Cells were plated in 10-cm plates as previously described. Cells were transfected with plasmids encoding cDNA for the Glosensor reporter (Promega, Madison, WI), receptor, and Gα-subunit at a ratio of 2:1:1 (2 μg: 1 μg: 1μg). The next day, cells were plated in black, clear-bottom, 384-well white plates. After aspiration of the medium on the day of the assay, cells were incubated for 60 minutes at 37°C with 20 μL of 5 mM luciferin substrate (GoldBio, St. Louis, MO) freshly prepared in assay buffer. For Gαs activity, 10 μL of drugs were added using the FLIPR Tetra® liquid-handling robot and read after 15 minutes in a Spectramax luminescence plate reader (Molecular Devices, San Jose, CA) with a 0.5 second signal integration time. For Gαi activity, 10 μL of drugs were added for a 15-minute incubation period. Subsequently, 10 μL of isoproterenol (final concentration of 200 nM) was added and incubated for an additional 15-minute period before reading.
Spectramax luminescence plate reader
Manufactured by Molecular Devices
The SpectraMax luminescence plate reader is a versatile instrument designed for detecting and quantifying luminescent signals in biological samples. It features a sensitive detection system capable of measuring a wide range of luminescent signals, making it suitable for a variety of assays and applications.
Automatically generated - may contain errors
2 protocols using spectramax luminescence plate reader
1
Glosensor Assay for Gα-Coupled Receptors
2
Glosensor Assay for Gα-Coupled Receptors
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were plated in 10-cm plates as previously described. Cells were transfected with plasmids encoding cDNA for the Glosensor reporter (Promega, Madison, WI), receptor, and Gα-subunit at a ratio of 2:1:1 (2 μg: 1 μg: 1μg). The next day, cells were plated in black, clear-bottom, 384-well white plates. After aspiration of the medium on the day of the assay, cells were incubated for 60 minutes at 37°C with 20 μL of 5 mM luciferin substrate (GoldBio, St. Louis, MO) freshly prepared in assay buffer. For Gαs activity, 10 μL of drugs were added using the FLIPR Tetra® liquid-handling robot and read after 15 minutes in a Spectramax luminescence plate reader (Molecular Devices, San Jose, CA) with a 0.5 second signal integration time. For Gαi activity, 10 μL of drugs were added for a 15-minute incubation period. Subsequently, 10 μL of isoproterenol (final concentration of 200 nM) was added and incubated for an additional 15-minute period before reading.
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!