Easy Detection Of Phosphorylated Substances: Phos-tag™

Source: Wako Laboratory Chemicals

Phos-tag™ phosphate-binding tag

Phos-tag™ is a novel phosphate-binding tag at neutral pH* (physiological pH) and was developed by Department of Functional Molecular Science at Hiroshima University. Wako Distributes specific electrophoretic procedure for the simultaneous analysis of a phosphoprotein isoform and its non-phosphorylated counterpart Manganese (II)-Phos-tag™ SDS-PAGE using thePhos-tag™ technologies are innovative methods using Pho-tag™ Agarose for separation, purification and enrichment of phosphorylated substances.

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Phos-tag™ Acrylamide provides a phosphate affinity SDS-PAGE for mobility shift detection of
phosphorylated protein. This method requires only a general mini-slab PAGE system.

  • Radioactivity is avoided.
  • Downstream procedure such as Western blot analysis and MS analysis are applicable.
  • This method can identify the time-course ratio of phosphorylated and non-phosphorylated proteins.

Phos-tag™ Agarose (AMP2- binding site = 3 -5 μmol / mL-gel) provides an efficient procedure for
separation and concentration of native phosphoproteins and phosphorylated peptides from
biological sample at physiological pH. It corresponds to a variety of column scales and
applications depending on the amount and the purpose.

  • The buffer contains no reducing agent and no detergent.
  • The procedure is almost the same as that for the general affinity column chromatography.
  • Phos-tag™ Agarose captures inorganic phosphate (HOPO3
  • 2-) and various phosphate dianions (ROPO₃²‾) bound to amino acid (Tyr, Thr, Ser, Asp, His, etc.), sugars, and lipids.

Phos-tag™ Biotin BTL-104 and BTL-105 provide a sensitive method for detection of
phosphorylated protein on PVDF membrane. BTL-111 has a long hydrophilic spacer, it has higher sensitivity than BTL-104.

  • Blocking treatment of PVDF membrane is not necessary.
  • Downstream procedure such as antibody reproving and MS analysis are applicable.
  • The procedure is similar to those using a HRP-conjugated antibody.

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