Notes: The NADPH/NADP and GSH/GSSG ratios were measured in cells to determine that CPT1A is required to maintain ROS homeostasis. (5039)

Notes: The ATP levels of human iPSC-derived cardiomyocytes were measured using the Mitochondrial ToxGlo™ Assay. To assess cell viability and caspase-3/7 activity, the ApoTox-Glo™ Triplex Assay was used and luminescence measured on a GloMax® plate reader for the caspase-3/7 assay. Cardiomyocytes were lysed and NAD+ and NADH measured from the same well using the NAD/NADH-Glo™ Assay to assess any changes in the NAD+/NADH ratio. (4952)

Notes: To determine relative NAD+ levels in bacteria, overnight cultures of E. coli strains with expression plasmids were diluted into LB broth, grown to mid-log phase and protein expression induced. After 1 hour, cultures were diluted to A₆₀₀ = 0.5, 500µl of cells pelleted, lysed and treated with 0.4M HCl for 15 minutes. After neutralizing the cells, the lysate was mixed with an equal volume of NAD/NADH-Glo™ Detection Reagent and luminescence measured continuously for 2 hours. The measurements were compared to an E. coli strain with a control vector to calculate relative NAD+. (4950)

Notes: This review article offers guidelines and protocols for assessing mitochondrial dysfunction and its role in human disease. The NAD/NADH-Glo™ Assay is mentioned in the section on mitochondrial NAD(P)H. (4949)

Notes: Human cultured cells were plated at 1,000 cells/well in 100µl of medium in 96-well plates. The next day, the growth medium was replaced with fresh medium or medium containing 4mM oxaloacetate or aspartate. After 3 days, cell viability was analyzed using the CellTiter-Glo® Luminescent Cell Viability Assay. To assess the ratio of NADPH to NADP+, cells were lysed, mixed with the NADP/NADPH-Glo® Assay reagents and luminescent measured after 30 minutes. (4948)

Notes: Cancer stem cells (CSCs; 10,000 cells/well in 96-well plates) were washed with PBS and 50µl of 1mM 2-Deoxy-d-Glucose (2DG) added. After 90 minutes, glucose uptake was measured using the Glucose Uptake-Glo™ Assay. CSCs were isolated from cell lines and plated at 10,000 cells/well in 96-well plates. After growing in defined medium supplemented with various amounts of glucose plus glutamine and growth factors for 2 days, cells were treated for 24 hours and 2µl samples removed and added to 98µl of PBS. Extracellular metabolites were assessed using the Glutamine/Glutamate-Glo™ Assay. Isolated CSCs were plated at 10,000 cells/well in 96-well plates, grown for 3 days, treated for 24 hours, medium removed and cells lysed. The lysates were treated to preserve NAD+ and NADH before mixing with the NAD/NADH-Glo™ Detection Reagent and luminescence measured. (4959)

Notes: Human primary hepatocytes at a density of 2 × 10⁴ were seeded into 96-well collagen-coated plates, incubated overnight and then stimulated with two compounds. After 48 hours, the levels of NAD+ was assessed using the NAD/NADH-Glo™ Assay. HepG2 and AML12 cells (5 × 10³) were treated with test compounds for 4 hours in a 384-well plate. Cell viability was measured using the CellTiter-Glo® Luminescent Cell Viability Assay and cell necrosis evaluated using the CytoTox-ONE™ Homogeneous Membrane Integrity Assay. (4953)

Notes: NAD⁺/NADH-Glo and NADP⁺/NADPH-Glo were used to determine levels of NAD+, NADH, NADP+ and NADPH. (4990)

Notes: NAD⁺/NADH-Glo and NADP⁺/NADPH-Glo were used to quantify NAD⁺, NADH, NADP⁺, and NADPH. (4992)

Notes: Various types of HeLa cell lines (Tet3G luciferase, LbNOX, mitoLbNOX, TPNOX or mitoTPNOX) cells were seeded at 400,000 cells per 6cm dish in 3ml of DMEM without pyruvate. After growing for 24 hours, the medium replaced, grown another 24 hours and the medium replaced again. After 2 hours, medium was removed, cells washed with ice-cold PBS, scraped and divided into two 200µl aliquots. One tube of lysate was left untreated, the other had ascorbic acid added to prevent oxidation of NAD(P)H into NAD(P)+. After heating at 60°C for 20 minutes, the levels of NAD+, NADH, NADP+ and NADPH were determined using the NAD/NADH-Glo™ or NADP/NADPH-Glo™ Assays. (4945)

Notes: ARPE-19 cells were seeded at 1 × 10⁴ cells per well in 96-well plates and incubated for 12 hours. Cells were treated with  H₂O₂ with or without olparib for 4 hours. Cellular NAD+ levels were measured with the NAD/NADH-Glo™ Assay.  Cellular ATP levels were measured with the CellTiter-Glo® Assay. (28292900)

Notes: The NAD/NADH-Glo™ Assay was used to assess whether or not overexpression of QPRT in K562 cells would increase cellular NAD levels and that was responsible for QPRT-associated resistance to imatinib. QPRT is in the de novo pathway for NAD synthesis from tryptophan. NAD levels did not rise upon QPRT upregulate but the decreased. (4829)

Notes: Six mosquitos per sample were frozen in liquid nitrogen and ground in a basic pH lysis buffer then heated to remove NAD+ then neutralized before measuring NADH with the NAD/NADH-Glo™ Assay System. To examine the effect of HNF4 on VLCAD and 3KCT gene expression, the authors employed gel shift assays and luciferase assays of the promoters in pGL4.10[luc2] and control with pGL4.73[hRluc/SV40] transfected into Drosophila S2 cells. Luciferase activities were measured with the Dual-Luciferase® Reporter Assay System on a GloMax® Instrument.  (4834)

Notes: The ratio of NADP+/NADPH was monitored upon BRCA1 knockdown in IOSE397 cells and FT33 fallopian tubes cells using the NADP/NADPH-Glo™ Assay.  The effect of BRCA1 knockdown on the HK2 promoter was evaluated in HEK293T cells using the Dual-Luciferase® Reporter Assay System. (4846)

Notes: In this study, the NADP/NADPH-Glo™ Assay was used to measure NADPH/NADP+ 48 hours after siRNA transfection. (4886)

Notes: Mouse islets were prepared from the pancreas by collagenase digestion and density gradient centrifugation, and then cultured for 2–5 days. The redox state of islets were assessed using batches of 20 islets preincubated for 40 minutes in 0.5mmol/L glucose before treating islets with 2–30mmol/L glucose for 15 minutes. The reaction was stopped and the cells sonicated before measuring NAD+, NADH, NADP+, and NADPH using the NAD/NADH-Glo™ and NADP/NADPH-Glo™ Assays. (4947)

Notes: The authors of this study explore the hypothesis that mitochondria pass glycolytic enzymes to the nucleus during early development to assist with producing substrates needed for epigenetic changes needed for activating the zygotic genome. Embryo extracts were analyzed with the ENLITEN® ATP Assay and the ADP-Glo™ Assay for ATP:ADP ratio; glutathione levels monitored with GSH-Glo™ Assay.  Extracts were also made as directed in the NAD/NADH-Glo™ Assay manual to determine NAD and NADH levels. (4831)

Notes: The NAD/NADH-Glo™ Assay was used to determine NAD/NADH ratios in E. coli using a referenced method for extraction. (4832)

Notes: Cell viability of IDH1 wild-type or mutant cell lines under treatment conditions was assessed using the CellTiter-Glo® Luminescent Cell Viability Assay. From these numbers, the IC₅₀ values were calculated. To evaluate apoptosis, 7,000–8,000 cells were treated with DMSO or compounds singly or in combination (12.5nmol/L NAMPT inhibitors and/or 200μmol/L temozolomide or 50μmol/L Z-VAD-FMK) for 6 or 24 hours and then measured using Caspase-Glo® 3/7 Assay. Changes in NAD+, NADH, NADP+, and NADPH were assessed using the NAD/NADH-Glo™ and NADP/NADPH-Glo™ Assays. (4946)

Notes: P7C3 compounds improve the production of NAD through the salvage pathway. NAD levels in rat brain homogenates were monitored with the NAD/NADH-Glo™ Assay using a referenced modification to the protocol. (4830)

Notes: NAD/NADH-Glo™ Assay was used to assess metabolic activity in cells. (4897)

Notes: This article referenced the NADP/NADPH-Glo™ Assay when describing products for bioluminescent analysis. (4885)

Notes: Mouse embryonic fibroblasts were treated with vehicle or DPQ for 1 hour followed by exposure to H₂O₂.  Intracellular NAD+ was measured with the NAD/NADH-Glo™ Assay and cellular ATP was measured with the CellTiter-Glo® Viability Assay. (4845)

Notes: B16 F10 murine melanoma cells were treated with fenofibrate and the ratios of NADP+/NADPH and GSH/GSSG were examined after PPARa knockdown or over-expression. The NADP+/NADPH levels did not change appreciably but the GSH/GSSG levels did fluctuate. The study used the NADP/NADPH-Glo™ and GSH/GSSG-Glo™ Assays. Assays were read on a GloMax® 20/20 Luminometer. (4852)

Notes: CellTiter-Glo® Cell Viability Assay was used to assess the vulnerability of myc-driven cancers to inhibitors of the NAMPT. Sensitivity was correlated to the level of myc expression. Apoptosis was assessed with the Caspase-Glo® 3/7 Assay. The NAD/NADH-Glo® Assay was used to provide a quantitative measure of NAD+ in the cells under study. (4835)