KLH-conjugated synthetic peptide. 
• Application Info: CHIP: 0,5 - 1 ug/CHIP
  Dot Blotting: 1:20.000
  Western Blotting: 1:1.000
  IF: 1:200
• Protein Name: histone H4
• Purification: Protein A purified
• Target Post-Translational Modification: Methyl-Lys20
• Preservative: 0.05% Sodium Azide and 0.05% ProClin 300.
• Gene Symbol: HIST4H4
• Predicted Species Reactivity: Human
• Application: CHIP, DB, WB, IF
• Predicted Homology Based on Immunogen Sequence: Mouse
• Image 1: Human Hela cells
  
  ChIP results obtained with the Aviva monoclonal antibody directed against H4K20me1. ChIP assays were performed using human HeLa cells, the Aviva monclonal antibody against H4K20me1 and optimized PCR primer sets for qPCR. ChIP was performed, using sheared chromatin from 1 million cells. A titration of the antibody consisting of 0.5, 1, 2 and 5 ug per ChIP experiment was analysed. IgG (1 ug/IP) was used as negative IP control. QPCR was performed with primers for the coding region of the active GAPDH and ACTB genes, used as positive controls, and for the GAPDH promoter and a region located 1 kb upstream of the GAPDH promoter, used as negative controls. Figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).
• Image 2: Human genome
  
  ChIP-seq results obtained with the Aviva monoclonal antibody directed against H4K20me1 r. ChIP was performed as described above using 0.5 ug of the Aviva antibody against H4K20me1. The IP'd DNA was subsequently analysed on an Illumina HiSeq. Library preparation, cluster generation and sequencing were performed according to the manufacturer's instructions. The 51 bp tags were aligned to the human genome using the BWA algorithm. Figure shows the peak distribution in genomic regions from chromosomes 12, 7, 14 and 3 surrounding the GAPDH, ACTB, FOS and EIF4A2 positive control genes (figure 2A, B, C and D, respectively).
• Image 3: H4K20me1
  
  Cross reactivity tests using the Aviva monoclonal antibody directed against H4K20me1. To check the specificity of the Aviva monoclonal antibody against H4K20me1 a Dot Blot was performed with peptides containing different modifications of histone H3 and H4 or the unmodified H4K20 sequence. One hundred to 0.2 pmol of peptide containing the respective histone modification were spotted on a membrane. The antibody was used at a dilution of 1:20,000. Figure 3 shows a high specificity of the antibody for the modification of interest.
• Image 4: Human osteosarcoma (U2OS) cells
  
  Immunofluorescence using the Aviva monoclonal antibody directed against H4K20me1. Human osteosarcoma (U2OS) cells were stained with the Aviva antibody against H4K20me1 and with DAPI. Cells were fixed with ice cold methanol for 10' and blocked with PBS/TX-100 containing 5% normal goat serum. Figure A: cells were immunofluorescently labeled with the H4K20me1 antibody (left), diluted 1:200 in blocking solution followed by an anti- mouse antibody conjugated to Alexa488 or with DAPI (right), which specifically labels DNA. Figure B: staining of the cells with the H4K20me1 antibody after incubation of the antibody with blocking peptide.
• Image 5: Human Hela cells
  
  Western blot analysis using the Aviva monoclonal antibody directed against H4K20me1. Western blot was performed on whole cell (25 ug, lane 1) and histone extracts (15 ug, lane 2) from HeLa cells, and on 1 ug of recombinant histone H2A, H2B, H3 and H4 (lane 3, 4, 5 and 6, respectively) using the Aviva monoclonal antibody against H4K20me1. The antibody was diluted 1:1,000 in TBS-Tween containing 5% skim milk. The marker (in kDa) is shown on the left, the position of the protein is indicated on the right