FM1-43
目录号 : GC67896FM1-43 是一种用于研究胞吞作用和胞吐作用的苯乙烯染料，可作为毛细胞机械传感器通道的渗透阻滞剂。FM1-43 还降低氨基糖苷类抗生素硫酸新霉素的耳毒性作用。
Cas No.:149838-22-2
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >95.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
本方案仅提供一个指导，应根据您的具体需要进行修改。
1. FM1-43细胞膜染色液制备
(1)配置DMSO储存液:储存液用DMSO配置,浓度1~5mM。
注:未使用的储存液分装保存在-20℃或-80℃避光保存，避免反复冻融。
(2)工作液制备: 使用合适的缓冲液(如:无血清培养基，HBSS或PBS)以获得5-20μM的FM工作溶液[2]。
注: 工作液终浓度需要根据不同细胞系和实验体系来优化，建议从推荐浓度开始，以10倍范围为区间进行最优浓度的摸索。
2.悬浮细胞染色
(1)悬浮细胞经4°C、1000-1500rpm离心3-5分钟，弃去上清液。用PBS清洗两次，每次5分钟。
(2)加入1mL的FM1-43工作液（推荐浓度10μM），室温孵育5-30分钟，不同的细胞最佳培养时间不同。
(3)细胞试管在1000-1500rpm离心5分钟。
(4) 孵育结束后，经1000-1500rpm离心5分钟，去除上清液，加入PBS清洗2-3次，每次5分钟。
(5)用无血清细胞培养基或PBS重悬细胞。通过荧光显微镜或流式细胞术观察。
3.粘壁细胞染色
(1)在无菌盖玻片上培养贴壁细胞。
(2从培养基中移走盖玻片，吸出过量的培养基，将盖玻片放在潮湿的环境中。
(3)从盖玻片的一角加入100uL的FM1-43工作液，轻轻晃动使染料均匀覆盖所有细胞。
(4)室温条件下孵育5-30分钟，不同的细胞最佳培养时间不同。
(5)吸弃工作液，用培养液洗盖玻片2~3次。
4.显微镜检测： FM1-43激发/发射光分别为480/598nm[3]。
注意事项：1）荧光染料均存在淬灭问题，请尽量注意避光，以减缓荧光淬灭。2）为了您的安全和健康，请穿实验服并戴一次性手套操作。
References:
[1]. J J Renger,C Egles, G Liu. A developmental switch in neurotransmitter flux enhances synaptic efficacy by affecting AMPA receptor activation.2001 Feb;29(2):469-84. doi: 10.1016/s0896-6273(01)00219-7.
[2]. Adriana Della Pietra, Nikita Mikhailov, Rashid Giniatullin. FM1-43 Dye Memorizes Piezo1 Activation in the Trigeminal Nociceptive System Implicated in Migraine Pain. 2023 Jan 14;24(2):1688. doi: 10.3390/ijms24021688.
FM1-43 is a water-soluble styrene dye with strong lipophilicity. FM 1-43 membrane probe is a reagent that can specifically bind to cell membranes and inner membrane organelles to generate fluorescence. It is widely used as endocytosis and excision membrane structure markers, and can be used to identify actively firing neurons and research Mechanisms of activity-dependent vesicle cycling[1]. In actively releasing neurotransmitters, the dye is internalized in recycled synaptic vesicles and the nerve terminals become brightly stained. Non-specific staining of cell surface membranes can be simply washed off before observation[2]. FM1-43 is nontoxic to cells, barely fluoresces in aqueous media, but fluoresces strongly after insertion into the outer leaflet of the cell membrane. FM1-43 also reduces the ototoxic effects of the aminoglycoside antibiotic neomycin sulfate[3].
FM1-43是一种亲脂性很强的水溶性苯乙烯染料。FM 1-43膜探针是一种能特异性地与细胞膜和内膜细胞器结合产生荧光的试剂，被广泛用于内吞和外切膜结构标志物，可用于识别活跃放电的神经元和研究活动依赖性囊泡循环的机制[1]。在主动释放神经递质的神经元中，染料在回收的突触囊泡内化，神经末梢变得明亮染色。细胞表面膜的非特异性染色可以在观察前简单地洗掉[2]。FM1-43对细胞无毒，在水性介质中几乎不发荧光，但在插入细胞膜的外小叶后发出强烈荧光。FM1-43 还降低氨基糖苷类抗生素硫酸新霉素的耳毒性作用[3]。
References:
[1]. Audrey C Brumback, et al. Using FM1-43 to study neuropeptide granule dynamics and exocytosis.2004 Aug;33(4):287-94. doi: 10.1016/j.ymeth.2004.01.002.
[2]. J J Renger,C Egles, G Liu. A developmental switch in neurotransmitter flux enhances synaptic efficacy by affecting AMPA receptor activation.2001 Feb;29(2):469-84. doi: 10.1016/s0896-6273(01)00219-7.
[3]. J E Gale, et al. FM1-43 dye behaves as a permeant blocker of the hair-cell mechanotransducer channel. J Neurosci. 2001 Sep 15;21(18):7013-25.
| Cas No. | 149838-22-2 | SDF | Download SDF |
| 分子式 | C30H49Br2N3 | 分子量 | 611.54 |
| 溶解度 | DMSO : 50 mg/mL (81.76 mM; ultrasonic and warming and heat to 60°C) | 储存条件 | Store at -20°C |
| General tips | 请根据产品在不同溶剂中的溶解度选择合适的溶剂配制储备液；一旦配成溶液，请分装保存，避免反复冻融造成的产品失效。 储备液的保存方式和期限：-80°C 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。 为了提高溶解度，请将管子加热至37℃，然后在超声波浴中震荡一段时间。 |
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| Shipping Condition | 评估样品解决方案：配备蓝冰进行发货。所有其他可用尺寸：配备RT，或根据请求配备蓝冰。 | ||
| 制备储备液 | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 1.6352 mL | 8.1761 mL | 16.3522 mL |
| 5 mM | 0.327 mL | 1.6352 mL | 3.2704 mL |
| 10 mM | 0.1635 mL | 0.8176 mL | 1.6352 mL |
| 第一步：请输入基本实验信息（考虑到实验过程中的损耗，建议多配一只动物的药量） | ||||||||||
| 给药剂量 | mg/kg |  |
动物平均体重 | g | |
每只动物给药体积 | ul | |
动物数量 | 只 |
| 第二步：请输入动物体内配方组成（配方适用于不溶于水的药物；不同批次药物配方比例不同，请联系GLPBIO为您提供正确的澄清溶液配方） | ||||||||||
| % DMSO % % Tween 80 % saline | ||||||||||
| 计算重置 | ||||||||||
计算结果:
工作液浓度： mg/ml；
DMSO母液配制方法： mg 药物溶于 μL DMSO溶液（母液浓度 mg/mL，
体内配方配制方法：取 μL DMSO母液，加入 μL PEG300，混匀澄清后加入μL Tween 80，混匀澄清后加入 μL saline，混匀澄清。
1. 首先保证母液是澄清的；
2.
一定要按照顺序依次将溶剂加入，进行下一步操作之前必须保证上一步操作得到的是澄清的溶液，可采用涡旋、超声或水浴加热等物理方法助溶。
3. 以上所有助溶剂都可在 GlpBio 网站选购。
Using FM1-43 to study neuropeptide granule dynamics and exocytosis
Methods 2004 Aug;33(4):287-94.PMID:15183177DOI:10.1016/j.ymeth.2004.01.002.
In the study of neuropeptide secretion and membrane trafficking, the fluorescent dye FM1-43 provides the ability to label selectively those structures that are undergoing exocytosis and endocytosis in living cells in real time. This review describes the unique properties of the FM dyes that make them ideal for studying neuropeptide granule dynamics and discusses various techniques that take advantage of FM dyes.
FM1-43 Dye Memorizes Piezo1 Activation in the Trigeminal Nociceptive System Implicated in Migraine Pain
Int J Mol Sci 2023 Jan 14;24(2):1688.PMID:36675204DOI:10.3390/ijms24021688.
It has been proposed that mechanosensitive Piezo1 channels trigger migraine pain in trigeminal nociceptive neurons, but the mechanosensitivity of satellite glial cells (SGCs) supporting neuronal sensitization has not been tested before. Moreover, tools to monitor previous Piezo1 activation are not available. Therefore, by using live calcium imaging with Fluo-4 AM and labeling with FM1-43 dye, we explored a new strategy to identify Piezo channels' activity in mouse trigeminal neurons, SGCs, and isolated meninges. The specific Piezo1 agonist Yoda1 induced calcium transients in both neurons and SGCs, suggesting the functional expression of Piezo1 channels in both types of cells. In Piezo1-transfected HEK cells, FM1-43 produced only a transient fluorescent response, whereas co-application with Yoda1 provided higher transient signals and a remarkable long-lasting FM1-43 'tail response'. A similar Piezo1-related FM1-43 trapping was observed in neurons and SGCs. The non-specific Piezo channel blocker, Gadolinium, inhibited the transient peak, confirming the involvement of Piezo1 receptors. Finally, FM1-43 labeling demonstrated previous activity in meningeal tissues 3.5 h after Yoda1 washout. Our data indicated that trigeminal neurons and SGCs express functional Piezo channels, and their activation provides sustained labeling with FM1-43. This long-lasting labelling can be used to monitor the ongoing and previous activation of Piezo1 channels in the trigeminal nociceptive system, which is implicated in migraine pain.
Monitoring secretory membrane with FM1-43 fluorescence
Annu Rev Neurosci 1999;22:1-10.PMID:10202529DOI:10.1146/annurev.neuro.22.1.1.
FM1-43 and similar styryl dyes have proven useful as probes for membrane trafficking because they reversibly stain membranes, are impermeable to membranes, and are more fluorescent when bound to membranes than when in solution. Because these dyes stain membranes in an activity-dependent manner, they are ideal for studies of neurotransmitter release mechanisms such as synaptic vesicle recycling, exocytosis, and endocytosis. FM dyes have been used in conjunction with other techniques such as fluorescent calcium indicator dyes and electrophysiological techniques to elucidate mechanisms of presynaptic calcium homeostasis and modulation of neurotransmitter release. Presynaptic membranes have been marked by FM dyes in studies of synaptogenesis and reinnervation. As a probe for endocytosed membranes, these dyes have been used to examine vacuole formation in yeast. These versatile membrane dyes are useful in a variety of applications.
FM1-43 Photoconversion and Electron Microscopy Analysis at the Drosophila Neuromuscular Junction
Bio Protoc 2017 Sep 5;7(17):e2523.PMID:29094061DOI:10.21769/BioProtoc.2523.
We developed a protocol for photoconversion of endocytic marker FM1-43 followed by electron microscopy analysis of synaptic boutons at the Drosophila neuromuscular junction. This protocol allows detection of stained synaptic vesicle even when release rates are very low, such as during the spontaneous release mode. The preparations are loaded with the FM1-43 dye, pre-fixed, treated and illuminated to photoconvert the dye, and then processed for conventional electron microscopy. This procedure enables clear identification of stained synaptic vesicles at electron micrographs.
FM1-43 reports plasma membrane phospholipid scrambling in T-lymphocytes
Biochem J 2000 Jul 1;349(Pt 1):255-60.PMID:10861236DOI:10.1042/0264-6021:3490255.
We have found using imaging techniques that stimulating Jurkat human leukaemic T-cells with ionomycin in the presence of FM1-43, a dye used to monitor exocytosis and endocytosis, causes large (6--10-fold) increases in FM1-43 fluorescence. These responses are too large to be caused by exocytosis. Instead, three lines of evidence suggest that FM1-43 is responding to phospholipid scrambling. First, ionomycin also stimulates increases in the fluorescence of annexin V, a phosphatidylserine-specific probe, while thapsigargin does not stimulate fluorescence increases of either probe. Secondly, cells that exhibit FM1-43 fluorescence increases after ionomycin stimulation stain with annexin V once FM1-43 is washed out. Thirdly, ionomycin stimulates uptake of 7-nitrobenz-2-oxa-1,3-diazole-labelled phosphatidylcholine, a specific assay for scramblase activity, whereas thapsigargin does not. We find that FM1-43 reports phospholipid scrambling with 'better' kinetics than annexin V, and does require extracellular Ca(2+) to report phospholipid scrambling. We suggest that FM1-43 may be a useful probe to study the dynamics of phospholipid scrambling. The results are the first demonstration that FM1-43 can respond significantly to a biological process other than vesicular trafficking.