IHC image of ab4630 staining Histone H2B (acetyl K85) 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 ab4630, 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. 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.
Performed under reducing conditions.Rabbit polyclonal to Histone H2B acetyl K85 (ab4630). All lanes contain 1ug of calf thymus Histone prep.Lane 1: ab4630 at 1/500Lane 2: ab4630 at 1/1000Lane 3: ab4630 at 1/500 + 2ug acetyl K85 blocking peptide (ab10121) Lane 4: ab4630 at 1/1000 + 2ug acetyl K85 blocking peptide (ab10121)Secondary antibody: Goat anti-rabbit (HRP) - ab6721 at 1/2000.
ICC/IF image of ab4630 stained human MCF7 cells. The cells were methanol fixed (5 min), permabilised in 0.1% PBS-Tween (20 min) and incubated with the antibody (ab4630, 5µg/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).