RH110基质酶 活性检测试剂盒 荧光法、Rh110 Matriptase Activity Assay Kit *Fluorimetric*详细如下: Rh110 Matriptase 检测试剂盒为筛选酶抑制剂和激活剂或使用荧光底物连续检测酶活性提供了一种方便的检测方法。在matriptase 裂解后,该底物产生Rh110 (Rhodamine 110) 荧光团,该荧光团具有亮绿色荧光,可在激发/发射=490 nm/520 nm 时检测到。Rh110 的更长波长的光谱和更高的消光系数提供了更高的灵敏度和更少的其他反应组分的干扰。该测定可以检测低至 0.15 ng/mL 的活性基质酶。 Matriptase(也称为 MT-SP1、ST14、TADG-15 和上皮蛋白)是来自 II 型跨膜丝氨酸蛋白酶家族的胰蛋白酶样蛋白酶。已知的基质蛋白酶底物谱包括细胞外基质蛋白、细胞粘附分子、离子通道、生长因子样蛋白和其他蛋白酶。它的作用可导致蛋白质加工、活化或降解。Matriptase 在几乎所有上皮细胞中广泛表达,并且在上皮来源的肿瘤中特别发现。 Add test compounds and diluted enzyme solution to the microplate wells. The suggested volume of enzyme solution for a 96-well plate is 40 µL/well and test compound is 10 µL/ well. Simultaneously set up the following control wells, as deemed necessary: Positive control contains the enzyme without test compound. Inhibitor control contains Matriptase enzyme and leupeptin. Vehicle control contains Matriptase enzyme and vehicle used in delivering test compound (e.g. DMSO, concentration not to exceed 1%). Test compound control contains assay buffer and test compound. Some test compounds have strong autofluorescence and may give false results. Substrate control contains assay buffer. Use the assay buffer to bring the total volume of all controls to 50 µL. Add 50 µL of Matriptase substrate solution into each well. For best accuracy, it is advisable to have the substrate solution equilibrated to the assay temperature (37ºC usually gives higher signal-to-background ratio). Mix the reagents completely by shaking the plate gently for 30 sec. For kinetic reading: Immediately start measuring fluorescence intensity at Ex/Em=490 nm/520 nm continuously and record data every 5 min. for 30 to 60 min. For end-point reading: Incubate the reaction at 37ºC for 30 to 60 min. Keep plate from direct light. Measure fluorescence intensity at Ex/Em=490 nm/520 nm.