Zymography is an electrophoretic technique, based on SDS-PAGE, that includes an enzyme substrate copolymerized with the polyacrylamide gel. Samples are prepared in Zymogram Sample Buffer without boiling to preserve the structure and activity of the enzyme. Following electrophoresis, the SDS is removed from the gel by washing in Zymogram Renature Buffer that contains a non-ionic detergent. The gels are then equilibrated in Zymogram Developing Buffer, which contains the divalent metal cation required for enzymatic activity. The zymogram is subsequently stained (commonly with Amido Black or Coomassie Brilliant Blue), and areas of digestion appear as clear bands against a darkly stained background where the substrate has been degraded by the enzyme.
• Zymogram Sample Buffer [2X]: 62.5mM Tris, 4% SDS, 25% Glycerol, 0.01% Bromophenol Blue, pH 6.8
• Zymogram Renature Buffer [10X]: 25% Triton® X-100
• Zymogram Development Buffer [10X]: 0.5M Tris, 2M NaCl, 50mM CaCl2, 0.2% Brij® 35, pH 7.5
- Zymogram Renature Buffer [10X]: 25% Triton® X-100