NAD/NADH-Glo Assay, Promega
Supplier: Promega Corporation
The NAD/NADH-Glo Assay is a bioluminescent, homogeneous single-reagent-addition assay for detecting total oxidized and reduced nicotinamide adenine dinucleotides and determining their ratio in biological samples or in defined enzyme reactions.
- Bioluminescent Assays to Detect NAD(P)+ and NAD(P)H Levels in Cells
- Homogeneous, single-reagent-addition assays
- Easily adapted for high throughput screening formats
The NAD/NADH-Glo Assay is a bioluminescent, homogeneous single-reagent-addition assay for detecting total oxidized and reduced nicotinamide adenine dinucleotides (NAD+ and NADH, respectively) and determining their ratio in biological samples or in defined enzyme reactions. An NAD Cycling Enzyme is used to convert NAD+ to NADH. In the presence of NADH, the provided reductase enzyme reduces a proluciferin reductase substrate to form luciferin. Luciferin then is quantified using Ultra-Glo Recombinant Luciferase, and the light signal produced is proportional to the amount of NAD+ and NADH in the sample. Cycling between NAD+ and NADH by the NAD Cycling Enzyme and Reductase increases assay sensitivity and provides selectivity for the nonphosphorylated NAD+ and NADH compared to the phosphorylated forms NADP+ and NADPH. The NAD Cycling Enzyme, Reductase and luciferase reactions are initiated by adding an equal volume of NAD/NADH-Glo Detection Reagent, which contains NAD Cycling Enzyme and Substrate, Reductase, Reductase Substrate and Ultra-Glo Recombinant Luciferase, to an NAD+- or NADH-containing sample. Detergent present in the reagent lyses cells, allowing detection of total cellular NAD+ and NADH in a multiwell format with addition of a single reagent. An accessory protocol is provided to allow separate measurements of NAD+ and NADH, and calculation of the NAD+ to NADH ratio. The simple add-mix-read protocol and scalable assay chemistry make the NAD/NADH-Glo Assay well suited to monitor effects of small molecule compounds on NAD and NADH levels in high-throughput formats.
NAD/NADH-Glo Assay, 10 mL contains enough reagents to perform 100 reactions in 96-well plates (100 μL of sample and 100 μL of Reagent), 400 assays in 384-well plates (25 μL of sample and 25 μL of Reagent), or 2000 assays in low-volume 384-well plates (5 μL of sample and 5 μL of Reagent). NAD/NADH-Glo Assay, 50 mL contains enough reagents to perform 500 reactions in 96-well plates (100 μL of sample and 100 μL of Reagent), 2000 assays in 384-well plates (25 μL of sample and 25 μL of Reagent), or 10,000 assays in low-volume 384-well plates (5 μL of sample and 5 μL of Reagent). Assay volumes can be varied depending on plate format as long as a 1:1 ratio of sample to NAD/NADH-Glo Detection Reagent is maintained.
In the presence of NADH, the provided reductase enzyme reduces a pro-luciferin reductase substrate to form luciferin. Ultra-Glo™ Recombinant Luciferase then quantifies luciferin, and the light signal produced is proportional to the amount of NAD+ and NADH in the sample. Cycling between NAD+ and NADH by the NAD Cycling Enzyme and Reductase provides selectivity for the nonphosphorylated NAD+ and NADH compared to the phosphorylated forms NADP+ and NADPH. The NAD Cycling Enzyme, reductase, and luciferase reactions are initiated by adding an equal volume of NAD/NADH-Glo Detection Reagent, which contains NAD Cycling Enzyme and Substrate, Reductase, Reductase Substrate, and Ultra-Glo Recombinant Luciferase, to an NAD+ or NADH-containing sample. Detergent is present in the reagent lyses cells for detection of total cellular NAD+ and NADH in a multiwell format with addition of a single reagent. An accessory protocol is provided to allow separate measurements of NAD+ and NADH and calculation of the NAD+ to NADH ratio. The add-mix-read protocol and scalable assay chemistry make the NAD/NADH-Glo Assay well suited to monitor effects of small molecule compounds on NAD and NADH levels in high-throughput formats.
Learn more
About VWR
Avantor is a vertically integrated, global supplier of discovery-to-delivery solutions for...