A bioluminescent luciferin
Coelenterazine is a substrate of various luciferases such as Renilla luciferase (Rluc) and Gaussia luciferase (Gluc), and is also a cofactor of aequorin. It is widely used and can be used for detection Calcium concentration in living cells, gene reporter assays, BRET (Bioluminescence Resonance Energy Transfer) studies, ELISA, HTS, and chemiluminescence detection of ROS levels in tissues or cells[1].
In vitro, coelenterazine enabled the dynamic monitoring of the RAW 264.7 cell oxidative burst [2].Coelenterazine chemiluminescence did not depend on the activity of cell-derived myeloperoxidase.It is a very sensitive and specific superoxide anion chemiluminescent probe[3]. Coelenterazine is a more reliable probe, although susceptible to storage-dependent high background chemiluminescence, but does not require prereductio[4].
In vivo, there were early, preclinical, changes in the localization and magnitude of coelenterazine chemiluminescent foci[2]. Commercially available coelenterazine（soluble in DMSO） is a sensitive substrate for in vivo bioluminescence imaging of coelenterazine luciferase[5].
参考文献:
[1]Jiang T, Du L, Li M. Lighting up bioluminescence with coelenterazine: strategies and applications. Photochem Photobiol Sci. 2016 Apr;15(4):466-80.
[2] Bronsart L, Nguyen L, et al. Reactive Oxygen Species Imaging in a Mouse Model of Inflammatory Bowel Disease. Mol Imaging Biol. 2016 Aug;18(4):473-8.
[3]Teranishi K. Non-invasive and accurate readout of superoxide anion in biological systems by near-infrared light. Anal Chim Acta. 2021 Sep 22;1179:338827.
[4] Kervinen M, P?tsi J, Finel M, Hassinen IE. Lucigenin and coelenterazine as superoxide probes in mitochondrial and bacterial membranes. Anal Biochem. 2004 Jan 1;324(1):45-51.?
[5]Lucas M, Solano F. Coelenterazine is a superoxide anion-sensitive chemiluminescent probe: its usefulness in the assay of respiratory burst in neutrophils. Anal Biochem. 1992 Nov 1;206(2):273-7.
Coelenterazine 是多种荧光素酶的底物，例如 Renilla 荧光素酶 (Rluc) 和 Gaussia 荧光素酶 (Gluc)，也是水母发光蛋白的辅助因子。用途广泛，可用于检测活细胞中的钙离子浓度、基因报告基因检测、BRET（生物发光共振能量转移）研究、ELISA、HTS，以及组织或细胞中ROS水平的化学发光检测[1]< /sup>.
在体外，腔肠素能够动态监测RAW 264.7细胞的氧化爆发[2]。腔肠素化学发光不依赖于细胞来源的髓过氧化物酶的活性。它是一种非常灵敏和特异的超氧阴离子化学发光探针[3].腔肠素是一种更可靠的探针，虽然对存储依赖性高背景化学发光敏感，但不需要预还原[4]。
在体内，腔肠素化学发光灶的定位和大小在早期、临床前发生了变化[2]。市售腔肠素（溶于DMSO）是腔肠素荧光素酶体内生物发光成像的敏感底物[5]。
| Cell experiment [1]: | |
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Cell lines |
RAW 264.7 cells |
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Preparation Method |
5 × 104 cRAW 264.7 cells were added to wells of a 96-well black plate in triplicate, cell counts were performed, and native coelenterazine was added to the sample (final coelenterazine concentration 0-100 μmol/l). Plates were imaged using an IVIS low-light imaging system |
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Reaction Conditions |
final coelenterazine concentration 0-100 μmol/l，60min |
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Applications |
RAW 264.7 chemiluminescent signal intensity positively correlates with coelenterazine concentration in culture. The production of chemiluminescence by RAW 264.7 cells in the absence of stimulation is consistent with the detection of superoxide anion production during oxidative phosphorylation. |
| Animal experiment [2]: | |
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Animal models |
10-26 weeks thirty-six mice (16 females, 20 males) |
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Preparation Method |
450 nl of virus (three sites) were injected into the cerebral cortex of mice, followed by coelenterazine injections at 28-day intervals (minimum = 26 days, maximum time = 74 days). Coelenterazine (dissolved in DMSO) and diluted to a concentration of 2.43 mM in sterile water (1 mg/ml) for cortical injection through a 34-gauge needle for imaging and electrophysiological recordings. |
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Dosage form |
0.2μl, 0.4μl and 1μl，the injection rate was 1.25 μL/min |
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Applications |
Coelenterazine strongly boosted activity in LMO3-expressing mice, photon production was proportional to emission rate, and maximal doses of coelenterazine moderately increased neural activity in naive mice and GLuc cohorts, which itself can be produced at high concentrations Nonspecific excitatory effects. |
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参考文献: [1]. Bronsart L, Nguyen L, Habtezion A, Contag C. Reactive Oxygen Species Imaging in a Mouse Model of Inflammatory Bowel Disease. Mol Imaging Biol. 2016 Aug;18(4):473-8. [2]. Gomez-Ramirez M, More AI,et al. The BioLuminescent-OptoGenetic in vivo response to coelenterazine is proportional, sensitive, and specific in neocortex. J Neurosci Res. 2020 Mar;98(3):471-480. |
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