Known to detect the recombinant form of human Histone H2A BBD. However, no reaction has yet been demonstrated against the endogenous protein in histone preparations. We believe that this is because of the low abundance of this variant in such preparations.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 3 µg/ml. Staining may not be specific for H2A.Bbd (see image).
1/500. Detects a band of approximately 12.5 kDa (predicted molecular weight: 12.5 kDa). As the WB shows, we can not find any reaction with any of the different histone extracts that we have used from different sources (calf, human, chicken). However, the antibody is very specific against the recombinant form of the protein.
Use at an assay dependent dilution. PubMed: 20943707
Atypical histone H2A which can replace conventional H2A in some nucleosomes and is associated with active transcription and mRNA processing. 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. Nucleosomes containing this histone are less rigid and organize less DNA than canonical nucleosomes in vivo. They are enriched in actively transcribed genes and associate with the elongating form of RNA polymerase. They associate with spliceosome components and are required for mRNA splicing. May participate in spermatogenesis.
Present in mature sperm.
Belongs to the histone H2A family.
The docking domain is responsible for the weaker heterodimerization with H2B.
Nucleus. Chromosome. Associated with the active X chromosome and with autosomes, while it is absent from the inactive X chromosome and excluded from Barr bodies.
Western blot - Anti-Histone H2A-Bbd antibody (ab4175)Juan Ausio
CE= chicken erythrocyte (10 and 2 micrograms) , HE= HeLa cells (10 micrograms) and 2A*= recombinant H2ABbd (the numbers underneath the lanes are the number of nanograms loaded in each lane).
The primary antibody dilution was 1:500 and the secondary was 1:3000.
As the WB shows, we can not find any reaction with any of the different histone extracts that we have used from different sources (calf, human, chicken). However, the antibody is very specific against the recombinant form of the protein.
Immunocytochemistry - Anti-Histone H2A-Bbd antibody (ab4175)This image is courtesy of Petra Hajkova, University of Cambridge
Mouse embryonic fibroblasts (MEFs) were fixed in 4%PFA, permeabilised in PBS 1%BSA 0.1%TX-100 and incubated with ab4175 (anti-H2A.Bbd) antibody o/n. The antibody appears to show nucleolar staining. The antibody gave the same staining pattern in different cell types. The dilution used was 3ug/ml.
Nashun B et al. Changes in the nuclear deposition of histone H2A variants during pre-implantation development in mice. Development137:3785-94 (2010).
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