Determination of NADH or NADPH Concentrations with the FL600TM Fluorescence Microplate Reader using Fluorescence or Absorbance Modes

Many biological assays have as their basis a link to the oxidative status of nicotinamide adenine dinucleotide (NAD) or nicotinamide adenine dinucleotide phosphate (NADP). Many dehydrogenase enzymes use these coenzymes to transfer hydrogen groups between molecules. Because the reduced forms of these molecules differ from the oxidized forms in their ability to absorb light, it is possible to quantitate reactions based on light absorbance at 340 nm or by the fluorescent emission of light at 445 nm. Here we describe the use of the FL600 fluorescence microplate reader to quantitate NADH or NADPH using either fluorescence or absorbance modes.