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钙离子荧光探针Fluo5N,AM

英文名称:Fluo-5N, AM *Cell permeant*
产品参数
Ex (nm)494Em (nm)516
分子量1127.92溶剂DMSO
存储条件在零下15度以下保存, 避免光照
产品概述

关键参数

仪器:荧光酶标仪

Ex:490

Em:525

Cutoff:515

推荐孔板:纯黑色

 

仪器:荧光显微镜

通道:FITC

推荐孔板:黑壁透明底

 

仪器:流式细胞仪

通道:FITC

 

产品介绍

钙离子荧光探针Fluo-5N,AM是美国AAT Bioquest生产的钙离子荧光探针,Fluo-5N是Fluo-4的类似物,具有较低的钙结合亲和力(Kd = ~90 uM),使其适用于检测1μM至1 mM范围内的细胞内钙水平,这将使Fluo-4的反应饱和。。 通过将溶解的指示剂直接加入含有培养细胞的培养皿中,可以将Fluo-5N AM酯直接加载到活细胞中。 它与氩离子激光源在488 nm激发相容,使Fluo-5N可用于共聚焦显微镜,流式细胞仪和微孔板筛选应用。 它的激发和发射波长分别为494和516 nm。 钙结合后,其荧光强度增加> 100倍。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

操作步骤

1.准备HHBS缓冲液,10%Pluronic®F-127溶液和25 mM Probenecid溶液。

2.在高质量无水DMSO中制备2 mM至5 mM Fluo-5N,AM *细胞渗透物*储备溶液。
2.1使用的Fluo-5N,AM *细胞渗透物*的量:1mg
2.2所需浓度:2 mM
2.3在合适的容器中,将1mg Fluo-5N,AM *细胞渗透物*与443.29μL无水DMSO混合。
 

3.使用10μMFluo-5N,AM *细胞渗透物* 4在HHBS中制备2X工作溶液,0.08%Pluronic®F-127和2 mM丙磺舒。
3.1终井内浓度的Fluo-5N,AM *细胞渗透物*:5μM
3.2Pluronic®F-127的终井内浓度:0.04%
3.3终孔内浓度的丙磺舒:1mM
3.4在合适的容器中混合16μLFluo-5N,AM *细胞渗透物*,25.6μL10%Pluronic®F-127和256μL25mM丙磺舒。接下来,添加HHBS或您选择的缓冲液,直到体积为3.2 mL。
注意:对于大多数细胞系,我们建议终浓度为Fluo-5N,AM *细胞渗透物*为4至5μM。
注意:推荐的Pluronic F-127井浓度终为0.02%至0.04%。
注意:推荐的终浓度为1至2.5 mM的Probenecid。
 

4.将100μL染料工作溶液加入已经含有100μL培养基的所需孔中。
4.1该步骤将染料工作溶液从2X稀释至1X,并将每种组分的终浓度调节至:5μMFluo-5N,AM *细胞渗透物*,0.04%Pluronic F-127,1mM丙磺舒。

 

5.孵育染料
5.1将染料加载板在细胞培养箱中孵育20-120分钟。
5.2将染料加载板在室温下孵育30分钟。
 

6.用1.0 mM Probenecid准备HHBS缓冲液(或您选择的缓冲液)。
6.1在合适的容器中加入160μL的25mM丙磺舒。 接下来,添加HHBS或您选择的缓冲液,直到体积为4 mL。
 

7.用HHBS缓冲液或您选择的缓冲液替换染料工作溶液,使用1.0 mM Probenecid。
7.1首先,从所需孔中除去200μL染料工作溶液和培养基。
7.2在相同的孔中加入200μL含有1.0mM丙磺舒的HHBS(或您选择的缓冲液)。
 

8.运行实验
8.1为您的样品添加所需的处理。
8.2以Ex / Em = 494/516 nm运行实验

 

参考文献

Calreticulin regulates TGF-β1-induced epithelial mesenchymal transition through modulating Smad signaling and calcium signaling
Authors: Yanjiao Wu, Xiaoli Xu, Lunkun Ma, Qian Yi, Weichao Sun, Liling Tang
Journal: The International Journal of Biochemistry & Cell Biology (2017)

Dexmedetomidine reduces hypoxia/reoxygenation injury by regulating mitochondrial fission in rat hippocampal neurons
Authors: Jia Liu, Qing Du, He Zhu, Yu Li, Maodong Liu, Shoushui Yu, Shilei Wang
Journal: Int J Clin Exp Med (2017): 6861--6868

Monosialoganglioside 1 may alleviate neurotoxicity induced by propofol combined with remifentanil in neural stem cells
Authors: Jiang Lu, Xue-qin Yao, Xin Luo, Yu Wang, Sookja Kim Chung, He-xin Tang, Chi Wai Cheung, Xian-yu Wang, Chen Meng, Qing Li
Journal: Neural Regeneration Research (2017): 945

Obtaining spontaneously beating cardiomyocyte-like cells from adipose-derived stromal vascular fractions cultured on enzyme-crosslinked gelatin hydrogels
Authors: Gang Yang, Zhenghua Xiao, Xiaomei Ren, Haiyan Long, Kunlong Ma, Hong Qian, Yingqiang Guo
Journal: Scientific Reports (2017): 41781

Di (2-ethylhexyl) phthalate-induced apoptosis in rat INS-1 cells is dependent on activation of endoplasmic reticulum stress and suppression of antioxidant protection
Authors: Xia Sun, Yi Lin, Qiansheng Huang, Junpeng Shi, Ling Qiu, Mei Kang, Yajie Chen, Chao Fang, Ting Ye, Sijun Dong
Journal: Journal of cellular and molecular medicine (2015): 581--594

The effect of mitochondrial calcium uniporter on mitochondrial fission in hippocampus cells ischemia/reperfusion injury
Authors: Lantao Zhao, Shuhong Li, Shilei Wang, Ning Yu, Jia Liu
Journal: Biochemical and biophysical research communications (2015): 537--542

Fungus induces the release of IL-8 in human corneal epithelial cells, via Dectin-1-mediated protein kinase C pathways.
Authors: Xu-Dong Peng, Gui-Qiu Zhao, Jing Lin, Nan Jiang, Qiang Xu, Cheng-Cheng Zhu, Jain-Qiu Qu, Lin Cong, Hui Li
Journal: International journal of ophthalmology (2014): 441--447

Propofol and remifentanil at moderate and high concentrations affect proliferation and differentiation of neural stem/progenitor cells
Authors: Qing Li, Jiang Lu, Xianyu Wang
Journal: Neural regeneration research (2014): 2002

Role of mitochondrial calcium uniporter in regulating mitochondrial fission in the cerebral cortexes of living rats
Authors: Nan Liang, Peng Wang, Shilei Wang, Shuhong Li, Yu Li, Jinying Wang, Min Wang
Journal: Journal of Neural Transmission (2014): 593--600

Increased expression of cell adhesion molecule 1 by mast cells as a cause of enhanced nerve--mast cell interaction in a hapten-induced mouse model of atopic dermatitis
Authors: M Hagiyama, T Inoue, T Furuno, T Iino, S Itami, M Nakanishi, H Asada, Y Hosokawa, A Ito
Journal: British Journal of Dermatology (2013): 771--778

 

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