Fluo-4,AM,细胞可透过性
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Fluo-4,AM,细胞可透过性
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Fluo-4,AM,细胞可透过性

标记钙指示剂是结合 Ca2+ 离子后荧光强度增加的分子。Fluo-3 应用在流式细胞分析上,如涉及笼状螯合剂光活化、第二信使和神经递质的实验以及细胞水平的药理学筛选,用于 Ca2+ 的空间动力学成像。Fluo-4了解更多信息
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货号数量
F1420110 x 50 μg
货号 F14201
价格(CNY)
3,781.00
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Ends: 31-Dec-2025
5,089.00
共减 1,308.00 (26%)
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数量:
10 x 50 μg
价格(CNY)
3,781.00
Online Exclusive
Ends: 31-Dec-2025
5,089.00
共减 1,308.00 (26%)
Each
添加至购物车
标记钙指示剂是结合 Ca2+ 离子后荧光强度增加的分子。Fluo-3 应用在流式细胞分析上,如涉及笼状螯合剂光活化、第二信使和神经递质的实验以及细胞水平的药理学筛选,用于 Ca2+ 的空间动力学成像。Fluo-4 是 fluo-3 的类似物,其中两个氯取代基被氟取代,导致在 488 nm 波长处的荧光激发增强,因此荧光信号水平更高。将溶解后的指示剂直接加入含有培养细胞的培养皿中,即可向细胞上样 AM 酯形式的这类钙离子指示剂。这些指示剂适用于荧光和共聚焦显微镜、流式细胞分析和微孔板筛选应用。

了解更多有关离子指示剂(包括钙、钾、pH 值和膜电位指示剂)的信息›

钙指示剂(AM 酯)规格:
•标签(Ca2+– 结合形式的激发/发射波长):Fluo-4 (494/506 nm)
• 结合 Ca2+ 后荧光强度增加:>100 倍
缓冲液中 Ca2+ 的 Kd:∼335 nM
• 结合 Ca2+ 后,荧光增加,波长稍有变化

使用 TPEN 控制重金属阳离子
此外,基于 BAPTA 的指示剂可结合多种重金属阳离子(例如 Mn2+、Zn2+、Pb2+),亲和力远高于 Ca2+。可以使用重金属选择性螯合剂 TPEN 来控制由这些离子引起的钙测量值扰动。

荧光钙指示剂的更多选择
我们提供大量的 Molecular Probes™ 钙指示剂供各种实验场景选择使用。更多信息请参阅《Molecular Probes™ 手册》中的可见光激发的荧光 Ca2+ 指示剂—第 19.3 节

对于紫外激发的 Ca2+ 指示剂、基于蛋白的 Ca2+ 指示剂、Ca2+ 指示剂的偶联物以及其他金属离子(即 Mg2+、Zn2+)的荧光指示剂,请参阅《Molecular Probes™ 手册》中的 Ca2+、Mg2+、Zn2+ 以及其他金属离子指示剂—第 19 章

我们推荐使用高质量的无水 DMSO 进行溶解,以达到 1 到 5 mM 储备液浓度。

仅供科研使用。不可用于诊断程序。
规格
检测方法荧光
染料类型基于荧光染料
激发/发射494/506 nm
数量10 x 50 μg
运输条件室温
适用于(应用)细胞活力、增殖与细胞成像, Cell Proliferation, Cellular Imaging
适用于(设备)共聚焦显微镜, 荧光显微镜, 高内涵分析仪, HTS 读数仪, 微孔板读数仪, 荧光成像仪
产品类型染料
Unit SizeEach
内容与储存
在冷冻冰箱(-5°C 至 -30°C)中避光储存。

常见问题解答 (FAQ)

为什么我看不到活细胞荧光指示剂信号有明显变化?

不管何种活细胞指示剂染料(如钙指示剂,pH指示剂,金属离子指示剂),请务必确保上样过程中无血清,否则血清会过早地切割带有AM酯基的染料并非特异性结合染料。在检测样品之前,请使用阳性对照优化染料浓度和染色时间来得到最佳的信号与背景比值。做阳性对照时缓冲液一定要包含已知浓度的自由离子和用于打开离子通道的离子载体(例如Fluo-4 AM一类的钙指示剂,它包括缓冲液中加入钙与卡西霉素,又比如pH指示剂,不同pH值的缓冲液与尼日利亚霉素相结合)。类似于CellROX Green 或H2DCFDA需要一个细胞活性氧(ROS)刺激作为阳性对照,例如甲萘醌。最后,确保成像系统具有灵敏的探测器。例如相对于显微镜或流式细胞仪,读板仪检测信号和背景差异的能力就比较低。

Can I fix my cells after loading with Fluo-4 AM dye for the detection of calcium?

No. Since Fluo-4 AM isn't covalently bound to any cellular components and fixation compromises the membrane, the dye would not be retained by the cell.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I am doing calcium flux imaging with your Fura-2 calibration kit, but am seeing a large variability in ratio in different places around the slide. I am correcting for uniform illumination, using the product as directed, and sealing the coverslip with nail polish.

The nail polish may be the problem. The Kd value (calcium sensitivity) changes depending upon the dye's environment. Nail polish has solvents that can leech under the coverslip and cause variability. We recommend either going without a sealing or sealing with melted paraffin painted on the coverslip edges with a cotton-tipped applicator (paraffin is hydrophobic and has no solvents).

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I need to label cells with Fluo-4, AM, for a calcium flux assay. How long after labeling will the dye be retained?

After loading dye into the cells, intracellular esterases remove the 'AM' moiety from the dye. When the 'AM' group is removed, the dye is able to bind calcium and fluoresce. Since the dye is not covalently bound to any cellular components, it may be actively effluxed from the cell. The rate of efflux is dependent upon the inherent properties of the cell, culture conditions and other factors. The dye may be retained for hours, days or even weeks or lost in a matter of minutes. The use of Probenecid (Cat. No. P36400) limits loss by active efflux.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Why don't I see a significant change in signal for my live-cell fluorescent indicator dye?

Regardless of the type of live-cell indicator dye (e.g., calcium indicators, pH indicator, metal ion indicators), make sure there is no serum during the loading step, which can prematurely cleave dyes with AM esters and bind dyes non-specifically. Always optimize the dye concentration and staining time with a positive control before you run your test samples, to give the best signal-to-background. Always run a positive control with a buffer containing free ions of known concentration and an ionophore to open pores to those ions (for instance, for calcium indicators like Fluo-4 AM, this would include a buffer with added calcium combined with calcimycin, or for pH indicators, buffers of different pHs combined with nigericin). Reactive oxygen indicators, such as CellROX Green or H2DCFDA would require a cellular reactive oxygen species (ROS) stimulant as a positive control, such as menadione. Finally, make sure your imaging system has a sensitive detector. Plate readers, for instance, have much lower detector efficiency over background, compared to microscopy or flow cytometry.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.