※ 引述《sheepfeather (...)》之铭言： : 读paper看到的新名词 想搞懂他是啥技术 : 网路上资料不是很清楚 kiwi也说得很模糊 : 希望有人愿意跟我说明一下基本原理或操作流程之类的 : 或是要去哪里才可以查询到相关资料都可以 : 感谢愿意回答的各位 ~ <(_""_)> Zymography is an electrophoretic technique, based on SDS-PAGE, that includes a substrate copolymerized with the polyacrylamide gel, for the detection of enzyme activity. Samples are prepared in the standard SDS-PAGE treatment buffer but without boiling, and without a reducing agent. Following electrophoresis, the SDS is removed from the gel (or zymogram) by incubation in unbuffered Triton X-100, followed by incubation in an appropriate digestion buffer, for an optimized length of time at 37°C. 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. These protocols, however, are subject to much adjustment. For instance, D. melanogaster digestive glycosidases generally survive reducing conditions (i.e. the presence of 2-mercaptoethanol or DTT) and and to an extent heating. Indeed, the separations following heating to 50°C tend to exhibit a substantial increase in band resolution, without appreciable loss of activity. --

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