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Keratin isolated from the human breast carcinoma cell line MCF-7.
Cytokeratins are a subfamily of intermediate filament proteins and are characterized by a remarkable biochemical diversity, represented in epithelial tissues by at least 20 different polypeptides. They range in molecular weight between 40 kDa and 68 kDa and isoelectric pH between 4.9 – 7.8. The individual cytokeratin polypeptides are numbered 1 to 20. The various epithelia in the human body usually express cytokeratins which are not only characteristic of the type of epithelium, but also related to the degree of maturation or differentiation within an epithelium. Cytokeratin subtype expression patterns are used to an increasing extent in the distinction of different types of epithelial malignancies. The cytokeratin antibodies are not only of assistance in the differential diagnosis of tumors using immunohistochemistry on tissue sections, but are also a useful tool in cytopathology and flow cytometric assays.
Our Abpromise guarantee covers the use of ab9023 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Flow Cyt||Use 2µg for 106 cells. ab170190-Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.|
|IHC-P||Use a concentration of 4 µg/ml.|
|ICC||Use at an assay dependent concentration.|
|IHC-Fr||1/100 - 1/200. For human colon 20min ice cold acetone fixation was carried out with 60min incubation with ab9023 at 37C.|
|WB||1/100 - 1/1000.|
The image shows different concentration of total protein from normal and tumour colon tissues.
This picture was kindly supplied as part of the review submitted by Semona Rupchand.
ab9023 staining MCF-7 breast cancer cells, grown in 8 well chamber slides, by ICC/IF. Cells were fixed with Acetone for 20 minutes at -20°C and permabilized with 0.1% Triton X-100 for 5 minutes at 4°C prior to blocking in 1% fish gelatin for 1 hour at RT. The primary antibody was diluted 1/100 and incubated with the sample for 1 hour. An Alexa Fluor® 488 conjugated donkey anti-mouse antibody diluted 1/1000 was used as the secondary. Slides were mounted using mounting medium containing DAPI.
Ab9023 staining cells from a human pancreatic cancer cell line by ICC/IF. The cells were PFA fixed and permeabilized in 0.1% Triton X-100 prior to blocking in 1% BSA for 30 minutes at 22°C. Ab9023 was diluted 1/100 and incubated with the sample for 1 hour at 22°C. An Alexa Fluor® 488 conjugated donkey anti-mouse antibody, diluted 1/1000, was used as the secondary.
This image is a courtesy of Anonymous Abreview
Overlay histogram showing MCF-7 cells stained with ab9023 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.5% PBS-Tween 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 (ab9023, 2µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H&L) (ab96879) at 1/250 dilution for 30 min at 22ºC. 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 0.5% PBS-Tween used under the same conditions.