Westergaard J L, Hackbarth C, Treuhaft M W, Roberts R C
J Immunol Methods. 1980;34(2):167-75. doi: 10.1016/0022-1759(80)90170-2.
A simple, sensitive technique for detecting proteolytic enzyme zones on electrophoretograms by making contact print zymograms is described. The method is applicable to electrophoretograms prepared on a variety of support media, immunoelectrophoretograms or isoelectric focusing patterns on various media. The contact print zymograms are prepared by placing unfixed, unstained electrophoretograms in contact with a thin film of casein which has diffused into a layer of agarose supported by a hydrophilic polyester film. After staining the casein film with Coomassie blue, the proteolytic zones are detected as clear zones against a blue background. The method can detect as little as 9 ng of trypsin. The utility of the method is illustrated by detection of the proteinase enzymes in thermophilic actinomycete antigen preparations separated by polyacrylamide electrophoresis, crossed immunoelectrophoresis and isoelectric focusing on agarose and granulated dextran supports.
本文描述了一种通过制作接触印相酶谱图来检测电泳图谱上蛋白水解酶区域的简单、灵敏技术。该方法适用于在各种支持介质上制备的电泳图谱、免疫电泳图谱或各种介质上的等电聚焦图谱。接触印相酶谱图是通过将未固定、未染色的电泳图谱与已扩散到由亲水性聚酯膜支撑的琼脂糖层中的酪蛋白薄膜接触而制备的。用考马斯亮蓝对酪蛋白膜染色后，蛋白水解区域在蓝色背景下呈现为清晰区域。该方法能检测低至9纳克的胰蛋白酶。通过检测经聚丙烯酰胺电泳、交叉免疫电泳以及在琼脂糖和粒状葡聚糖支持物上进行等电聚焦分离的嗜热放线菌抗原制剂中的蛋白酶，说明了该方法的实用性。
