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Applications: ChIP (1 - 2 µg/ChIP)
ELISA (1:50)
DB (1:20,000)
Assay Conditions: ChIP results obtained with the antibody directed against H2Apan
ChIP assays were performed using human HeLa cells, the antibody against H2Apan (Cat. # 25226) and optimized PCR primer sets for qPCR. ChIP was performed on sheared chromatin from 10,000 cells using the SX-8G IP-Star automated system. A titration of the antibody consisting of 1, 2, 5, and 10 µg per ChIP experiment was analyzed. IgG (5 µg/IP) was used as negative IP control. QPCR was performed with primers for the GAPDH promoter and for the inactive MYOD gene. Figure 1 shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).Determination of the titer
To determine the titer of the antibody, an ELISA was performed using a serial dilution of the antibody directed against H2Apan (Cat. # 25226), crude serum and flow-through in antigen coated wells. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:1,800. Cross reactivity test using the antibody directed against H2Apan
A Dot Blot analysis was performed to test the cross reactivity of the antibody against H2Apan (Cat. # 25226) with the peptide used for immunization of the rabbit and other peptides containing unmodified sequences of different histones. 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 3 shows a high specificity of the antibody for the H2A peptide.
Background: Histones are the main constituents of the protein part of chromosomes of eukaryotic cells. They are rich in the amino acids arginine and lysine and have been greatly conserved during evolution. Histones pack the DNA into tight masses of chromatin. Two core histones of each class H2A, H2B, H3 and H4 assemble and are wrapped by 146 base pairs of DNA to form one octameric nucleosome. Histones play a central role in the regulation of transcription, DNA repair, DNA replication and chromosomal stability. These different functions are established via a complex set of post-translational modifications which either directly or indirectly alter chromatin structure and DNA accessibility to facilitate transcriptional activation or repression or other nuclear processes.
Concentration: 50 µg/79 µl
Description: Polyclonal antibody raised in rabbit against histone H2A using a KLH-conjugated synthetic peptide containing an unmodified sequence from the C-terminal part of the protein.
Format: Aqueous buffer solution
Formulation: PBS containing 0.05% azide and 0.05% ProClin 300
Immunogen: synthetic peptide
Purification: Affinity purified
Storage Stability: Store at -80°C for up to 2 years. Centrifuge after first thaw to maximize product recovery. Aliquot to avoid repeated freeze/thaw cycles. Aliquots may be stored at -20°C for at least one month.
Uniprot: Q7L7L0
Warnings: Avoid freeze/thaw cycles.
Biosafety Level: Not applicable (BSL-1)
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