基于凝胶的明胶酶谱法检测细胞培养中的基质金属蛋白酶活性

Gel-Based Gelatin Zymography to Examine Matrix Metalloproteinase Activity in Cell Culture.

作者信息

Chhabra Aastha, Rani Vibha

机构信息

Department of Biotechnology, Jaypee Institute of Information Technology, Noida, Uttar Pradesh, India.

Peptide and Proteomics Division, Defence Institute of Physiology and Allied Sciences (DIPAS), DRDO, Delhi, India.

出版信息

Methods Mol Biol. 2018;1731:83-96. doi: 10.1007/978-1-4939-7595-2_9.

DOI:10.1007/978-1-4939-7595-2_9

PMID:29318546

Abstract

Gelatin zymography, first described by Heussen and Dowdle in the 1980s, is a widely used valuable tool in research and diagnostics. The technique identifies gelatinases by the degradation of their preferential substrate as well as by their molecular weight (kDa). We here describe detailed methodology for the detection of pro- and active- forms of both MMP-2 (gelatinase A) and MMP-9 (gelatinase B) in cells using norepinephrine-stimulated H9c2 cardiomyoblasts as model. An easy to follow step-by-step protocol has been carefully written for reliable results. We also suggest an acceptable method for quantification of gelatin zymograms.

摘要

明胶酶谱法最早由赫森和多德尔于20世纪80年代描述，是一种在研究和诊断中广泛应用的重要工具。该技术通过明胶酶对其优先底物的降解以及它们的分子量（千道尔顿）来识别明胶酶。我们在此描述了使用去甲肾上腺素刺激的H9c2心肌母细胞作为模型来检测细胞中MMP-2（明胶酶A）和MMP-9（明胶酶B）的前体形式和活性形式的详细方法。为了获得可靠的结果，我们精心编写了一个易于遵循的分步方案。我们还提出了一种可接受的明胶酶谱定量方法。