IHC image of Cytokeratin 15 staining in a formalin fixed, paraffin embedded normal human breast tissue section*, performed on a Leica Bond™ 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 ab80522, 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.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

ab80522 stained in A431cells. The cells were fixed with 4% paraformaldehyde (10min) at room temperature and incubated with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% triton for 1 hour at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab80522 at 5 µg/ml and ab6046 (Rabbit polyclonal to beta tubulin) at 1ug/ml overnight at +4°C. The secondary antibodies were ab150117 (colored green) used at 1 ug/ml and ab150087 (pseudo-colored red) used at 2 µg/ml for 1 hour at room temperature. DAPI was used to stain the cell nuclei (colored blue) at a concentration of 1.43 µM for 1 hour at room temperature.

ab80522 staining Cytokeratin 15 in mouse colon tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde; antigen retrieval was by heat mediation. Samples were incubated with primary antibody (undiluted) for 10 hours.

See Abreview

ab8969 staining keratin 15 in rat hair folicles by immunocytochemistry/ immunofluorescence. Cells were paraformaldehyde fixed and permeabilized in 0.25% Triton X-100 prior to blocking in 1% BSA for 10 minutes at 21°C. The primary antibody was diluted 1/100 and incubated with the sample for 1 hour at 21°C. A TRITC conjugated mouse anti-mouse antibody, diluted 1/85, was used as the secondary.

See Abreview