钙离子荧光探针Fluo3AM 超纯级 CAS 121714-22-5

钙离子荧光探针Fluo-3, AM是美国AAT生产的用于钙通量测定的试剂，钙通量测定是用于筛选G蛋白偶联受体（GPCR）的药物发现中的优选方法。 我们的Quest Fluo-8 和Rhod-4 系列钙检测试剂是亮的绿色和红色钙指示剂。 我们的其他钙指标如Fluo-3，Fura-2，Indo-1，Rhod-5N和Rhod-2 AM也具有与市场上相比高的质量。百萤生物是AAT Bioquest 的中国代理商，为您提供优质的钙离子荧光探针。

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钙离子篇：时间轴式讲解应用于钙离子检测的探针

实验方案

储备溶液配制

在高质量无水DMSO制备2至5 mM 的Fluo-3 AM中储备溶液。

工作溶液配制

Fluo-3 AM工作溶液
1.在实验当天，将Fluo-3AM溶解在DMSO中，或将指示剂储备溶液的等分试样解冻至室温。

2.在含有0.04%Pluronic®F-127的缓冲液（例如Hanks和Hepes缓冲液）中制备2至20µM Fluo-3 AM工作溶液。对于大多数细胞系，建议使用最终浓度为4-5μM的Fluo-3 AM。细胞负载所需指示剂的确切浓度请根据经验确定。

注：非离子洗涤剂Pluronic F-127用于提高Fluo-3 AM的水溶性。可从百萤购买各种Pluronic F-127溶液。
注：如果你的细胞含有有机阴离子转运蛋白，可以将丙磺舒（1-2mM）添加到染料工作溶液中（最终井内浓度为0.5-1mM），以减少酯化指示剂的泄漏。多种形式的丙磺舒产品，包括水溶性、钠盐和稳定溶液，可从百萤购买。

操作步骤

以下是我们推荐的将AM酯加载到活细胞中的方案。本方案仅提供指南，实际应根据您的具体需求进行修改。

1.在生长培养基中培养细胞过夜。
2.第二天，将1X Fluo-3 AM工作溶液添加到孔板中。
注意：如果你的化合物干扰血清，在染色前用新鲜的HHBS缓冲液代替生长培养基。
3.将加入染料的孔板在37°C的细胞培养箱中培养30至60分钟。
注：将染料培养2小时以上可以提高某些细胞系的信号强度。
4.用HHBS或您选择的缓冲液（含有阴离子转运蛋白抑制剂，如1mM丙磺舒）代替染料工作溶液，以去除任何多余的探针。
5.根据需要添加刺激物，并同时使用配备有FITC滤波片组的荧光显微镜或包含可编程液体处理系统（如FDSS、FLIPR或FlexStation）的荧光酶标仪在490/525nm，Cutoff=515nm处检测荧光强度。

图示

将U2OS细胞以40,000个细胞/100μL/孔接种过夜，置于96孔黑色壁/透明底板中。 除去生长培养基，将细胞分别与100μLFluo-3 AM，Fluo-4 AM和Fluo-8 AM在HHBS中以4μM浓度在37℃，5％CO 2中温育。 孵化器1小时。 用200μLHHBS洗涤细胞两次，然后使用FITC通道用荧光显微镜（Olympus IX71）成像。

试剂应用文献

AMPA receptors in the synapse turnover by monomer diffusion
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Discrimination of Dormant and Active Hematopoietic Stem Cells by G0 Marker Reveals Dormancy Regulation by Cytoplasmic Calcium
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Ketamine Increases Proliferation of Human iPSC-Derived Neuronal Progenitor Cells via Insulin-Like Growth Factor 2 and Independent of the NMDA Receptor
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MRGPRX4 is a bile acid receptor for human cholestatic itch
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TRPA1-dependent reversible opening of tight junction by natural compounds with an $\alpha$, $\beta$-unsaturated moiety and capsaicin
Authors: Kanda, Yusuke and Yamasaki, Youhei and Sasaki-Yamaguchi, Yoshie and Ida-Koga, Noriko and Kamisuki, Shinji and Sugawara, Fumio and Nagumo, Yoko and Usui, Takeo
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A new electro-optical approach for conductance measurement: an assay for the study of drugs acting on ligand-gated ion channels
Authors: Menegon, A and Pitassi, S and Mazzocchi, N and Redaelli, L and Rizzetto, R and Roll and JF and Poli, C and Imberti, M and Lanati, A and Grohovaz, F
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Altered spontaneous calcium signaling of in situ chondrocytes in human osteoarthritic cartilage
Authors: Gong, Xiaoyuan and Xie, Wenbin and Wang, Bin and Gu, Lingchuan and Wang, Fuyou and Ren, Xiang and Chen, Cheng and Yang, Liu
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Bystander effects elicited by single-cell photo-oxidative blue-light stimulation in retinal pigment epithelium cell networks
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Bystander effects elicited by single-cell photo-oxidative blue-light stimulation in retinal pigment epithelium cell networks
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Journal: Cell Death Discovery (2017): 16071

High-throughput screen detects calcium signaling dysfunction in typical sporadic autism spectrum disorder
Authors: Schmunk, Galina and Nguyen, Rachel L and Ferguson, David L and Kumar, Kenny and Parker, Ian and Gargus, J Jay
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