Phos-tag™: phosphoprotein detection, purification, separation

A series of reagents utilizing Phos-tag™ from Fujifilm Wako can be used in the electrophoretic separation of phosphoproteins by SDS-PAGE, specific detection of phosphoproteins via Western Blot, highly sensitive detection of phosphorylated compounds via mass spectrometry, purification of phosphorylated proteins and selective staining of phosphoproteins in gels.

• SuperSep Phos-tag™ is a ready-to-use precast polyacrylamide gel, which contains zinc as a supplemental metal. High reproducibility and compatibility with Bio-Rad® and Life Technologies® tanks.
• Phos-tag™ Acrylamide AAL-107 is used for the electrophoretic separation of phosphorylated proteins using a self-made acrylamide gel. The acrylamide Phos-tag™ ligand enables the detection of phosphoproteins using phosphate affinity electrophoresis. The separation is based on a conventional SDS-PAGE. The phosphorylated proteins are withheld by the interaction with the Phos-tag™ ligand incorporated into the gel. Thus, phosphorylated molecules can be separated from non-phosphorylated molecules. After the electrophoretic separation the gel can be used for further downstream analysis such as Western blotting or MALDI-TOF MS.
• Phos-tag™ Agarose is a functional molecule which is used for the separation, purification and concentration of phosphorylated proteins. It is suitable for the separation of phosphorylated peptides and native phosphoproteins from biological samples under neutral conditions. This method is based on Immobilized Metal Affinity Chromatography (IMAC) during which a phosphate-binding dinuclear zinc(II) complex attached to a highly cross-linked agarose granule (Phos-tag™ Agarose) represents the stationary phase.
• Phos-tag™ Biotin is a functional molecule which is used for the detection of phosphorylated proteins on a PVDF membrane. The specific detection of phosphorylated proteins can be achieved without any phosphor-specific antibodies using Western blot analysis. This visualization of the bands requires streptavidin-conjugated horseradish peroxidase (HRP) and chemiluminescent detection reagents.
• Phos-tag™ Fluorescent Gel Stain Reagents are applicable for SDS-PAGE gels. They are equipped with the functional molecule Phos-tag™ that captures free phosphate groups. By staining the gel after SDS-PAGE, phosphorylated proteins can be specifically detected. The products include Yellow, Magenta, Cyan and Aqua with different fluorescence wavelengths.