Indirect immunoperoxidase staining of human colon adenocarcinoma paraffin tissue section with ab9221. Dilution 1:50 and microwave pretreatment. Specific staining of the epithelial tumor cells. No reactivity in connective tissues.

Immunohistochemistry on a frozen tissue sections of human kidney  with ab9221 (1:500).

Indirect immunoperoxidase staining of human small intestine paraffin tissue section with ab9221. Dilution 1:50 and microwave pretreatment. Specific staining of the epithelial cells. No reactivity in the connective tissues.

Immunohistochemistry on frozen tissue sections of human colon with ab9221 (1:500).

Indirect immunoperoxidase staining of human placenta paraffin tissue section with ab9221. Dilution 1:50 and microwave pretreatment. Specific staining of the epithelial cells. No reactivity in the connective tissues.

ab9221 staining Cytokeratin 19 in human skin (sweat glands).
Left panel: with primary antibody at 1 ug/ml. Right panel: isotype control.
Sections were stained using an automated system (DAKO Autostainer Plus ), at room temperature: sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffers citrate pH6.1 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.

Overlay histogram showing MCF-7 cells stained with ab9221 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab9221, 1µg/1x10⁶ cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in MCF-7 cells fixed with methanol (5 min)/permeabilized in 0.1% PBS-Triton used under the same conditions.

ab9221 staining cytokeratin 19 in three-day-old Zebrafish tissue by Immunohistochemistry (Frozen sections).

Immunohistochemistry on frozen section of human liver bile duct epithelium