Coelenterazine 400a |
Catalog No.GC43290
One-Click Copy Product Info
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Coelenterazine 400a, a derivative of natural coelenterazine, functions as a substrate for Renilla luciferase (Rluc), with a maximum emission wavelength of 400nm.
Products are for research use only. Not for human use. We do not sell to patients.
Cas No.: 70217-82-2
Sample solution is provided at 25 µL, 10mM.
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Coelenterazine 400a, a derivative of natural coelenterazine[1], functions as a substrate for Renilla luciferase (Rluc)[2], with a maximum emission wavelength of 400nm. In the presence of Rluc, Coelenterazine 400a reacts with molecular oxygen, forming high-energy intermediates and emitting light as a result[3][4]. Its minimal interference with fluorescent acceptors makes it an ideal choice for bioluminescence resonance energy transfer (BRET) assays, commonly used to evaluate protein-protein interactions[5][6].
References:
[1]. Sousa J, Magalhães CM, González-Berdullas P, et al. Comparative Investigation of the Chemiluminescent Properties of a Dibrominated Coelenterazine Analog. Int J Mol Sci. 2022 Jul 30;23(15):8490. doi: 10.3390/ijms23158490. PMID: 35955625; PMCID: PMC9369366.
[2]. Markova SV, Vysotski ES. Coelenterazine-dependent luciferases. Biochemistry (Mosc). 2015 Jun;80(6):714-32. doi: 10.1134/S0006297915060073. PMID: 26531017.
[3]. Min CG, Ferreira PJO, Pinto da Silva L. Theoretically obtained insight into the mechanism and dioxetanone species responsible for the singlet chemiexcitation of Coelenterazine. J Photochem Photobiol B. 2017 Sep;174:18-26. doi: 10.1016/j.jphotobiol.2017.07.012. Epub 2017 Jul 20. PMID: 28750319.
[4]. Vacher M, Fdez Galván I, Ding BW, et al. Chemi- and Bioluminescence of Cyclic Peroxides. Chem Rev. 2018 Aug 8;118(15):6927-6974. doi: 10.1021/acs.chemrev.7b00649. Epub 2018 Mar 1. PMID: 29493234.
[5]. Ming-Liang Yuan, Tian-Yu Jiang, Lu-Pei Du, et al. Luminescence of coelenterazine derivatives with C-8 extended electronic conjugation. Chinese Chemical Letters. 2016, 27(04): 550-554 https://doi.org/10.1016/j.cclet.2016.02.011
[6]. Li J, Wang X, Dong G, et al. Novel furimazine derivatives for nanoluciferase bioluminescence with various C-6 and C-8 substituents. Org Biomol Chem. 2021 Sep 22;19(36):7930-7936. doi: 10.1039/d1ob01098k. PMID: 34549229.
Experimental Protocol for Coelenterazine 400a in BRET assay
1.Preparation of Stock Solution: Dissolve Coelenterazine 400a in 100% ethanol or methanol to prepare a stock solution with a concentration of 1mM. Aliquote the stock solution and store at -20°C or -80°C in the dark.
2.Preparation of Sample: Culture cells in an appropriate medium until they reach 70-80% confluence. Use suitable transfection reagents to co-transfect cells with Renilla luciferase and fluorescent acceptor plasmids. After transfection, continue to culture the cells for 24-48 hours to ensure adequate protein expression.
3.Preparation of Substrate Solution: Prior to use, dilute the Coelenterazine 400a stock solution to a concentration of 100μM with cell culture medium.
4.BRET Assay: Wash the transfected cells and resuspend them in phosphate-buffered saline (PBS). Add 90µL of the cell suspension to each well of a 96-well white microplate. Add 10µL substrate solution to each well to achieve a final concentration of 10µM.
5.Fluorescence Measurement: After adding Coelenterazine 400a, measure the luminescence intensity using a multimode microplate reader.
This protocol only provides a guideline and should be modified according to your specific needs.
References:
[1]. Bery N, Rabbitts TH. A Cell-based Screening Method Using an Intracellular Antibody for Discovering Small Molecules Targeting Hard-to-drug Proteins. Bio Protoc. 2022 Feb 20;12(4):e4324. doi: 10.21769/BioProtoc.4324. PMID: 35340285; PMCID: PMC8899552.
| Cas No. | 70217-82-2 | SDF | |
| Canonical SMILES | O=C1C(CC2=CC=CC=C2)=NC3=C(CC4=CC=CC=C4)NC(C5=CC=CC=C5)=CN31 | ||
| Formula | C26H21N3O | M.Wt | 391.5 |
| Solubility | 1mg/mL in DMSO, Ethanol: 0.5 mg/mL, Methanol: 0.5 mg/mL | Storage | Store at -20°C,protect from light |
| General tips | Please select the appropriate solvent to prepare the stock solution according to the
solubility of the product in different solvents; once the solution is prepared, please store it in
separate packages to avoid product failure caused by repeated freezing and thawing.Storage method
and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored
at -20°C, please use it within 1 month. To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time. |
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| Shipping Condition | Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request. | ||
| Prepare stock solution | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 2.5543 mL | 12.7714 mL | 25.5428 mL |
| 5 mM | 510.9 μL | 2.5543 mL | 5.1086 mL |
| 10 mM | 255.4 μL | 1.2771 mL | 2.5543 mL |
Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)
g
μL
Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)
Calculation results:
Working concentration: mg/ml;
Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )
Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL saline, mix and clarify.
Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.
Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.
3. All of the above co-solvents are available for purchase on the GlpBio website.
Quality Control & SDS
- View current batch:
- Purity: >97.00% Appearance: A solid
- COA (Certificate of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Average Rating: 5 (Based on Reviews and 37 reference(s) in Google Scholar.)