Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human testicular seminoma tissue labelling DHX36 with ab70269 at 1/1000 (1µg/ml). Detection: DAB.
All lanes : Anti-DHX36 antibody (ab70269) at 0.1 µg/mlLane 1 : HeLa whole cell lysate at 50 µgLane 2 : HeLa whole cell lysate at 15 µgLane 3 : 293T whole cell lysate at 50 µgLane 4 : NIH3T3 whole cell lysate at 50 µgdeveloped using the ECL technique
Immunoprecipitation of HeLa whole cell lysate (1mg) using ab70269 at 3 µg/mg (lane 1) or control IgG (lane 2). 1/4 of the immunoprecipitate was loaded on to a gel, analysed by Western blot and revealed using ab70269 at 0.1 µg/ml.
ICC/IF image of ab70269 stained HeLa 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 (ab70269, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 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.
IHC image of ab70269 staining in Human testis 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 ab70269, 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.