A Homogeneous Bioluminescent Assay for High-Throughput Antimicrobial Screening...

Abstract

Frank Fan, Braeden Butler, Terry Riss and Keith Wood
Promega Corporation, 2800 Woods Hollow Road, Madison WI 53711

ATP-based detection of microbial cells represents a key application of luciferase/luciferin bioluminescence assay. Conventional methods require two steps: application of a lysis reagent to release microbial ATP, followed by a detection reagent to elicit bioluminescence. We have developed an assay that combines the lytic reagent with luciferase/luciferin, thus allowing sensitive
detection of microbial cells in a single-step. The assay system utilizes a thermostable luciferase to enable extraction of ATP from bacterial cells and to support a stable “glow-type” luminescent signal. In addition, we developed a formulation to achieve rapid and efficient extraction of ATP from a variety of microbial cells. The combination of these two essential elements enabled the design of a homogeneous single-reagent system (BacTiter-Glo™ Reagent) for performing ATP assays on cultured cells.