Immunohistochemistry analysis of Cdc4 showing staining in the cytoplasm and nucleus of paraffin-embedded human cervical carcinoma tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a Cdc4 polyclonal antibody (401500) diluted in 3% BSA-PBS at a dilution of 1:50 overnight at 4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.
|Tested species reactivity||Human, Mouse|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide derived from an internal region of the human and mouse Cdc4|
|Purification||Antigen affinity chromatography|
|Storage buffer||PBS, pH 7.4|
|Contains||0.1% sodium azide|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||3-5 µg/million cells|
|Immunohistochemistry (Paraffin) (IHC (P))||1:10-1:100|
|Immunoprecipitation (IP)||Assay Dependent|
|Western Blot (WB)||Assay Dependent|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
Fbw7/Cdc4 is a member of the F box protein family which are characterized by an approximately 40 amino acid motif, the F box. The F box proteins constitute one of the four subunits of ubiquitin protein ligase complex called SCFs (SKP1-cullin-F box), which function in phosphorylation-dependent ubiquitination. The F box proteins are divided into 3 classes: Fbws containing WD40 domains, Fbls containing leucine rich repeats, and Fbxs containing either different protein-protein interaction modules or no recognizable motifs. Fbw7 belongs to the Fbws class; in addition to an F box, this protein contains 7 tandem WD40 repeats. It binds directly to cyclin E and probably targets cyclin E for ubiquitin mediated degradation. Mutations of this gene are detected in ovarian and breast cancer cell lines. Alternative splicing of this gene generates 2 transcript variants diverging at the 5' termini.
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