更快、更灵敏的氧化酶生成过氧化氢的酶谱检测。以二胺氧化酶为例。

Faster and sensitive zymographic detection of oxidases generating hydrogen peroxide. The case of diamine oxidase.

机构信息

Department of Chemistry & Centre CERMO-FC, Université du Québec à Montréal, CP 8888, Branch A, Montreal, Québec, H3C 3P8, Canada.

Department of Biochemical Sciences "A. Rossi-Fanelli", Sapienza University of Rome, 00185, Rome, Italy.

出版信息

Anal Biochem. 2022 Jul 1;648:114676. doi: 10.1016/j.ab.2022.114676. Epub 2022 Mar 29.

DOI:10.1016/j.ab.2022.114676

PMID:35364058

Abstract

The existing zymography method for the detection of diamine oxidase (DAO) activity has been improved by a new staining procedure with the aim to ameliorate its sensitivity. Both procedures used SDS-PAGE gels containing uniformly distributed entrapped peroxidase (that wouldn't migrate during electrophoresis). The new approach with 3,5-dichloro-2-hydroxybenzenesulfonate (DCHBS) as peroxidase substrate and with 4-amino-antipyrine as color stabilizer allows a more sensitive detection of DAO when compared to the previously reported o-phenylenediamine (o-PDA) as peroxidase substrate. The newly improved method appears faster, simple and environmentally friendly. It can be used for most of oxidases releasing hydrogen peroxide as reaction product.

摘要

现有的用于检测二胺氧化酶（DAO）活性的酶谱法通过一种新的染色方法得到了改进，目的是提高其灵敏度。两种方法均使用 SDS-PAGE 凝胶，其中包含均匀分布的固定化过氧化物酶（在电泳过程中不会迁移）。新方法使用 3,5-二氯-2-羟基苯磺酸钠（DCHBS）作为过氧化物酶的底物，4-氨基安替比林作为显色稳定剂，与之前报道的邻苯二胺（o-PDA）作为过氧化物酶的底物相比，可更灵敏地检测 DAO。新改进的方法更快、更简单、更环保。它可用于大多数氧化酶，这些氧化酶将过氧化氢作为反应产物释放。