Total ATP content of SH-SY5Y cells was determined using a bioluminescence assay (ViaLighTM HT, Cambrex Bio Science, Walkersville, MD, USA) according to the instruction of the manufacturer, as previously described [35, 36 (link)]. SH-SY5Y cells were plated in 5 replicates into a white 96-well cell culture plate at a density of 2x104 cells/well. The bioluminescent method measures the formation of light from ATP and luciferin by luciferase. The emitted light was linearly related to the ATP concentration and was measured using the multilabel plate reader VictorX5 (Perkin Elmer).
Vialigh ht
Manufactured by Cambrex
Sourced in United States
The ViaLightTM HT is a high-throughput cell viability assay kit designed for the rapid quantification of cell proliferation and cytotoxicity. The kit utilizes a luminescent signal that is directly proportional to the number of viable cells present in the sample.
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Lab products found in correlation
4 protocols using vialigh ht
1
Measuring ATP Content in SH-SY5Y Cells
2
Mitochondrial Bioenergetics in Hippocampus
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Mitochondria were isolated from hippocampus as previously described.49 (link) Oxygen consumption rate was measured in isolated mitochondria from cortex and hippocampus using a Seahorse Bioscience (North Billerica, MA, USA) XF24Analyzer. ATP content from isolated mitochondria was determined by a bioluminescence assay (ViaLighTM HT; Cambrex Bio Science, Lonza, Basel, Switzerland).
3
Bioluminescence Assay for Total ATP Quantification
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Total ATP content was determined using a bioluminescence assay (ViaLighTM HT; Cambrex Bio Science) according to the instructions of the manufacturer, as previously described [26 (link)]. Cells were plated in 5 replicates into a white 96-well cell culture plate at a density of 1.5 × 104 cells/well. The bioluminescent method measures the formation of light from ATP and luciferin by luciferase. The emitted light was linearly related to the ATP concentration and was measured using multilabel plate reader VictorX5 (Perkin Elmer).
4
ATP, NAD+, and NADH Quantification in Fibroblasts
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The total ATP content from synchronized human skin fibroblasts was determined using a bioluminescence assay (ViaLighTM HT; Cambrex Bio Science, Walkersville, MD USA) according to the instruction of the manufacturer. Cells were plated in 8 replicates into a 96-wells cell culture plate at a cell density sufficient to reach 40–50% of confluency on the following day. The enzyme luciferase, which catalyzes the formation of light from ATP and luciferin was used. The emitted light is linearly related to the ATP concentration and is measured using a multilabel plate reader VictorX5 (Perkin Elmer).
To measure NAD+ and NADH, the two molecules were separately extracted using an acid-base extraction method (HCL 0.1 mol/l–NAOH 0.1 mol/l). To determinate both NAD+ and NADH, an assay was used that is based on passing the electron from ethanol through reduced pyridine nucleotides to MTT [3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] in a PES (phenazine ethosulfate)-coupled reaction resulting in the formation of a purple precipitate (formazan) that, once dissolved, can be quantified at 595 nm (VictorX5, Perkin Elmer).
To measure NAD+ and NADH, the two molecules were separately extracted using an acid-base extraction method (HCL 0.1 mol/l–NAOH 0.1 mol/l). To determinate both NAD+ and NADH, an assay was used that is based on passing the electron from ethanol through reduced pyridine nucleotides to MTT [3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] in a PES (phenazine ethosulfate)-coupled reaction resulting in the formation of a purple precipitate (formazan) that, once dissolved, can be quantified at 595 nm (VictorX5, Perkin Elmer).
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