ICC/IF image of ab77590 stained HeLa cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 0.3M glycine in 0.1% PBS-Tween (no animal sera) for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab77590 at 10µg/ml overnight at +4°C. The secondary antibody (pseudo-colored green) was Alexa Fluor® 488 donkey anti- goat (ab150133) IgG used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (pseudo-colored red) at a 1/200 dilution for 1h at room temperature. DAPI was used to stain the cell nuclei (pseudo-colored blue) at a concentration of 1.43µM for 1hour at room temperature.
ab77590 at 2.5 µg/ml staining ASS1 in Human Kidney by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Steamed antigen retrieval with citrate buffer pH 6 was performem, antibody revealed by AP-staining.
IHC image of ab77590 staining in human kidney 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 ab77590, 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.