Donna Leippe, Mary Sobol, Jolanta Vidugiriene, *Wenhui Zhou, Gediminas Vidugiris, *Troy Good, *Laurent Bernad, *Poncho Meisenheimer, James Cali
Promega Corporation, 2800 Woods Hollow Rd, Madison, WI 53711;
*Promega BioSciences, LLC, 227 Granada Dr, San Luis Obispo, CA 93401
We have developed a bioluminescent method for monitoring changes in NAD, NADP, NADH and NADPH levels using a homogeneous, one-step, add-and-read protocol for biochemical or cell-based detection. The assays are sensitive, capable of detecting ≤50 nM of these dinucleotides, and have large maximum signal windows (S/B) of ≥100, providing advantages over currently available colorimetric and fluorescent detection methods. The high sensitivity, one-step reagent addition, and no requirement for sample processing, permit an in-well protocol for the assay of cells directly in culture plate wells. Changes in cellular nicotinamide dinucleotide levels can be rapidly monitored in a high-throughput format, facilitating the study of metabolic pathways and the development of therapeutics that target cancer cell metabolism.