All batches of ab130951 are tested in Peptide Array against peptides to different Histone H2B modifications. Six dilutions of each peptide are printed on to the Peptide Array in triplicate and results are averaged before being plotted on to a graph. Results show strong binding to Histone H2B - GlcNAc S112 peptide (ab166685), indicating that this antibody recognises the Histone H2B - GlcNAc S112 modification. This product shows <20% cross reactivity in peptide array with the unmodified peptide and does not cross react with GlcNAc S36, GlcNAc T101 or GlcNAc S47.ab166685 - Histone H2B - GlcNAc S112ab166686 - Histone H2B - unmodifiedab166688 - Histone H2B - GlcNAc S36
Anti-Histone H2B (glcnac S112) antibody (ab130951) at 1 µg/ml + Calf Thymus Histone Preparation Nuclear Lysate at 0.25 µgSecondaryGoat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilutiondeveloped using the ECL techniquePerformed under reducing conditions.
IHC image of ab130951 staining Histone H2B (glcnac S112) in human colon formalin fixed paraffin embedded tissue sections, performed on a Leica Bond. 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 ab130951, 1µg/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. No primary antibody was used in the negative control (shown on the inset).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.
ICC/IF image of ab130951 stained MCF-7 cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab130951 at 1µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- rabbit (ab96899) IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.