@entity19396 (cADPR) is a novel cyclic @entity377 derived from @entity2125 that has now been established as a general @entity141 messenger in a wide variety of cells. Despite the obvious importance of monitoring its cellular levels under various physiological conditions, its measurement has been technically difficult and requires specialized reagents. In this study a widely applicable high-sensitivity assay for cADPR is described. @entity857 - @entity13338 cyclase normally catalyses the synthesis of cADPR from @entity2125 , but the reaction can be reversed in the presence of high concentrations of @entity4571 , producing @entity2125 from cADPR stoichiometrically. The resultant @entity2125 can then be coupled to a cycling assay involving @entity167 dehydrogenase and @entity45299 . Each time @entity2125 cycles through these coupled reactions, a molecule of highly fluorescent @entity30368 is generated. The reaction can be conducted for hours, resulting in more than a thousand-fold amplification of cADPR. Concentrations of cADPR in the nanomolar range can be measured routinely. The unique ability of @entity857 - @entity13338 cyclase to catalyse the reverse reaction provides the required specificity. Using this assay, it is demonstrated that cADPR is present in all tissues tested and that the levels measured are directly comparable with those obtained using a radioimmunoassay. All the necessary reagents are widely available and the assay can be performed using a multiwell fluorescence plate reader, providing a high-throughput method for monitoring cADPR levels. This assay should be valuable in elucidating the messenger role of cADPR in cells.

A novel cycling assay for cellular XXXX with nanomolar sensitivity.

@entity2125 :: ('MESH:D009243', 'Chemical') :: ['NAD(+)'] @entity30368 :: ('CHEBI:51602', 'Chemical') :: ['resorufin'] @entity167 :: ('CHEBI:16236', 'Chemical') :: ['alcohol'] @entity377 :: ('CHEBI:36976', 'Chemical') :: ['nucleotide'] @entity13338 :: ('CHEBI:26565', 'Chemical') :: ['ribosyl'] @entity19396 :: ('MESH:D036563', 'Chemical') :: ['cADP-ribose'] @entity45299 :: ('1738', 'Gene') :: ['diaphorase'] @entity141 :: ('CHEBI:29108', 'Chemical') :: ['Ca(2+)'] @entity857 :: ('MESH:D000244', 'Chemical') :: ['ADP'] @entity4571 :: ('MESH:D009536', 'Chemical') :: ['nicotinamide']
@entity19396 :: (MESH:D036563,Chemical) :: ['cADP-ribose']
