Protein Kinase A【子货号：V135A，包装：1 x 20μl，，运保温度：-20℃】
cAMP-Glo™ ONE Buffer【子货号：V166A，包装：1 x 2.5ml，，运保温度：-20℃】
MgCl2, 1M【子货号：V330A，包装：1 x 1ml，，运保温度：-20℃】
cAMP, 1mM【子货号：V642B，包装：1 x 100μl，，运保温度：-20℃】
Kinase-Glo® Substrate【子货号：V672A，包装：1 x 1 vial，，运保温度：-20℃】
Kinase-Glo® Buffer【子货号：V673A，包装：1 x 10ml，，运保温度：-20℃】
描述：
说明
cAMP-Glo™ Max Assay (cAMP-Glo™ Max 检测试剂盒) 是一种均质的、利用生物发光原理检测细胞内环腺苷酸(cAMP) 水平的高通量方法。若某种化合物能调节与腺苷酸环化酶偶联的G 蛋白偶联受体(GPCR)，则会明显地改变细胞内的cAMP 水平。cAMPGlo™ Max Assay 用于监测细胞内的cAMP 水平在以下应答反应中的改变：激动剂、拮抗剂、及受试化合物对GPCR 的效应。这种检测方法的原理是：cAMP 激活蛋白激酶A(PKA) 全酶的活性，降低ATP含量，从而引起ATP 偶联的萤光素酶反应的光信号降低。
cAMP-Glo™ Max Assay 将裂解缓冲液与cAMP 反应缓冲液合二为一，成为cAMP-Glo™ ONE Buffer (cAMP-Glo™ ONE 缓冲液)。这种新的规格简化了实验操作过程，减少了完成检测所需的时间。新的ONE Buffer 以5 倍浓度提供，为细胞培养液的起始体积提供更多的灵活性。
The cAMP-Glo™ Max Assay is a homogeneous, bioluminescent and high-throughput assay to measure cyclic AMP (cAMP) levels in cells. Compounds that modulate GPCRs coupled with adenylate cyclase typically alter intracellular cAMP levels. The cAMP-Glo™ Max Assay monitors cAMP levels in cells in response to the effect of agonists, antagonists or test compounds on G protein-coupled receptors (GPCRs). The assay is based on the principle that cyclic AMP (cAMP) stimulates protein kinase A (PKA) holoenzyme activity, decreasing available ATP and leading to decreased light production in a coupled luciferase reaction.
This improved version combines the lysis and cAMP reaction buffers into the cAMP-Glo™ ONE Buffer. This new format streamlines the protocol and reduces the time needed to complete the assay. The new ONE Buffer is supplied at a 5X concentration, which provides increased flexibility for starting cell culture volumes.
The cAMP-Glo™ Max Assay can be performed in 96-, 384- or 1536-well plates. The cells are induced with a test compound for an appropriate period of time to modulate cAMP levels. After induction, cells are lysed, and the cAMP released stimulates protein kinase A in the reagent (Figure 1). The Kinase-Glo® Reagent is then added to terminate the PKA reaction and detect the remaining ATP via a luciferase reaction. Plates are read using a microplate-reading luminometer. The half-life for the luminescent signal is greater than 4 hours allowing ample time to read the plates and eliminates the need for luminometers with reagent injectors.
Features - Benefits
Fast and Easy to Use:
Improved—Lysis and cAMP detection steps combined (cAMP-Glo™ ONE Buffer).
ONE Buffer—5X concentration provides better flexibility for starting cell culture volumes.
Assay can be completed in approximately 30 minutes. Excellent Signal-to-Noise Ratios:
Best signal:background ratio of all the cAMP assays.
Signal:Background >200 (with cAMP), >15 (on cells).
Easily scalable to 1536-well plate formats and beyond. Proven Luminescent Technology:
Powered by Ultra-Glo™ Recombinant Luciferase.
No interference by fluorescent compounds.
Non-radioactive.
Applications
High-throughput analysis of library compounds to identify modulators of GPCR activity.
Easy quantitation of cAMP levels in broad range of cell types.
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