5'-Adp → 5'-Adp

Participants

Reaction Steps

ADP substrate (>99.999% pure with respect to ATP), magnesium acetate and extractant reagent (0.15% cationic detergent and 0.25% tertiary diamine per final volume) is supplied from source (10) in sufficient quantity to provide 0.1 mM ADP and 10 mM magnesium ions in the reaction vessel, the magnet deenergised and stirrer operated and extraction/ADP conversion allowed to proceed for a set period between of 1 or 5 minutes before the stirrer is stopped, the magnet energised and the liquid removed to the luminometer flow cell (11) using the peristaltic tubing (3) and reversible pumps (4) and (12) while simultaneously mixing it with an effective volume of luciferase/luciferin reagent of depleted adenylate kinase content from source (13). Using an adenylate kinase modified Celsis LDR reagent of otherwise standard amounts of active ingredients this volume is about one half of the volume of the sample. Light emitted in the flow cell is related to the amount of E. coli captured by reference to standard curves obtained by performance of the assay with known numbers of cells.

Provenance & Evidence