An improved assay for proteases and polysaccharidases employing a cartilage proteoglycan substrate entrapped in polyacrylamide particles.
An improved assay for proteases and polysaccharidases employing a cartilage proteoglycan substrate entrapped in polyacrylamide particles.
复制标题
使用包埋在聚丙烯酰胺颗粒中的软骨蛋白聚糖底物对蛋白酶和多糖酶进行改进的测定。
DOI：
10.1016/0003-2697(80)90400-5
复制
发表时间：
1980
影响因子：
2.9
通讯作者：
WoessnerJr,JF
中科院分区：
文献类型：
--
作者：
Nagase,H;WoessnerJr,JF
一种灵敏的比色法测定蛋白酶和某些多糖酶是基于从牛鼻软骨中消化蛋白多糖。这种高分子量的底物被困在聚丙烯酰胺凝胶的空隙中。然后，凝胶被分散成小颗粒。酶可以扩散到这些颗粒中，消化产物也可以扩散出去。消化后，将颗粒离心，上清液中的消化产物通过与变色染料1，9-二甲基亚甲基蓝反应或用咔唑试剂测定其糖醛酸含量来定量。来自四个主要类别的每一种酶都可以在1-200毫微克的水平上定量。该方法特别适用于降解基质蛋白多糖的软骨蛋白酶的研究。
A sensitive colorimetric assay for proteases and certain polysaccharidases is based on the digestion of proteoglycan from bovine nasal cartilage. This high-molecular-weight substrate is trapped in the interstices of a polyacrylamide gel. The gel is then dispersed as small particles. Enzymes can diffuse into these particles and digestion products can diffuse out. Following digestion, the particles are centrifuged off and the digestion products in the supernatant are quantitated by reaction with the metachromatic dye 1,9-dimethylmethylene blue or by assay of their uronic acid content with carbazole reagent. Proteases from each of the four major classes can be quantitated at levels of 1–200 ng. The method is particularly suitable for the study of cartilage proteases that degrade matrix proteoglycans.