The chemicals used in the luciferase assay were as follows: FMN-Na (Alfa Aesar, J66949.09), NADPH (Cayman Chemical Company, 9000743), ATP (Larova GmbH, ATP_100ML), D-luciferin (Cayman Chemical Company, 25836), coelenterazine (Cayman Chemical Company, 16123), coelenterazine H (Promega, S2011), frimazine (Aoblous, AOB36539), hispidin (Cayman Chemical Company, 10012605), octanaldehyde (Fisher Scientific, O004425ML), decyl aldehyde (Fisher Scientific, AC154971000), dodecyl aldehyde (fisher scientific), octanoic acid (Fisher Scientific, O002725ML), decanoic acid (Fisher Scientific, AC167271000), dodecanoic acid (Fisher Scientific, S25377), tetradecanoic acid (Fisher Scientific, AAA1206730). D-luciferin, furimazine, and hispidin were dissolved in DMSO as 10 mM stocks. coelenterazine and coelenterazine H were dissolved in ethanol as 10 mM stocks. Long-chain fatty aldehydes and acids were dissolved in ethanol as 500 mM stocks. Dodecyl aldehyde was not soluble in 100% ethanol at the concentration of 500 mM; we used the suspension with vortexing each time.
Coelenterazine
Manufactured by Cayman Chemical
Coelenterazine is a bioluminescent substrate that emits light when combined with the enzyme luciferase. It is a key component in various assays and research applications that utilize bioluminescence as a detection method.
Automatically generated - may contain errors
Lab products found in correlation
Coelenterazine h, by Promega (1 mentions)
Nadph, by Cayman Chemical (1 mentions)
Octanoic acid, by Thermo Fisher Scientific (1 mentions)
D luciferin, by Cayman Chemical (1 mentions)
Spectramax l, by Molecular Devices (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
2 protocols using coelenterazine
1
Luciferase Assay Reagents Protocol
2
Monitoring GPCR-Effector Interactions using NanoBiT
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The NanoBiT system, which employs split luciferase consisting of small and large fragments (SmBiT and LgBiT, respectively) and was previously utilized to monitor interactions of GPCRs with ß-arrestin or specific Gα/βγ complexes in response to GPCR activation (34) was used to characterize ApoA1-ApoM nanoparticles containing S1P. Briefly, HEK293A cells maintained in DMEM (GIBCO) supplemented with 10% FCS and Pen-Strep, were dispersed into 6 well plates and allowed to adhere overnight. For functional studies, cells were transfected with appropriate combinations of reporter plasmids. S1PR1-SmBiT and LgBiT-beta-arrestin fusion proteins or Gα i -SmBiT combined with Gβ 1 and LgBiT-Gγ 1 were employed as described previously (34) . After 24hrs, cells were harvested, resuspended with the luciferase substrate Coelenterazine (Cayman Chem; 50 µM), dispersed into white opaque-bottom 96-well plates (Greiner) and maintained at RT for 2 hours to quench background. S1P containing samples were added by multi-channel pipetting and the plate was immediately analyzed for luminenescence for 30 minutes in a SpectraMax L 96-well plate reader (Molecular Devices).
Data was integrated as described (34) .
Data was integrated as described (34) .
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!