Western blot was performed on whole cell extracts (25 µg, lane 1) and histone extracts (15 µg, lane 2) from HeLa cells, and on 1 µg of recombinant histone H2A, H2B, H3 and H4 (lane 3, 4, 5 and 6, respectively) using H4pan Polyclonal Antibody(bs-53138R). The antibody was diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. The position of the protein of interest is indicated on the right; the marker (in kDa) is shown on the left.
BS-53138R Image
HeLa cells were stained with H4pan Polyclonal Antibody (bs-53138R) and with DAPI. Cells were fixed with 4% formaldehyde for 10' and blocked with PBS/TX-100 containing 1% BSA. The cells were immunofluorescently labeled with the H4 antibody (middle) diluted 1:1,000 in blocking solution followed by an anti-rabbit antibody conjugated to Alexa488. The left panel shows staining of the nuclei with DAPI. A merge of the two stainings is shown on the right.
BS-53138R Image
To determine the titer of the antibody, an ELISA was performed using a serial dilution of H4pan Polyclonal Antibody (bs-53138R) in antigen coated wells. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:3,000.
BS-53138R Image
ChIP assays were performed using HeLa cells, H4pan Polyclonal Antibody (bs-53138R) and optimized PCR primer pairs for qPCR. ChIP was performed using sheared chromatin from 1 million cells on the IP-Star Compact automated system. A titration consisting of 0.5, 1, 2 and 5 µg of antibody per ChIP experiment was analyzed. IgG (1 µg/IP) was used as a negative IP control. Quantitative PCR was performed with primers specific for the promoters of the active GAPDH and c-fos genes, and for the inactive MYOD1 gene and the Sat2 satellite repeat. Figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).
Polyclonal antibody raised in rabbit against histone H4 using a KLH-conjugated synthetic peptide containing an unmodified sequence from the C-terminal part of the protein
Description: 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.