Fast track antibodies constitute a diverse group of products that have been released to accelerate your research, but are not yet fully characterized. They have all been affinity purified and show high titre values against the immunizing peptide (by ELISA).
Use at an assay dependent concentration. : This antibody gave a positive result in ELISA against the immunizing peptide (ab18504).
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
1/500. Detects a band of approximately 15 kDa (predicted molecular weight: 14 kDa).
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.
Linker histones are involved in the formation of higher order structure in chromatin and the maintenance of overall chromatin compaction. Whilst the core histones are highly conserved across a wide range of organisms, the linker histones are less conserved.
This Fast-Track antibody is not yet fully characterised. These images represent
inconclusive preliminary data.
Western blot - Anti-Histone H2B (phospho S32) antibody (ab10476)
All lanes : Anti-Histone H2B (phospho S32) antibody (ab10476) at 1/500 dilution
Lane 1 : Hela control Lane 2 : Gamma irraditaed hela 1h Lane 3 : Gamma irraditaed hela 2h Lane 4 : Hela control with Human Histone H2B (phospho S32) peptide (ab18504) at 1 µg/ml Lane 5 : Gamma irraditaed hela 1h with Human Histone H2B (phospho S32) peptide (ab18504) at 1 µg/ml Lane 6 : Gamma irraditaed hela 2h with Human Histone H2B (phospho S32) peptide (ab18504) at 1 µg/ml Lane 7 : Hela control with Human Histone H2B peptide (ab18507) at 1 µg/ml Lane 8 : Gamma irraditaed hela 1h with Human Histone H2B peptide (ab18507) at 1 µg/ml Lane 9 : Gamma irraditaed hela 2h with Human Histone H2B peptide (ab18507) at 1 µg/ml
Lysates/proteins at 25 µg per lane.
Secondary All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab6721) at 1/5000 dilution
ICC/IF image of ab10476 stained HeLa cells. The cells were 4% PFA fixed (10 min), permabilised in 0.1% PBS-Tween (20 min) and incubated with the antibody (ab10476, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal goat serum / 0.3M glycine was used to block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue). This antibody also gave a positive IF result in HepG2, Hek293 and MCF7 cells fixed in 4% PFA at 1ug/ml and Hela, Hek293, HepG2 and MCF7 cells fixed in 100% methanol at 1ug/ml. However, this Fast-Track antibody is not yet fully characterised. This image represents inconclusive preliminary data.
IHC image of ab10476 staining in Human Breast adenocarcinoma formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab10476, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
Immunocytochemistry/ Immunofluorescence - Anti-Histone H2B (phospho S32) antibody (ab10476)Image courtesy of an abreview submitted by Dr. Kirk McManus, Univ. of Manitoba/Cancer Care MICB, Canada
ab10476 (1/200) staining Histone H2B phospho S32 in Hela cells (green). Cells were fixed in paraformaldehyde, permeabilised with 0.5% Triton X100 and counterstained with DAPI in order to highlight the nucleus (red). for further experimental details please refer to Abreview.
Li J et al. Nuclear PKC-? facilitates rapid transcriptional responses in human memory CD4+ T cells through p65 and H2B phosphorylation. J Cell Sci129:2448-61 (2016).
Read more (PubMed: 27149922) »
Lau AT et al. Phosphorylation of histone H2B serine 32 is linked to cell transformation. J Biol Chem : (2011).
Read more (PubMed: 21646345) »