Phos binding reagent (Phosbind) acrylamide

Phosbind acrylamide is an innovative phosphate-binding reagent which contains MnCl2. It is designed for the electrophoretic separation and detection of phosphorylated versus non-phosphorylated proteins. Operating optimally at neutral (physiological) pH, its mechanism is based on selective interaction with phosphate groups. This reagent enables protein phosphorylation analyses via SDS-PAGE, particularly suitable for protein targets within the 30–130 kDa molecular weight range(marker F4005). Compared to conventional phosphorylation detection methods, this approach reveals phosphorylation-dependent electrophoretic mobility shift without reliance on specific antibodies. For precise interpretation, optimal separation is achieved using standard Tris-glycine running buffer (K1197).

Protein phosphorylation is an important covalent post-translational modification that can alter the structural conformation of a protein, which then regulates the function, location and specific binding of the target protein. Many cellular processes are regulated by the reversible phosphorylation of proteins and 30% of the proteins are likely to be phosphorylated at some point during their existence.

The determination of the phosphorylation state of proteins is important for defining protein kinase substrates and revealing the activation state of signal transduction pathways. Methods for determining the phosphorylation status of proteins are thus have important implications in the understanding of diverse biological and pathophysiological processes, such as signal transduction pathways, cancer and other diseases.

Phos binding reagents (Phosbind) are specific and selective phosphate-binding reagents, and exhibit preferential ionic interactions with phosphorylated ions on phosphorylated proteins or peptides at neutral pH. Phos binding reagent is a dinuclear metal (Zn²⁺ or Mn²⁺) complex.

Phosbind Acrylamide is a specific reagent for separation of phosphorylated proteins using SDS-PAGE method.

The product contains Phosbind Acrylamide and MnCI2 components, of which MnCI2 is 100 mg.

1. Recognition of all phosphorylated forms of Tyr/Ser/Thr.

2. Followed by Western blotting and Mass analysis.

3. Simultaneous detection of phosphorylated/non-phosphorylated proteins using the total antibody without the anti phosphorylated protein.

4. Simply add Phos binding reagent acrylamide and MnCl2 solution* to acrylamide solution in the preparation of SDS-PAGE gel.

(*: Add 5.0 mM Phos binding reagent Acrylamide Aqueous Solution and 10 mM MnCl2 solution in equal amount in the gel).

1. M. J. Hubbard, P. Cohen. On target with a new mechanism for the regulation of protein phosphorylation. Trends Biochem. Sci. 18: 172 (1993).

2. B. Agnew et al. Compositions and methods for detection and isolation of phosphorylated molecules. US Patent # 7,102,005. (September 5, 2006).