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Mca-APK（Dnp）ACE2荧光底物
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货号 | 13555 | 存储条件 | 在零下15度以下保存, 避免光照 |
| 规格 | 1 mg | 价格 | 2412 | |
| Ex (nm) | 322 | Em (nm) | 381 | |
| 分子量 | 696.67 | 溶剂 | DMSO | |
| 产品详细介绍 | ||||
简要概述
ACE2（血管紧张素转换酶2）是一种金属蛋白酶，需要将二价阳离子置于活性位点才能进行催化。它具有围绕其三价功能的多种生理作用：肾素-血管紧张素系统的负调节剂，氨基酸转运的促进剂以及严重的急性呼吸系统综合症冠状病毒（SARS-CoV）和SARS-CoV-2受体。ACE2最近被确定为SARS-CoV-2受体，是冠状病毒疾病的传染源，在免疫，炎症，ACE2和心血管疾病之间提供了关键的联系。尽管与SARS-CoV具有密切的进化关系，但SARS-CoV-2的受体结合域在几个关键氨基酸残基上有所不同，从而与人ACE2受体具有更强的结合亲和力，这可能是SARS-CoV-2致病性更大的原因。由SARS-CoV-2结合后，ACE2功能的丧失是由内吞作用以及蛋白水解切割和加工的激活驱动的。ACE2系统是防止心力衰竭（包括心肌梗塞和高血压）以及肺部疾病和糖尿病的重要保护途径。ACE2控制肠营养不良和血管通透性已成为肺动脉高压和糖尿病性心血管并发症的重要机制。Mca-APK（Dnp）ACE2底物是一种FRET肽，用于测量细胞和组织中的酶活性。它使用DNP作为淬灭剂分子来淬灭甲氧基香豆素（Mca）的荧光。当酶切割脯氨酸-赖氨酸残基并恢复Mca的荧光时，这种相互作用被消除。这种荧光底物比常用的基于HPLC分离的方法具有更大的灵活性和更高的通量。
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产品说明书
样品分析方案
简要概述
准备测试样品和空白样品（50 µL）
根据需要用抑制剂处理样品
加入Mca-APK（Dnp）ACE2底物工作溶液（50 µL）
在37°C下孵育样品
在Ex / Em = 320/430 nm（截止= 420 nm）下检测荧光
储备溶液配制
用适量的DMSO准备5-10 mM的储备溶液。
注意：将溶液分装后在-20°C下，避光保存。
在1 mL分析缓冲液中加入5 µL Mca-APK（Dnp）ACE2底物储备液（10 mM），制成Mca-APK（Dnp）ACE2底物工作液。
注意：应根据经验测量底物的适当浓度。
注意：最终浓度为50 µM。
注意：100 mM含1 M NaCl，pH约7.5的Tris-HCl可用作检测缓冲液。也可以将600 µM的ZnCl2添加到缓冲液中。
操作步骤
- 准备并添加50 µL测试样品和空白对照。
- 向样品和空白样品中加入50 µL Mca-APK（Dnp）ACE2底物工作溶液。
- 在37°C下孵育样品。
- 用荧光酶标仪在Ex / Em = 320/430 nm（截止= 420 nm）下检测荧光增加。
对于动力学读数：立即开始连续不断地测量荧光强度，并每5分钟记录一次数据，持续30-120分钟。
对于终点读数：在避光的条件下，将反应在所需温度下孵育30至120分钟。然后检测荧光强度
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