Acrylodan
(Synonyms: 6-丙烯酰基-2-二甲氨基萘) 目录号 : GC61505
Acrylodan可与硫醇反应，对Cys34周围的结合口袋内的局部环境双极性和动力学敏感。
Cas No.:86636-92-2
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >98.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Acrylodan, reacted with thiols, is sensitive to the local environmental dipolarity and dynamics within the binding pocket surrounding Cys34[1][2].
Guidelines (Following is our recommended protocol. This protocol only provides a guideline, and should be modified according to your specific needs).Purification and Labeling of Act1: 1.20-fold molar excess of acrylodan (Anaspec) was added to the polymerized actin and incubated on ice overnight. 2.The labeled actin is centrifuged (2 h, 4 °C, 100,000 × g) then the pellet was resuspended in 10 mm Tris, pH 7.5, 0.5 mm β-mercaptoethanol, 0.2 mm CaCl2, 50 μm ATP and dialyzed against the same buffer for 70 h at 4 °C. 3.The supernatant was collected and the actin concentration and degree of labeling were calculated using the extinction coefficient 26,600 M-1 cm-1 for actin at 290 nm, 18,500 M-1 cm-1 or acrylodan at 385 nm, and 71,000 M-1 cm-1 for Alexa Fluor 488 at 494 nm. 4.Acrylodan-labeled Act1 (AcrylAct1NAT) is dramatically different than that of EDTA-unfolded AcrylAct1 (AcrylAct1I3), which has a very similar emission spectrum to CCT-bound AcrylAct1.
[1]. Sarah F Stuart, et al. A two-step mechanism for the folding of actin by the yeast cytosolic chaperonin. J Biol Chem. 2011 Jan 7;286(1):178-84. [2]. K Flora, et al. Unfolding of acrylodan-labeled human serum albumin probed by steady-state and time-resolved fluorescence methods. Biophys J. 1998 Aug;75(2):1084-96.
| Cas No. | 86636-92-2 | SDF | |
| 别名 | 6-丙烯酰基-2-二甲氨基萘 | ||
| Canonical SMILES | C=CC(C1=CC=C2C=C(C=CC2=C1)N(C)C)=O | ||
| 分子式 | C15H15NO | 分子量 | 225.29 |
| 溶解度 | 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 | 4.4387 mL | 22.1936 mL | 44.3872 mL |
| 5 mM | 0.8877 mL | 4.4387 mL | 8.8774 mL |
| 10 mM | 0.4439 mL | 2.2194 mL | 4.4387 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。
Acrylodan-labeled intestinal fatty acid-binding protein to measure concentrations of unbound fatty acids
Methods Mol Biol 2007;400:27-43.PMID:17951725DOI:10.1007/978-1-59745-519-0_3.
The concentration of long-chain (14-18 carbons) fatty acids (FA) free in solution (unbound) is difficult to measure directly because of the low aqueous solubility of these common dietary FA. One indirect and convenient way to measure the concentration of unbound FA is a method using the fluorescent-(Acrylodan) labeled intestinal FA-binding protein (ADIFAB). Under appropriate conditions, ADIFAB fluorescence measures unbound FA, regardless of any third phase such as albumin, FA-binding proteins, or membranes. With knowledge of the total amount of FA in the system and the assumption that the amount of FA bound to ADIFAB is negligible, equilibrium constants or partition coefficients for FA in equilibrium with the third phase can be calculated. Herein, the use of ADIFAB is described to measure unbound FA concentration using oleic acid as a typical long-chain FA. Attempts were not made to calibrate the accuracy of ADIFAB for FA concentration, but to investigate its reliability and reproducibility under differing buffer conditions. It is shown that ADIFAB fluorescence is sensitive to biologically prevalent ions and that calibration curves must be constructed for conditions that do not closely match those previously published. The results with in vitro systems suggest that there will be caveats with the application of ADIFAB to measure FA concentrations in vivo, where the precise environment of the probe is not known or cannot be tightly controlled.
Acrylodan can label amino as well as sulfhydryl groups: results with low-density lipoprotein, lipoprotein[a], and lipid-free proteins
Biochemistry 1993 Sep 7;32(35):9215-20.PMID:8369288DOI:10.1021/bi00086a029.
Human plasma lipoprotein[a] and autologous low-density lipoprotein were reacted with the fluorescent probe 6-acryloyl-2-(dimethylamino)naphthalene (Acrylodan) previously reported to be specific for sulfhydryl groups. Reaction kinetics were biphasic in both cases. The reaction of bovine serum albumin with Acrylodan was also biphasic. Monophasic kinetics were observed when protein free sulfhydryl groups were blocked by carboxamidomethylation prior to Acrylodan reaction. A significant increase in total fluorescence was observed in the reaction of Acrylodan with proteins containing no free sulfhydryl groups and with polylysine. The rates of these reactions were highly sensitive to pH. Fluorescence changes due to dissolution of probe into hydrophobic protein or lipid domains were minimal as was reaction of probe with phospholipid head groups. When isolated from acrylodan-labeled Lp[a], apo[a], which contains no free sulfhydryl groups, contained covalently bound Acrylodan. These results suggest that Acrylodan can modify the lysine residues of lipid-free proteins and may modify not only the free sulfhydryl groups of low-density lipoprotein and lipoprotein[a] but also reactive amino groups. We conclude that under these conditions, the use of this probe to quantify free sulfhydryl groups in these lipoproteins is infeasible.
Fluorescent responses of acrylodan-labeled plasma gelsolin
Arch Biochem Biophys 1993 Apr;302(1):31-6.PMID:8385903DOI:10.1006/abbi.1993.1176.
Under nondenaturing conditions, 1 mol of horse plasma gelsolin reacts with 1.9 +/- 0.5 mol (mean +/- SD, n = 6) of the sulfhydryl-specific fluorescent reagent 6-acryloyl-2-dimethylaminonaphthalene (Acrylodan). The degree of labeling in 6 M guanidine-HCl increases to about 3 mol of Acrylodan per mole of gelsolin. Viscosity studies show that the modified gelsolin retains its ability to sever F-actin filaments. Circular dichroism spectra in the peptide bond absorption region are indistinguishable for labeled and unmodified gelsolin at room temperature. The thermal stability of gelsolin, as monitored by circular dichroism, is unaffected by reaction with Acrylodan. While circular dichroism spectra of acrylodan-labeled gelsolin recorded at room temperature are not influenced significantly by Ca2+, fluorescence studies reveal a number of Ca(2+)-dependent changes in the protein. Ca2+ causes a decrease and red-shift in fluorescence emission, an increase in sensitivity to quenching by I-, and a decrease in polarization of the fluorescence of acrylodan-labeled gelsolin. Together, these changes in fluorescence properties indicate there to be an increased exposure of the label to the solvent when gelsolin binds Ca2+. Fluorescence polarization experiments in which acrylodan-labeled gelsolin is titrated with actin emphasize that Ca2+ is required for these two proteins to interact.
A fluorescent derivative of the oligomycin-sensitivity conferring protein (acrylodan-OSCP). Evidence for polarity changes in the environment of CYS118 of OSCP upon binding to mitochondrial F1
Biochem Biophys Res Commun 1987 Jan 15;142(1):31-7.PMID:2880585DOI:10.1016/0006-291x(87)90447-5.
The fluorescent probe, 6-acryloyl-2-dimethylaminonaphtalene (Acrylodan) was reacted with the oligomycin-sensitivity conferring protein (OSCP). Acrylodan bound covalently to the single cysteinyl residue of the protein. Acrylodan-OSCP was fully competent in conferring oligomycin sensitivity to the mitochondrial F0-F1 ATPase complex. The fluorescence emission peak of acrylodan-OSCP was blue-shifted compared to that of an acrylodan-mercaptoethanol adduct, which means that Acrylodan experiences a hydrophobic environment in OSCP. Binding of acrylodan-OSCP to the isolated F1 was accompanied by a red shift of fluorescence. It was achieved in less than 1 s at 25 degrees C. The titration curve revealed one high affinity OSCP binding site per F1. Acrylodan-OSCP appears to be an interesting tool for studying the dynamics of structural changes within the mitochondrial ATPase complex.
Kinetics of acrylodan-labelled cAMP-dependent protein kinase catalytic subunit denaturation
Protein J 2013 Oct;32(7):519-25.PMID:24048767DOI:10.1007/s10930-013-9511-4.
Fluorescence spectroscopy was used to study denaturation of cAMP-dependent protein kinase catalytic subunit labeled with an Acrylodan moiety. The dye was covalently bound to a cystein residue introduced into the enzyme by replacement of arginine in position 326 in the native sequence, located near the enzyme active center. This labeling had no effect on catalytic activity of the enzyme, but provided possibility to monitor changes in protein structure through measuring the fluorescence spectrum of the dye, which is sensitive to changes in its environment. This method was used to monitor denaturation of the protein kinase catalytic subunit and study the kinetics of this process as well as influence of specific ligands on stability of the protein. Stabilization of the enzyme structure was observed in the presence of adenosine triphosphate, peptide substrate RRYSV and inhibitor peptide PKI[5-24].