Histone H2A Antibody (Acetyl-Lys5/7/11) (OADC00261)

Data Sheet
 
Product Number OADC00261
Product Page www.avivasysbio.com/histone-h2a-antibody-acetyl-lys5-7-11-oadc00261.html
Name Histone H2A Antibody (Acetyl-Lys5/7/11) (OADC00261)
Conjugation Unconjugated
NCBI Gene Id 3012; 317772; 8335; 8338
Host Rabbit
Clonality Polyclonal
Concentration 1,09 mg/mL
Gene Full Name histone cluster 1, H2ae
Alias Symbols H2A.1, H2A.2, H2A/a, H2AC4, H2AFA, HIST1H2AE
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 H2A family. Transcripts from this gene lack polyA tails; instead, they contain a palindromic termination element. This gene is found in the large histone gene cluster on chromosome 6p22-p21.3.
Reconstitution and Storage Store at -20C. Avoid repeated freeze/thaw cycles.
Datasheets/Manuals Printable datasheet for anti-HIST1H2AE (OADC00261) antibody
Specificity Polyclonal antibody raised in rabbit against the region of histone H2A.Z containing the acetylated lysines 5, 7 and 11, using a KLH-conjugated synthetic peptide. 
Application Info CHIP: 1 ug per IP
ELISA: 1:500
Dot Blotting: 1:20,000
Western Blotting: 1:1,000
Immunofluorescence: 1:500
Protein Name histone H2A type 1-B/E
Purification Affinity purified
Target Post-Translational Modification Acetyl-Lys5/7/11
Preservative 0.05% Sodium Azide and 0.05% ProClin 300.
Gene Symbol HIST1H2AE
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 antibody directed against H2A.Zacr. Figure 1.A ChIP assays were performed using human HeLa cells, the Aviva antibody against H2A.Zac and optimized PCR primer pairs for qPCR. ChIP was performed with the ""Auto Histone ChIP-seq" kit on the IP-Star automated system, using sheared chromatin from 1,000,000 cells. A titration consisting of 1, 2, 5 and 10 ug of antibody per ChIP experiment was analyzed. IgG (2 ug/IP) was used as a negative IP control. Figure 1.B ChIP assays were performed using human K562 cells, the Aviva antibody against H2A.Zac and optimized PCR primer sets for qPCR. ChIP was performed on sheared chromatin from 100,000 cells. A titration of the antibody consisting of 0.2, 0.5, 1 and 2 ug per ChIP experiment was analysed. IgG (1 ug/IP) was used as negative IP control. Quantitative PCR was performed with primers specific for the promoter of the active genes CCT5 and EIF4A2, used as positive controls, and for the coding region of the inactive MYT1 gene and the Sat2 satellite repeat, 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
K562 cells
ChIP-seq results obtained with the Aviva antibody directed against H2A.Zacr. ChIP was performed on sheared chromatin from 100,000 K562 cells using 1 ug the Aviva antibody against H2A.Zac. The IP'd DNA was subsequently analysed with an Illumina Genome Analyzer. Library preparation, cluster generation and sequencing were performed according to the manufacturer's instructions. The 36 bp tags were aligned to the human genome using the ELAND algorithm. Figure shows the peak distribution along the complete sequence and a 1.5 Mb region of the X-chromosome and in two regions surrounding the EIF4A2 and CCT5 positive control genes, respectively.
Image 3
H2A.Zacr
Cross reactivity tests using the Aviva antibody directed against H2A.Zacr. To test the cross reactivity of the Aviva antibody against H2A.Zac, a Dot Blot analysis was performed with peptides containing different multiple acetylations and the unmodified H2A.Z. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:20,000. Figure 4 shows a high specificity of the antibody for the modification of interest.
Image 4
H2A.Zac
Determination of the antibody titer. To determine the titer of the antibody, an ELISA was performed using a serial dilution of the Aviva antibody against H2A.Zac. The antigen used was a peptide containing the histone modifications of interest. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:56,600.
Image 5
Hela cells
Immunofluorescence using the Aviva antibody directed against H2A.Zacr. HeLa cells were stained with the Aviva antibody against H2A.Zac and with DAPI. Cells were fixed with 4% formaldehyde for 10' and blocked with PBS/TX-100 containing 5% normal goat serum and 1% BSA. The cells were immunofluorescently labeled with the H2A.Zac antibody (left) diluted 1:500 in blocking solution followed by an anti-rabbit antibody conjugated to Alexa488. The middle panel shows staining of the nuclei with DAPI. A merge of the two stainings is shown on the right.
Image 6
Human Hela cells
Western blot analysis using the Aviva antibody directed against H2A.Zacr. Western blot was performed on whole cell extracts (25 ug, lane 1) from HeLa cells, and on 1 ug of recombinant histone H2A, H2B, H3 and H4 (lane 2, 3, 4 and 5, respectively) using the Aviva antibody against H2A.Zac. The antibody was diluted 1:1,000 in TBS-Tween containing 5% skim milk. The position of the protein of interest is indicated on the right, the marker (in kDa) is shown on the left.
 

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