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Furimazine (PBI 3939) Sale

目录号 : GC32913

Furimazine是一种咪唑并吡嗪酮类的发光底物。在哺乳动物细胞中,NanoLuc(Nluc)与Furimazine联用,产生更亮的发光,比带Coelenterazine的Oluc-19高250万倍。

Furimazine (PBI 3939) Chemical Structure

Cas No.:1374040-24-0

规格 价格 库存 购买数量
10mM (in 1mL DMSO)
¥3,533.00
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1mg
¥2,409.00
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5mg
¥3,442.00
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10mg
¥5,163.00
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25mg
¥9,293.00
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50mg
¥13,938.00
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Sample solution is provided at 25 µL, 10mM.

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Quality Control & SDS

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实验参考方法

Kinase experiment:

Random libraries are generated by error-prone PCR (average of 2-3 mutations per clone). Library 1 (phase 1; template=Oluc-N166R) is screened (4,400 variants) with Coelenterazine. Library 2 (phase 2; template=C1A4E) is screened (4,400 variants) with 11 novel Coelenterazine analogues: 3840, 3841, 3842, 3857, 3880, 3881, 3886, 3887, 3889, 3897, and 3900. The 11 analogues represent substitutions at positions 2, 6, and 8 and are considered to be representative of the entire set of 24 compounds; 2,200 variants are screened with compounds 3896 and 3894. All hits (improved luminescence) are screened again with the remaining Coelenterazine analogues. Library 3 (phase 3; template=C1A4E+Q18L/K33N/F54I/F68Y/L72Q/M75K/I90V) was screened in the context of a mouse Id-X-HaloTag (where X=library) using Coelenterazine and Furimazine. Library screens are performed on a Freedom robotic workstation as follows: induced bacterial cultures (in 96-well microtiter plates) are lysed with a buffer containing 300 mM HEPES pH 8, 200 mM thiourea, 0.3X Passive Lysis Buffer, 0.3 mg/mL lysozyme, and 0.002 units of RQ1 DNase. Assay reagent containing 1 mM CDTA, 150 mM KCl, 10 mM DTT, 0.5% (v/v) Tergitol, and 20 μM substrate is then added to equal volumes of lysate. Samples are measured on a GENios Pro luminometer[1].

References:

[1]. Hall MP, et al. Engineered luciferase reporter from a deep sea shrimp utilizing a novel imidazopyrazinone substrate. ACS Chem Biol. 2012 Nov 16;7(11):1848-57.

产品描述

Furimazine is an imidazopyrazinone substrate. NanoLuc (Nluc) paired with Furimazine produced 2.5 million-fold brighter luminescence in mammalian cells relative to Oluc-19 with Coelenterazine.

Furimazine is a Coelenterazine analogue.The apparent Km for purified NanoLuc (Nluc) using either Furimazine or Coelenterazine is ~10 μM, while the maximum luminescence (i.e., apparent Vmax) is ~30-fold higher for Furimazine than for native Coelenterazine. Measurement of luminescence intensity from cells expressing Nluc reveals that maximal signal is attained at about 10-20 μM Furimazine[1].

[1]. Hall MP, et al. Engineered luciferase reporter from a deep sea shrimp utilizing a novel imidazopyrazinone substrate. ACS Chem Biol. 2012 Nov 16;7(11):1848-57.

Chemical Properties

Cas No. 1374040-24-0 SDF
Canonical SMILES O=C1C(CC2=CC=CO2)=NC3=C(CC4=CC=CC=C4)NC(C5=CC=CC=C5)=CN31
分子式 C24H19N3O2 分子量 381.43
溶解度 DMSO : ≥ 100 mg/mL (262.17 mM) 储存条件 Store at 2-8°C
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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。
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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。

溶解性数据

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1 mg 5 mg 10 mg
1 mM 2.6217 mL 13.1086 mL 26.2171 mL
5 mM 0.5243 mL 2.6217 mL 5.2434 mL
10 mM 0.2622 mL 1.3109 mL 2.6217 mL
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Research Update

Novel Furimazine derivatives for nanoluciferase bioluminescence with various C-6 and C-8 substituents

Org Biomol Chem 2021 Sep 22;19(36):7930-7936.PMID:34549229DOI:10.1039/d1ob01098k.

Nanoluciferase (NLuc) is the emerging commercially available luciferase considering its small size and superior bioluminescence performance. Nevertheless, this bioluminescence system has some limitations, including narrow emission wavelength and single substrate. Herein, a series of novel Furimazine derivatives at the C-6 and C-8 positions of the imidazopyrazinone core have been designed and synthesized for extension of the bioluminescence substrates. It should be noted that two compounds, molecules A2 (2-(furan-2-ylmethyl)-6-(4-(hydroxymethyl)phenyl)-8-(phenylthio)imidazo[1,2-a]pyrazin-3(7H)-one) and A3 (2-(furan-2-ylmethyl)-6-(4-amino-3-fluorophenyl)-8-(phenylthio)imidazo[1,2-a]pyrazin-3(7H)-one), display reasonable bioluminescence properties for in vitro and in vivo biological evaluations. In particular, compound A3 can broaden the application of NLuc bioluminescence techniques, especially for in vivo bioluminescent imaging.

A Series of Furimazine Derivatives for Sustained Live-Cell Bioluminescence Imaging and Application to the Monitoring of Myogenesis at the Single-Cell Level

Bioconjug Chem 2022 Mar 16;33(3):496-504.PMID:35184558DOI:10.1021/acs.bioconjchem.2c00035.

Bioluminescence (BL) imaging, which utilizes light emitted through the enzymatic reaction of luciferase oxidizing its substrate luciferin, enables sensitive and noninvasive monitoring of life phenomena. Herein, we developed a series of caged Furimazine (FMZ) derivatives by introducing a protective group at the C-3 position and a hydroxy group at the C-6 phenyl ring to realize long-term live-cell BL imaging based on the NanoLuc (NLuc)/NanoKAZ (NKAZ)-FMZ system. The membrane permeability and cytotoxicity of the substrates were evaluated and related to their hydrophobicity. Among the series, the derivative with the bulkiest protective group (adamantanecarbonyl group) and a hydroxy substituent (named Ad-FMZ-OH) showed significantly prolonged and constant BL signal in cells expressing NLuc compared to the native FMZ substrate. This derivative enabled continuous BL imaging at the single-cell level for 24 h. Furthermore, we applied Ad-FMZ-OH to BL imaging of myocyte fusion and succeeded in the consecutive and sensitive monitoring at a single-cell level over a day. In summary, NLuc/NKAZ-caged FMZ derivatives have the potential to be applied to live-cell BL imaging of various life phenomena that require long-term observation.

Novel NanoLuc substrates enable bright two-population bioluminescence imaging in animals

Nat Methods 2020 Aug;17(8):852-860.PMID:32661427DOI:10.1038/s41592-020-0889-6.

Sensitive detection of two biological events in vivo has long been a goal in bioluminescence imaging. Antares, a fusion of the luciferase NanoLuc to the orange fluorescent protein CyOFP, has emerged as a bright bioluminescent reporter with orthogonal substrate specificity to firefly luciferase (FLuc) and its derivatives such as AkaLuc. However, the brightness of Antares in mice is limited by the poor solubility and bioavailability of the NanoLuc substrate Furimazine. Here, we report a new substrate, hydrofurimazine, whose enhanced aqueous solubility allows delivery of higher doses to mice. In the liver, Antares with hydrofurimazine exhibited similar brightness to AkaLuc with its substrate AkaLumine. Further chemical exploration generated a second substrate, fluorofurimazine, with even higher brightness in vivo. We used Antares with fluorofurimazine to track tumor size and AkaLuc with AkaLumine to visualize CAR-T cells within the same mice, demonstrating the ability to perform two-population imaging with these two luciferase systems.

Novel NanoLuc-type substrates with various C-6 substitutions

Bioorg Med Chem Lett 2020 May 1;30(9):127085.PMID:32171617DOI:10.1016/j.bmcl.2020.127085.

NanoLuc (NLuc)-furimazine bioluminescence system offers several advantages over established systems, including improved stability, smaller size, and >150-fold enhancement in bioluminescence. Herein, we designed and synthesized a series of bioluminescent substrates with varying at the C-6 position of Furimazine for NLuc-furimazine bioluminescence system. Among all derivatives, compounds A6 and A11 provided excellent bioluminescence characteristics compared with Furimazine in vitro and in vivo. We believe that these new NLuc substrates can broaden the application of NLuc bioluminescence techniques, especially in vivo bioluminescent imaging.

Nanoluciferase signal brightness using Furimazine substrates opens bioluminescence resonance energy transfer to widefield microscopy

Cytometry A 2016 Aug;89(8):742-6.PMID:27144967DOI:10.1002/cyto.a.22870.

Fluorescence and bioluminescence resonance energy transfer (FRET, BRET) techniques are powerful tools for studying protein-protein interactions in cellular assays. In contrast to fluorescent proteins, chemiluminescent proteins do not require excitation light, known to trigger autofluorescence, phototoxicity, and photobleaching. Regrettably, low signal intensity of luciferase systems restricts their usage as they require specialized microscopes equipped with ultra low-light imaging cameras. In this study, we report that bioluminescence quantification in living cells using a standard widefield automated microscope dedicated to screening and high content analysis is possible with the newer luciferase systems, Nanoluciferase (Nluc). With such equipment, we showed that robust intramolecular BRET can be measured using a combination of Nluc and yellow fluorescent protein (YFP). Using the human Superoxide Dismutase 1 (SOD1) dimer model, we next validated that intermolecular BRET could be quantified at a single cell level. The enhanced signal brightness of Nluc enabling BRET imaging to widefield microscopy shows strong potential to open up single cell protein-protein interactions studies to a wider audience. © 2016 International Society for Advancement of Cytometry.