TMA-DPH
(Synonyms: TMA-DPH(疏水膜荧光探针),N,N,N-Trimethyl-4-(6-phenyl-1,3,5-hexatrien-1-yl)phenylammonium (p-toluenesulfonate)) 目录号 : GC45058
A fluorescent probe
Cas No.:115534-33-3
Sample solution is provided at 25 µL, 10mM.
;)
,-1-7$$$37316672.jpg');)
;)
;)
$$$36806353.jpg');)
;33(7)%EF%BC%9A546-561$$$37156877.jpg');)
;)
;)
;)
%201124-1138.$$$35908550.jpg');)
%20766-780.$$$37100057.jpg');)
%20418-432.$$$36893736.jpg');)
%EF%BC%9A-240-255.$$$35108512.jpg');)
533-545$$$36543525.jpg');)
%201-13.$$$36600053.jpg');)
;)
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.细胞膜染色液制备
(1)配置DMSO或EtOH 储存液:储存液用DMSO或EtOH配置,浓度1~5mM。
注意: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)稀释储存液，配制浓度为0.5~5μM的工作液。
注意: 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.细胞染色
（1）．悬浮细胞: 经4°C、1000-1500rpm离心收集细胞，加入 PBS 洗涤两次，每次5分钟。使用终浓度为2μM的TMA-DPH工作液重悬细胞，室温下避光进行短期孵育(10S)，而后立即将未洗过的细胞铺板后转移到显微镜下观察[1]。
贴壁细胞;弃去培养基，加入胰蛋白酶消化细胞。经4°C、1000-1500rpm离心弃去上清后，加入 PBS 洗涤两次，每次5分钟。
（2）.贴壁细胞：弃去培养基，使用PBS 洗涤两次，每次5分钟。
1).细胞质膜染色：细胞在室温下与终浓度为2μM的TMA-DPH进行短期孵育(10S)。然后将未洗过的玻片转移到显微镜下观察。
2).细胞内化膜标记染色：细胞移至载玻片烧瓶中，根据实验需求，在37°条件下用终浓度为2μM的TMA-DPH孵育所需的时间。孵育完成后在PBS中轻轻摇动载玻片几秒钟进行洗涤，用以消除外周标记[1]。
4. 显微镜检测：TMA-DPH的激发/发射光分别为355/430nm。
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]. Illinger, D., and Kurhy, J.G. The kinetic aspects of intracellular fluorescence labeling with TMA-DPH support the maturation model for endocytosis in L929 cells. Journal of Cell Biology 125(4), 783-794 (1994).
TMA-DPH is a fluorescent probe used for measuring membrane fluidity in artificial and living membrane systems.[1],[2],[3],[4] The cylindrical shape of TMA-DPH confers high sensitivity to reorientation resulting from changes in surrounding lipids. TMA-DPH has excitation and emission maxima at 355 and 430 nm, respectively.
Reference:
[1]. Subramanian, V., Knight, J.S., Parelkar, S., et al. Design, synthesis, and biological evaluation of tetrazole analogs of cl-amidine as protein arginine deiminase inhibitors. Journal of Medicinal Chemistry 58(3), 1337-1344 (2015).
[2]. Illinger, D., and Kurhy, J.G. The kinetic aspects of intracellular fluorescence labeling with TMA-DPH support the maturation model for endocytosis in L929 cells. Journal of Cell Biology 125(4), 783-794 (1994).
[3]. Illinger, D., Duportail, G., Mely, Y., et al. A comparison of the fluorescence properties of TMA-DPH as a probe for plasma membrane and for endocytic membrane. Biochimica et Biophysica Acta 1239(1), 58-66 (1995).
[4]. Chazotte, B. Labeling the plasma membrane with TMA-DPH. Cold Spring Harb.Protoc. 2011(5), (2011).).
| Cas No. | 115534-33-3 | SDF | |
| 别名 | TMA-DPH(疏水膜荧光探针),N,N,N-Trimethyl-4-(6-phenyl-1,3,5-hexatrien-1-yl)phenylammonium (p-toluenesulfonate) | ||
| 化学名 | N,N,N-trimethyl-4-(6-phenyl-1,3,5-hexatrien-1-yl)-benzenaminium, 4-methylbenzenesulfonate | ||
| Canonical SMILES | C[N+](C)(C)C1=CC=C(/C=C/C=C/C=C/C2=CC=CC=C2)C=C1.CC3=CC=C(S([O-])(=O)=O)C=C3 | ||
| 分子式 | C21H24N•C7H7O3S | 分子量 | 461.6 |
| 溶解度 | 1mg/mL in methanol, or in DMF , 10mg/mL in DMSO | 储存条件 | Store at -20°C, protect from light,unstable in solution, ready to use. |
| 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。 |
||
| 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 | |||
 |
1 mg | 5 mg | 10 mg |
| 1 mM | 2.1664 mL | 10.8319 mL | 21.6638 mL |
| 5 mM | 0.4333 mL | 2.1664 mL | 4.3328 mL |
| 10 mM | 0.2166 mL | 1.0832 mL | 2.1664 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。
Diphenylhexatriene membrane probes DPH and TMA-DPH: A comparative molecular dynamics simulation study
Biochim Biophys Acta 2016 Nov;1858(11):2647-2661.PMID:27475296DOI:10.1016/j.bbamem.2016.07.013.
Fluorescence spectroscopy and microscopy have been utilized as tools in membrane biophysics for decades now. Because phospholipids are non-fluorescent, the use of extrinsic membrane probes in this context is commonplace. Among the latter, 1,6-diphenylhexatriene (DPH) and its trimethylammonium derivative (TMA-DPH) have been extensively used. It is widely believed that, owing to its additional charged group, TMA-DPH is anchored at the lipid/water interface and reports on a bilayer region that is distinct from that of the hydrophobic DPH. In this study, we employ atomistic MD simulations to characterize the behavior of DPH and TMA-DPH in 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and POPC/cholesterol (4:1) bilayers. We show that although the dynamics of TMA-DPH in these membranes is noticeably more hindered than that of DPH, the location of the average fluorophore of TMA-DPH is only ~3-4Å more shallow than that of DPH. The hindrance observed in the translational and rotational motions of TMA-DPH compared to DPH is mainly not due to significant differences in depth, but to the favorable electrostatic interactions of the former with electronegative lipid atoms instead. By revealing detailed insights on the behavior of these two probes, our results are useful both in the interpretation of past work and in the planning of future experiments using them as membrane reporters.
Fluorescent probes DPH, TMA-DPH and C17-HC induce erythrocyte exovesiculation
J Membr Biol 2002 Nov 1;190(1):75-82.PMID:12422273DOI:10.1007/s00232-002-1025-5.
An experimental approach has been developed to study human erythrocyte vesiculation, using the fluorescent probes diphenylhexatriene (DPH), trimethylamino-diphenylhexatriene (TMA-DPH) and heptadecyl-hydroxycoumarin (C17-HC). Acetylcholinesterase (AChE) enzyme activity measurements confirmed the presence of exovesicles released from erythrocyte membranes labeled with DPH, TMA-DPH or C17-HC. The fluorescence intensity and anisotropy values obtained showed that the amphiphilic probes TMA-DPH and C17-HC are preferentially incorporated in the exovesicles (when compared with DPH). There is a significant decrease of the cholesterol content of the exovesicle suspensions with time, independently of the fluorescence probe used, reaching undetectable cholesterol levels for the samples incubated for 48 hr. The ratios between the concentration of cholesterol released in the exovesicles after 1 hr incubation with DPH, TMA-DPH or C17-HC and the probe concentration used in the incubation were 84.7, 3.82 and 0.074, respectively. The size of the released vesicles was evaluated by dynamic light scattering spectroscopy. Some hypotheses are proposed that could explain the resemblance and differences between the results obtained for erythrocytes labeled with each probe, considering the present knowledge of membrane vesiculation mechanisms, lipid microdomains (rafts), erythrocyte membrane phospholipid asymmetry and AChE inhibition by TMA-DPH and C17-HC. This work demonstrates that the fluorescent probes DPH, TMA-DPH and C17-HC induce rapid erythrocyte exovesiculation; their use can lead to new methodologies for the study of this still poorly understood mechanism.
Properties of a phosphatidylcholine derivative of diphenyl hexatriene (DPH-PC) in lymphocyte membranes. A comparison with DPH and the cationic derivative TMA-DPH using static and dynamic fluorescence
Membr Biochem 1993 Jan-Mar;10(1):17-27.PMID:8510559DOI:10.3109/09687689309150249.
Using static and dynamic fluorescence we studied the fluorescence properties of a phosphatidylcholine analog of 1,6-diphenyl-1,3,5-hexatriene (DPH-PC) incorporated in lymphocyte plasma membranes with respect to DPH and its cationic derivative 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH), in order to study if phospholipid derivatives of DPH may be used to investigate structural and physicochemical properties of specific membrane lipid domains. DPH-PC and TMA-DPH showed similar fluorescence polarization values that were significantly higher with respect to DPH, suggesting a localization of the fluorescent portion of DPH-PC in a more ordered region of the membrane which was probably due to the elecrostatic interactions between phospholipid head-groups. The localization of the fluorescent moiety of DPH-PC near the membrane surface was also supported by the study of the fluorescence decay of the three probes using frequency-domain fluorometry. The main lifetime component of DPH-PC was rather similar to that of TMA-DPH (6.74 versus 6.24, ns) but considerably lower with respect to DPH (10.52 ns), in agreement with data obtained from exponential analysis. In lymphocyte membranes obtained from concanavalin A treated cells, a significant decrease of fluorescence polarization has been shown with DPH and its phosphatidylcholine derivative, but not with TMA-DPH. In liposomes obtained from total lipids extracted from lymphocyte membranes, a decrease of fluorescence polarization has been observed only with DPH. Our results suggest that DPH-PC localizes the fluorescent portion of its molecule in membrane microenvironments of different properties with respect to those probed by DPH and TMA-DPH. The use of DPH-phospholipid derivatives and other DPH-probes may represent an useful tool to study plasma membrane heterogeneity in biological membranes.
Labeling the plasma membrane with TMA-DPH
Cold Spring Harb Protoc 2011 May 1;2011(5):pdb.prot5622.PMID:21536758DOI:10.1101/pdb.prot5622.
INTRODUCTION TMA-DPH (trimethylamine-diphenylhexatriene) is a fluorescent membrane probe that has classically been used to label the outer leaflet of a membrane bilayer, to label the outer leaflet of the plasma membrane in cells, and to report on membrane dynamics using the techniques of fluorescence polarization and/or fluorescence lifetime. This probe has also been used to follow exocytosis and endocytosis of labeled plasma membranes. The interaction of the aqueous environment with mitochondrial inner membrane dynamics has also been studied following the fluorescence polarization and the lifetime of TMA-DPH. This protocol describes the use of TMA-DPH to label the plasma membrane.
Plasma membrane fluidity in isolated rat hepatocytes: comparative study using DPH and TMA-DPH as fluorescent probes
J Gastroenterol Hepatol 1989 May-Jun;4(3):221-7.PMID:2491149DOI:10.1111/j.1440-1746.1989.tb00829.x.
The study of membrane fluidity is a rapidly expanding field of research and its interest in hepatology has been stressed recently. The present study is the first report concerned with the determination of membrane fluidity of isolated rat hepatocytes. The data have been compared with those obtained in plasma membrane fractions and subfractions (basolateral or canalicular) derived from homogenates. The fluorescent probes used to measure the fluidity were diphenylhexatriene (DPH) a 'classical' probe, and its derivative trimethylammoniodiphenylhexatriene (TMA-DPH) at 25 degrees C and at 37 degrees C. The values obtained with DPH were lower than those with TMA-DPH, probably due to the localization of the probes in different regions of the phospholipid bilayer. In addition, DPH revealed significant differences in the fluorescence polarization values obtained in isolated hepatocytes compared with membrane fractions, which was in contrast to TMA-DPH, where the respective values were of the same order of magnitude. This behaviour is probably due to the mobility of DPH in the membrane core and its rapid internalization into the cell, whereas TMA-DPH remains anchored for a long time on the cell surface. These findings suggest that TMA-DPH is a better probe than DPH for measuring the fluorescence polarization of whole isolated hepatocytes and that the use of different probes might be of help in exploring different zones of the membrane bilayer.