Cycling assay for nicotinamide adenine dinucleotide in human plasma
|Enzymatic reactant recycling assay
|Philipp Brunnbauer, Felix Krenzien
|31 October 2018
|Charité - Universitätsmedizin Berlin
A cost-effective, high-throughput assay for measuring extracellular NAD+ in blood and other human body fluids with a high sensitivity, ranging into low nanomolar concentrations.
A research group (Philipp Brunnbauer, Felix Krenzien and Annekatrin Leder) in the department of Experimental Surgery from the Charité-Universitätsmedizin Berlin have developed a novel robust assaying method for the detection of low extracellular NAD+ (eNAD+) levels in various body fluids, focusing on human heparinized plasma. This tool provides a simple, time and cost-effective method, that can be re-created in almost every lab using off-the-shelf components in contrast to other standard analytical methods, such as elaborate high-pressure liquid chromatography-mass spectrometry (LC-MS).
The novel standardized method is suitable for high throughput measurements of extracellular NAD+ (and other members of the pyridine metabolome) levels in human body fluid. This highly sensitive colorimetric assay is capable of quantifying extracellular NAD+ concentrations in the low micromolar into the nanomolar range. The underlying assay protocol is ideal for the development of a direct-to-consumer lab testing product or the establishment of a laboratory standard for population-wide screening efforts (eNAD+ in ageing).
The underlying methodology is based on the standard enzymatic alcohol dehydrogenase dependent NAD+ cycling reaction and kinetics, which was modified in an innovative fashion to accurately and reliably measure in particular very low quantities of eNAD+ in plasma samples. Notably, assay conditions can be adapted and miniaturized for high-throughput analysis and for other bodily fluids.
This opportunity is available for licensing agreements or acquisition of the IP rights.
Assay characteristics such as assay linearity, measurement range and reproducibility were systematically evaluated according to best practices for biochemical measurement techniques against synthetic eNAD+ samples and were validated in real human heparinised blood samples, demonstrating clinical utility.
Provisional US patent application at the USPTO under application number 62/903,111 (09/20/2019). Name: “Methods, kits and devices for measuring extracellular pyridine nucleotide.”
P. Brunnbauer and A. Leder et al., "The nanomolar sensing of nicotinamide adenine dinucleotide in human plasma using a cycling assay in albumin modified simulated body fluids". Scientific Reports. 8. 2018-12-01. doi:10.1038/s41598-018-34350-6.
- Brunnbauer, Philipp; Leder, Annekatrin; Kamali, Can; Kamali, Kaan; Keshi, Eriselda; Splith, Katrin; Wabitsch, Simon; Haber, Philipp; Atanasov, Georgi; Feldbrügge, Linda; Sauer, Igor M. (2018-10-31). "The nanomolar sensing of nicotinamide adenine dinucleotide in human plasma using a cycling assay in albumin modified simulated body fluids". Scientific Reports. 8 (1): 1–15. doi:10.1038/s41598-018-34350-6. ISSN 2045-2322.